GC-MS Profiling Research Articles

Gas chromatographic–mass spectrometric profile of non-polar fraction and high-performance thin-layer chromatographic analysis of methanolic fraction with simultaneous quantifications of protocatechuic acid and quercetin in Carissa carandas L. fruits

Extraction of crude drugs by using different solvents provides polarity-based fractions containing specific type of secondary metabolites. Carissa carandas L. fruits were extracted and fractionated, petroleum ether extract was processed by a fatty acid methyl ester (FAME) technique for characterization by gas chromatography—mass spectrometry (GC—MS) analysis, and the remaining part was extracted with methanol for high-performance thin-layer chromatography (HPTLC) analysis, followed by simultaneous quantitative determination of protocatechuic acid and quercetin in methanolic fractions. A validated method for the simultaneous quantification of protocatechuic acid and quercetin was developed and is being reported for the first time in C. carandas L. fruits to the best of our knowledge. Petroleum ether and methanol fractions were found to be the best for the highest possible recovery of targeted analytes. Chromatographic elution of FAME compounds generated from petroleum ether extract was evaluated by GC—MS profiling. Nineteen fatty acid compounds were separated with the highest quantity of octacosanal (13.31%). On the other hand, a polar fraction was processed by HPTLC profiling. For achieving good separation, a mobile phase of toluene-ethyl acetate-formic acid (6:3:1; V/V) was used. Densitometric determination was carried out at 310 nm in the reflection/absorption mode. The calibration curves were linear in the range of 100–600 ng per spot for protocatechuic acid and quercetin. During the analysis, the dried raw material from C. carandas L. fruits showed the presence of protocatechuic acid (0.04%) and quercetin (0.05%). The proposed method is simple, precise, specific, and accurate. The statistical analysis of the data obtained proves that the method is reproducible and selective, which can be used for the routine analysis of the reported phenolic compounds in crude drug and extracts. The simultaneous quantification of these phenolic compounds has not yet been reported in C. carandas L. fruits which may be utilized for the proper standardization of the drug.

Metabolic perturbations caused by depletion of nephronophthisis factor Anks6 in mIMCD3 cells.

Nephronophthisis (NPH) is an inherited form of cystic kidney disease with various extrarenal manifestations accounting for the largest amount of endstage renal disease in childhood. Patient mutations of Anks6 have also been found to cause NPH like phenotypes in animal models. However, little is known about functionality of Anks6. We investigated the impact of Anks6 depletion on cellular metabolism of inner medullary collecting duct cells by GC-MS profiling and targeted LC-MS/MS analysis using two different shRNA cell lines for tetracycline-inducible Anks6 downregulation, namely mIMCD3 krab shANKS6 i52 and mIMCD3 krab shANKS6 i12. In combination, we could successfully identify 158 metabolites of which 20 compounds showed similar alterations in both knockdown systems. Especially, large neutral amino acids, such as phenylalanine, where found to be significantly downregulated indicating disturbances in amino acid metabolism. Arginine, lysine and spermidine, which play an important role in cell survival and proliferation, were found to be downregulated. Accordingly, cell growth was diminished in tet treated mIMCD3 krab shANKS6 i52 knockdown cells. Deoxynucleosides were significantly downregulated in both knockdown systems. Hence, PARP1 levels were increased in tet treated mIMCD3 krab shANKS6 i52 cells, but not in tet treated mIMCD3 krab shANKS6 i12 cells. However, yH2AX was found to be increased in the latter. In combination, we hypothesise that Anks6 affects DNA damage responses and proliferation and plays a crucial role in physiological amino acid and purine/pyrimidine metabolism.

Deciphering the phylogeny of violets based on multiplexed genetic and metabolomic approaches

Molecular phylogenetics based on nucleotide sequence comparisons has profoundly influenced plant taxonomy. A comprehensive chemotaxonomical approach based on GC-MS and UHPLC-HRMS profiling was evaluated for its ability to characterize a large collection of plants all in the violet family Violaceae (n = 111) and thus decipher the taxonomy. A thorough identification of violets is challenging due to their natural hybridization and phenotypic variability. Phylogenetic inference performed on ribosomal internal transcribed spacer sequences using maximum likelihood and neighbor-joining distance methods allowed the clear identification of 58% of the collection. Metabolomic approaches with multivariate data analysis were performed on SPME/GC-MS chromatograms of volatile compounds emitted by fresh mature flowers and on UHