GC-QTOF MS Research Articles

Integrating 16 S rRNA gene sequencing and metabolomics analysis to reveal the mechanism of Angelica sinensis oil in alleviating ulcerative colitis in mice

Angelica sinensis (Oliv.) Diels (AS) is a commonly used herbal medicine and culinary spice known for its gastrointestinal protective properties. Angelica sinensis oil (AO) is the main bioactive component of AS. However, the therapeutic effects and mechanisms of AO on the gastrointestinal tract remain unclear. In this study, we aim to investigated the potential of AO in restoring gut microbiota disorder and metabolic disruptions associated with ulcerative colitis (UC). A systematic chemical characterization of AO was conducted using GC×GC-Q TOF-MS. A UC mouse model was established by freely drinking DSS to assess the efficacy of AO. Utilizing 16 S rRNA sequencing in combination with untargeted metabolomics analysis of serum, we identified alterations in gut microbiota, differential metabolites, and pathways influenced by AO in UC treatment, thereby elucidating the therapeutic mechanism of AO in UC management. Pharmacodynamic results indicated that AO effectively inhibited the content of inflammation mediators, such as Interleukin-1β, Interleukin-6 and tumor necrosis factor-α, and proserved colon tissue integrity in UC mice. Furthermore, AO significantly downregulated the abundance of pathogenic bacteria (Bacteroidetes, Proteobacteria, and Desulfobacteriaceae) while increasing the abundance of beneficial bacteria (Firmicutes, Blautia, Akkermansia, and Lachnospiraceae). Metabolomics analysis highlighted significant disruptions in endogenous metabolism in UC mice, with a notable restoration of SphK1 and S1P levels following AO administration. Besides, we discovered that AO regulated the balance of sphingolipid metabolism and protected the intestinal barrier, potentially through the SphK1/MAPK signaling pathway. Overall, this study indicated that AO effectively ameliorates the clinical manifestations of UC by synergistically regulating gut microbe and metabolite homeostasis. AO emerges as a potential functional and therapeutic ingredient for UC treatment.

The volatile oil of Hyssopus cuspidatus Boriss. (HVO) ameliorates OVA-induced allergic asthma via inhibiting PI3K/Akt/JNK/P38 signaling pathway and maintaining airway barrier integrity

Ethnopharmacological relevanceHyssopus cuspidatus Boriss., a classic Uyghur medicine, is used to treat inflammatory lung diseases such as asthma. But the therapeutic effect and mechanism of the volatile oil of Hyssopus cuspidatus Boriss.(HVO) in asthma therapy remain unclear. Aim of the studyWe aim to characterize the constituents of HVO, investigate the therapeutic effect in OVA-induced allergic asthmatic mice and further explore the molecular mechanism. Materials and methodsIn this study, we applied two-dimensional gas chromatography quadrupole time-of-flight mass spectrometry (GC × GC-QTOF MS) to identify the ingredients of HVO. We established OVA-induced asthmatic model to investigate the therapeutic effect of HVO. To further explore the potential molecular pathways, we used network pharmacology approach to perform GO and KEGG pathways enrichment, and then built an ingredient-target-pathway network to identify key molecular pathways. Finally, LPS-induced RAW 264.7 macrophages and OVA-induced asthmatic model were used to validate the potential signaling pathways. ResultsGC × GC-QTOF MS analysis revealed the presence of 123 compounds of HVO. The sesquiterpenes and monoterpenes are the main constituents. The in vivo study indicated that HVO suppressed OVA-induced eosinophilic infiltration in lung tissues, inhibited the elevation of IgE, IL-4, IL-5, and IL-13 levels, downregulated the expressions of phosphorylated PI3K, Akt, JNK and P38, and maintained epithelial barrier integrity via reducing the degradation of occludin, Zo-1, Zo-2, and E-cadherin. The in vitro study also revealed an inhibition of NO release and downregulation of phosphorylated PI3K, Akt, JNK and P38 levels. ConclusionHVO alleviates airway inflammation in OVA-induced asthmatic mice by inhibiting PI3K/Akt/JNK/P38 signaling pathway and maintaining airway barrier integrity via reducing the degradation of occludin, Zo-1, Zo-2, and E-cadherin.

Sandalwood Oils of Different Origins Are Active In Vitro against Madurella mycetomatis, the Major Fungal Pathogen Responsible for Eumycetoma.

In the search for new bioactive agents against the infectious pathogen responsible for the neglected tropical disease (NTD) mycetoma, we tested a collection of 27 essential oils (EOs) in vitro against Madurella mycetomatis, the primary pathogen responsible for the fungal form of mycetoma, termed eumycetoma. Among this series, the EO of Santalum album (Santalaceae), i.e., East Indian sandalwood oil, stood out prominently with the most potent inhibition in vitro. We, therefore, directed our research toward 15 EOs of Santalum species of different geographical origins, along with two samples of EOs from other plant species often commercialized as "sandalwood oils". Most of these EOs displayed similar strong activity against M. mycetomatis in vitro. All tested oils were thoroughly analyzed by GC-QTOF MS and most of their constituents were identified. Separation of the sandalwood oil into the fractions of sesquiterpene hydrocarbons and alcohols showed that its activity is associated with the sesquiterpene alcohols. The major constituents, the sesquiterpene alcohols (Z)-α- and (Z)-β-santalol were isolated from the S. album oil by column chromatography on AgNO3-coated silica. They were tested as isolated compounds against the fungus, and (Z)-α-santalol was about two times more active than the β-isomer.

Comparative study on different extraction methods of oil extract from Mentha asiatica Boriss. and optimization of the extraction process by response surface methodology

AbstractIn the current study, the lipophilic solvent (n‐hexane) extraction, steam distillation (SD) and supercritical CO2 fluid extraction (SC–CO2) methods were used to extract the volatile components of Mentha asiatica Boriss. Comparisons were made between the extraction yield, chemical component and biological activity. Gas chromatography quadrupole‐time‐of‐flight mass spectrometry (GC–QTOF–MS) and semi‐quantitative analysis of chemical contents by flame ionization detector (GC–FID) were used to study the chemical ingredients in Mentha asiatica Boriss. aromatic extract. The oil extract obtained from lipophilic solvent (n‐hexane) extraction (1.27 ± 0.03%, w/W) showed the highest yield, followed by supercritical CO2 fluid extraction (1.15 ± 0.04%, w/W), the extraction rate of SD was (0.37 ± 0.01%，w/W). The oil extract of Mentha asiatica Boriss. contained a total of 70 components that were identified. In the following, the primary volatile compounds of the essential oil produced using the three extraction techniques: α‐thujene, camphene, sabinene, β‐pinene, β‐cymene, limonene, β‐terpinene, camphol, α‐terpinene, carvenone, carvone, piperitone oxide, thymol, myrtenyl acetate, dihydrojasmone, caryophyllene, germacrene D and caryophyllene oxide. Findings from this study indicate that the three methods of essential oil extraction produced oils with varying antioxidant capacities against DPPH and ABTS free radicals. Against Staphylococcus aureus, Candida albicans, Bacillus subtilis and Escherichia coli, all of the essential oils simultaneously showed clear inhibitory actions. Response surface methodology (RSM) and the Box–Benhnken design with three factors and three levels were both utilized to optimize the distillation process of essential oils. The appropriate extraction parameters were a 3 h extraction period, a solid–liquid ratio of 1:10 and a 3 h soaking time. The highest yield of essential oil was 0.38%. The impact of extraction parameters on essential oil yield was examined using the SD method. The findings indicated that the main factors affecting the yield of volatile oil extracted by SD were soaking time > ratio of solid to liquid > extraction time.

Gas chromatography mass spectrometry couple with quadrupole time-of-flight (GC-QTOF MS) as a powerful tool for profiling of oxygenated sesquiterpenes in agarwood oil

Agarwood (Aquilaria malaccensis) is very well known as the most expensive wood in the world due to its wide applications in perfumery, cosmetic traditional medicine, and religious ceremonies. The study aimed to give an in-depth characterisation focusing on marker compounds in A. malaccensis from different places in Malaysia. The establishment of an oxygenated sesquiterpenes chemical profile of the fungus-infected agarwood oil was achieved by gas chromatography mass spectrometry (GC–MS) coupled with quadrupole time (QTOF) technique. Aroma compounds were identified as sesquiterpenes and oxygenated sesquiterpenes where agarospirol was found in samples of all locations (3.12%, 3.54%, 3.36% and 2.26% from Melaka, Pahang, Kelantan A and Kelantan B respectively) and also N-hexadecanoic acid as one of the major compounds. Both compounds were further isolated by Prep-GC and confirmed by NMR. This study provides a reference for agarwood oil analysis from different origins in Malaysia.

Person-portable gas chromatography-toroidal ion trap mass spectrometry analysis of coffee bean volatile organic compounds

Analysis of roasted coffee bean volatile organic compounds (VOCs) was performed using person-portable gas chromatography–toroidal ion trap mass spectrometry (ppGC–TMS). Chromatographic separation performance and mass spectra were compared and contrasted with benchtop gas chromatography–quadrupole time-of-flight mass spectrometry (GC–QTOFMS) and gas chromatography–quadrupole mass spectrometry (GC–QMS). The tentative identification of about 50 analytes was established in benchtop analysis, based on accurate MS data, MS spectrum matching and retention indices, and contrasted with 20 individual analyte peaks similarly recognised in ppGC–TMS analysis. This study suggests the suitability of ppGC–TMS analysis for rapid coffee VOC profiling with minimal sample preparation, using selective key chemical markers, with potential use for on-site process monitoring and roasting optimisation.

Profiling of volatile substances by direct thermal desorption gas chromatography high-resolution mass spectrometry for flagging a characterising flavour in cigarette tobacco

This paper describes an analytical method that supports the implementation of articles 9 and 10 of the WHO Framework Convention on Tobacco Control (FCTC) regarding the provisions on the reduction of the palatability and attractiveness of tobacco products regarding flavour ingredients. This study aimed to develop a screening method to identify cigarettes that may have a characterising flavour to support the implementation of the ban of characterising flavours of tobacco products, as laid down in the US and EU law. An analytical method combining direct thermal desorption and GC–QTOF MS was developed for acquiring the profile of volatile and semi-volatile substances in tobacco. A database of flavour additives was created comprising 133 compounds. A group of cigarettes without a declared characterising flavour was used to establish a reference profile of flavouring chemicals commonly present in tobacco products. A reference profile was modelled both by the means of principal component analysis (PCA) and based on the calculation of threshold values specified as 95th percentile of measured compounds’ relative responses. Cigarettes and roll-your-own tobacco labelled as flavoured were analysed to evaluate the discrimination power of the method. A constructed model of the reference cigarettes allowed the differentiation of the flavoured tobacco products from the reference group. The method allows drawing conclusions on the chemical profiles of flavour constituents of tobacco products at even sensorial subliminal concentration levels and is suitable for both the initial screening of products on the market for characterising flavours and for confirmatory purposes after sensory analysis.Graphical abstract

Comparing Stable Isotope Enrichment by Gas Chromatography with Time-of-Flight, Quadrupole Time-of-Flight, and Quadrupole Mass Spectrometry.

Stable isotope tracers are applied for in vivo and in vitro studies to reveal the activity of enzymes and intracellular metabolic pathways. Most often, such tracers are used with gas chromatography coupled to mass spectrometry (GC-MS) owing to its ease of operation and reproducible mass spectral databases. Differences in isotope tracer performance of the classic GC-quadrupole MS instrument and newer time-of-flight instruments are not well studied. Here, we used three commercially available instruments for the analysis of identical samples from a stable isotope labeling study that used [U-13C6] d-glucose to investigate the metabolism of the bacterium Rothia mucilaginosa with respect to 29 amino acids and hydroxyl acids involved in primary metabolism. The prokaryote R. mucilaginosa belongs to the family of Micrococcaceae and is present and metabolically active in the airways and sputum of cystic fibrosis patients. Overall, all three GC-MS instruments (low-resolution GC-SQ MS, low-resolution GC-TOF MS, and high-resolution GC-QTOF MS) can be used to perform stable isotope tracing studies for glycolytic intermediates, tricarboxylic acid (TCA) metabolites, and amino acids, yielding similar biological results, with high-resolution GC-QTOF MS offering additional capabilities to identify the chemical structures of unknown compounds that might show significant isotope enrichments in biological studies.

Identification of new, very long-chain polyunsaturated fatty acids in fish by gas chromatography coupled to quadrupole/time-of-flight mass spectrometry with atmospheric pressure chemical ionization.

The characterization of very long-chain (>C24) polyunsaturated fatty acids (VLC-PUFAs), which are essential in the vision, neural function, and reproduction of vertebrates, is challenging because of the lack of reference standards and their very low concentrations in certain lipid classes. In this research, we have developed a new methodology for VLC-PUFA identification based on gas chromatography coupled to quadrupole/time-of-flight mass spectrometry with an atmospheric pressure chemical ionization source (GC-APCI-QTOF MS). The mass accuracy attainable with the innovative QTOF instrument, together with the soft ionization of the APCI source, provides valuable information on the intact molecule, traditionally lost with electron ionization sources due to the extensive fragmentation suffered. We have identified, for the first time, VLC-PUFAs with chains up to 44 carbons in eyes, brain, and gonads of gilthead sea bream, a commercially important fish in the Mediterranean. The added value of ion mobility-mass spectrometry (IMS), recently developed in combination with GC-QTOF MS, and the contribution of the collisional cross section (CCS) parameter in the characterization of novel VLC-PUFAs (for which reference standards are not available) have been also evaluated. The methodology developed has allowed assessing qualitative differences between farmed and wild fish, and opens new perspectives in a still scarcely known field of research.

Unraveling salt responsive metabolites and metabolic pathways using non-targeted metabolomics approach and elucidation of salt tolerance mechanisms in the xero-halophyte Haloxylon salicornicum

Haloxylon salicornicum is a xero-halophyte growing in saline and arid regions of the world. Metabolite profiling was carried out in shoot of both control and salinity treated (400 mM NaCl) samples by GC-QTOF–MS and HPLC-DAD analysis to decipher the salinity tolerance mechanism in this xero-halophyte. The present study investigates the alteration in metabolite profile of H. salicornicum that support the salinity tolerance of the plant. The metabolomic analysis of H. salicornicum shoot identified 56 metabolites, of which 47 metabolites were significantly changed in response to salinity. These metabolites were mainly included in the category of amino acids, organic acids, amines, sugar alcohols, sugars, fatty acids, alkaloids, and phytohormones. In response to salinity, most of the amino acids were down-regulated except alanine, phenylalanine, lysine, and tyramine, which were up-regulated in H. salicornicum. In contrast to amino acids, most sugars and organic acids were up-regulated in response to salinity. Correlation and pathway enrichment analysis identified important biological pathways playing significant roles in conferring salt tolerance of H. salicornicum. These biological pathways include amino sugar and nucleotide sugar metabolism, citrate cycle (TCA cycle), starch and sucrose metabolism, phenylalanine metabolism, cysteine, methionine, glycine, serine, and threonine metabolism, etc. The data presented here suggest that the modulations of various metabolic pathways facilitate H. salicornicum to survive and grow optimally even under high salinity condition. This study offers comprehensive information on metabolic adaptations and overall salt tolerance mechanisms in H. salicornicum. The information gained through this study will provide guidance to plant breeders and molecular biologists to develop salinity tolerant crop varieties.

Targeting the administration of ecdysterone in doping control samples

PurposeThe phytosteroid ecdysterone was recently reported to enhance performance in sports and may thus be considered as a substance of relevance in anti-doping control. To trace back an administration of ecdysterone from urine samples, analytical properties have been investigated to assess its integration into initial testing procedures (ITP) in doping control laboratories.MethodsAnalytical properties of ecdysterone were evaluated using gas chromatography-quadrupole/time-of-flight-mass spectrometry (GC–QTOF–MS) and liquid chromatography (LC)–QTOF–MS. Its metabolism and elimination in human were studied using urines collected after administration.ResultsThe detectability of ecdysterone by GC–MS (after derivatization) and/or LC–MS(/MS) has been demonstrated and sample preparation methods were evaluated. Dilute-and-inject for LC–MS(/MS) or solid phase extraction using Oasis HLB for GC–MS or LC–MS were found most suitable, while liquid–liquid extraction was hampered by the high polarity of ecdysteroids.Most abundantly, ecdysterone was detected in the post administration urines as parent compound besides the metabolite desoxy-ecdysterone. Additionally, desoxy-poststerone was tentatively assigned as minor metabolite, however, further investigations are needed.ConclusionAn administration of ecdysterone can be targeted using existing procedures of anti-doping laboratories. Ecdysterone and desoxy-ecdysterone appeared as suitable candidates for integration in ITP. Using dilute-and-inject a detection of the parent compound was possible for more than 2 days after the administration of a single dose of ~ 50 mg.

Coupled multidimensional GC and odor activity value calculation to identify off-odors in thermally processed muskmelon juice

Predominant contributors to thermally-induced off-notes in muskmelon juice were investigated by combination of sensory evaluation, GC–olfactometry (GC–O), multidimensional GC, odor activity value (OAV) calculation and addition experiments. “Fermented” and “sulfurous” were revealed as the dominant heat-induced offensive off-notes, and fundamental heat-led changes in odorant compositions were observed. Comparative aroma extract dilution analysis revealed that the off-flavors were mainly caused by newly heat-formed compounds with extremely low odor threshold and unpleasant smells. These included volatile sulfur compounds with “rotten” odors, Strecker aldehydes having “fermented” notes, and heterocyclics exhibiting “roasted” smells. Specifically, dimethyl sulfide, methional, methanethiol, dimethyl trisulfide, dimethyl disulfide, and acetaldehyde were identified as the predominant off-note contributors, among which DMS was the most important one, showing the highest OAV. Multiple extraction techniques, GC–O coupling to GC × GC–QTOF–MS and OAV calculation are powerful tools for accurate location and reliable identification of trace and key off-odorants in natural complex matrices.

Suspect screening analysis of the occurrence and removal of micropollutants by GC-QTOF MS during wastewater treatment processes.

The presence of micropollutants in wastewater treatment plant (WWTP) poses potential risks to the aquatic system and human health. This study comprehensively characterized the presence, elimination, and transformation of micropollutants in the WWTP using the suspect screening approaches by employing the gas chromatography coupled with quadrupole time-of-flight mass spectrometry. Overall, 5724 features were identified in influent, while only 3418 features existed in effluents. Hierarchical cluster analysis (HCA) was used to group features based on intensity profiles to investigate the behavior and fate of detected features. Four representative trends were defined with the corresponding features. Among 117 compounds identified during the treatment process, there were 99 compounds in influent, 10 biological transformation products (TPs), 3 ozone TPs, and 5 ultraviolet TPs. Furthermore, the removal efficiencies for the detected compounds were determined. The study findings highlight the importance of the removal of emerging pollutants and TPs such as fipronil, fipronil sulfide, and fipronil sulfone.

Comprehensive investigation of pesticides in Brazilian surface water by high resolution mass spectrometry screening and gas chromatography–mass spectrometry quantitative analysis

In this work, a comprehensive investigation on the occurrence of pesticides in the Paraná 3 hydrographic basin of Paraná State, Brazil, was made by application of wide-scope screening based on ultra-high performance liquid chromatography (LC) and gas chromatography (GC) both coupled to quadrupole time-of-flight mass spectrometry (QTOF MS). The use of two complementary techniques, such as GC-QTOF MS and LC-QTOF MS, allowed screening a large number of compounds with different polarity and volatility. This screening approach was applied to 17 samples, enabling the detection of fifty-two pesticides and six metabolites. In a second step, an specific research was made on the herbicide atrazine, one of the most frequent compounds in samples, and its major transformation products (TPs), which were quantitatively analyzed by dispersive liquid-liquid microextraction (DLLME) followed by GC–MS measurement. Twenty-one agricultural streams from the Paraná 3 hydrographic basin were sampled twice in 2017, each time along six successive weeks. Additional samples were also collected after rain events exceeding 10 mm. In total, 407 samples were quantitatively analyzed by DLLME/GC–MS. Atrazine concentrations did not exceed the maximum permitted concentration of 2 μg L−1 according to Brazilian legislation, and only one surface water sample, collected after precipitation events, was slightly above this value (2.89 μg L−1). The maximum concentrations for the TPs desethylatrazine and deisopropylatrazine were 0.80 and 1.22 μg L−1, respectively. Based on the quantification results, a map was produced showing the occurrence of atrazine and its TPs in the area under study. This is the first time that the presence of agrochemicals is evaluated in the Paraná 3 hydrographic basin.

GC-QTOFMS with a low-energy electron ionization source for advancing isotopologue analysis in 13C-based metabolic flux analysis.

For the study of different levels of (intra)cellular regulation and condition-dependent insight into metabolic activities, fluxomics experiments based on stable isotope tracer experiments using 13C have become a well-established approach. The experimentally obtained non-naturally distributed 13C labeling patterns of metabolite pools can be measured by mass spectrometric detection with front-end separation and can be consequently incorporated into biochemical network models. Here, despite a tedious derivatization step, gas chromatographic separation of polar metabolites is favorable because of the wide coverage range and high isomer separation efficiency. However, the typically employedelectron ionization energy of 70 eV leads to significant fragmentation and consequently only low-abundant ions with an intact carbon backbone. Since these ions are considered a prerequisite for the analysis of the non-naturally distributed labeling patterns and further integration into modeling strategies, a softer ionization technique is needed. In the present work, a novel lowenergy electron ionization source is optimized for the analysis of primary metabolites and compared with a chemical ionization approach in terms of trueness, precision, and sensitivity.

Group-Type Analysis of Hydrocarbons and Sulfur Compounds in Thermally Stressed Merox Jet Fuel Samples

Methods to investigate the response of hydrocarbons and sulfur compounds to different extents resulting from thermal oxidation of jet fuels refined via the mercaptan oxidation (Merox) process have been developed. Relative comparison of hydrocarbon contents including n-paraffins, isoparaffins, olefins, naphthenes, and aromatics (PIONA) was performed by using comprehensive two-dimensional gas chromatography hyphenated with a quadrupole-accurate mass time-of-flight mass spectrometer (GC × GC–Q-TOFMS). A simple approach for quantification of sulfur compound groups was then developed using both GC × GC and GC hyphenated with flame photometric detection (GC × GC–FPD; GC–FPD). The approach largely separated the sulfur compounds into two distinct regions comprising of lower polarity S1 (thiophenes, thiophenols, thiols, disulfides, sulfides, and alkyl thianaphthenes) and higher polarity S2 (sulfones and phenylsulfides) groups. The reliability of this approach was evaluated using higher resolution GC × GC–FPD, show...

In vitro antioxidant and, \u03b1-glucosidase inhibitory activities and comprehensive metabolite profiling of methanol extract and its fractions from Clinacanthus nutans

BackgroundThis study was aimed to evaluate antioxidant and α-glucosidase inhibitory activity, with a subsequent analysis of total phenolic and total flavonoid content of methanol extract and its derived fractions from Clinacanthus nutans accompanied by comprehensive phytochemical profiling.MethodsLiquid-liquid partition chromatography was used to separate methanolic extract to get hexane, ethyl acetate, butanol and residual aqueous fractions. The total antioxidant activity was determined by 2,2-diphenyl-1-picrylhydrazy (DPPH) radical scavenging and ferric reducing antioxidant power assay (FRAP). The antidiabetic activity of methanol extract and its consequent fractions were examined by α-glucosidase inhibitory bioassay. The chemical profiling was carried out by gas chromatography coupled with quadrupole time-of-flight mass spectrometry (GC Q-TOF MS).ResultsThe total yield for methanol extraction was (12.63 ± 0.98) % (w/w) and highest fractionated value found for residual aqueous (52.25 ± 1.01) % (w/w) as compared to the other fractions. Significant DPPH free radical scavenging activity was found for methanolic extract (63.07 ± 0.11) % and (79.98 ± 0.31) % for ethyl acetate fraction among all the fractions evaluated. Methanol extract was the most prominent in case of FRAP (141.89 ± 0.87 μg AAE/g) whereas most effective reducing power observed in ethyl acetate fraction (133.6 ± 0.2987 μg AAE/g). The results also indicated a substantial α-glucosidase inhibitory activity for butanol fraction (72.16 ± 1.0) % and ethyl acetate fraction (70.76 ± 0.49) %. The statistical analysis revealed that total phenolic and total flavonoid content of the samples had the significant (p < 0.05) impact on DPPH free radical scavenging and α-glucosidase inhibitory activity.ConclusionCurrent results proposed the therapeutic potential of Clinacanthus nutans, especially ethyl acetate and butanol fraction as chemotherapeutic agent against oxidative related cellular damages and control the postprandial hyperglycemia. The phytochemical investigation showed the existence of active constituents in Clinacanthus nutans extract and fractions.

Identification of bioactive compounds with GC–Q-TOF–MS in the extracts from Clinacanthus nutans using subcritical carbon dioxide extraction

Subcritical carbon dioxide Soxhlet extraction of biologically active compounds from Clincanthus nutans was investigated by full factorial design to identify and optimize the factors (particle size and co-solvent) affecting extract yield, antioxidant activity, total phenolic content, total flavonoid content, and α-glucosidase inhibitory activity. An average of 3.103% yield, 98.90% antioxidant activity, 49.40 mg/g (GAE) TPC, 43.76 mg/g (RE), and 88.58% AGI activity can be achieved using the optimum levels of independent variables. The GC-Q-TOF MS identification of optimized extract shown that different classes of phytoconstituents were successfully separated by CO2-Soxhlet to produce potential antioxidant and α-glucosidase inhibitory activity.

Avoiding hard chromatographic segmentation: A moving window approach for the automated resolution of gas chromatography–mass spectrometry-based metabolomics signals by multivariate methods

Gas chromatography–mass spectrometry (GC–MS) produces large and complex datasets characterized by co-eluted compounds and at trace levels, and with a distinct compound ion-redundancy as a result of the high fragmentation by the electron impact ionization. Compounds in GC–MS can be resolved by taking advantage of the multivariate nature of GC–MS data by applying multivariate resolution methods. However, multivariate methods have to be applied in small regions of the chromatogram, and therefore chromatograms are segmented prior to the application of the algorithms. The automation of this segmentation process is a challenging task as it implies separating between informative data and noise from the chromatogram. This study demonstrates the capabilities of independent component analysis–orthogonal signal deconvolution (ICA–OSD) and multivariate curve resolution–alternating least squares (MCR–ALS) with an overlapping moving window implementation to avoid the typical hard chromatographic segmentation. Also, after being resolved, compounds are aligned across samples by an automated alignment algorithm. We evaluated the proposed methods through a quantitative analysis of GC–qTOF MS data from 25 serum samples. The quantitative performance of both moving window ICA–OSD and MCR–ALS-based implementations was compared with the quantification of 33 compounds by the XCMS package. Results shown that most of the R2 coefficients of determination exhibited a high correlation (R2>0.90) in both ICA–OSD and MCR–ALS moving window-based approaches.

Mass spectral fragmentation of trimethylsilylated small molecules.

Mass spectrometry-based untargeted metabolomics detects many peaks that cannot be identified. While advances have been made for automatic structure annotations in LC-electrospray-MS/MS, no open source solutions are available for hard electron ionization used in GC-MS. In metabolomics, most compounds bear moieties with acidic protons, for example, amino, hydroxyl, or carboxyl groups. Such functional groups increase the boiling points of metabolites too much for use in GC-MS. Hence, in GC-MS-focused metabolomics, derivatization of these groups is essential and has been employed since the 1960s. Specifically, trimethylsilylation is known as mild and universal method for GC-MS analysis. Here, we comprehensively compile accurate mass fragmentation rules and pathways of trimethylsilylated small molecules from 80 research articles over the past 5 decades, including diagnostic fragment ions, neutral losses, and typical ion ratios, for alcohols, carboxylic acids, amines, amino acids, sugars, steroids, thiols, and phosphates. These fragmentation rules were subsequently validated by specificity and sensitivity assessments using the NIST 14 nominal mass library and a new in-house GC-QTOF MS library containing 589 accurate mass spectra. From 556 tested fragmentation patterns, 228 rules yielded true positive hits within 4 mDa mass accuracy. These rules can be applied to assign substructures for mass spectra computation and unknown identification. © 2016 Wiley Periodicals, Inc. Mass Spec Rev 37:245-257, 2018.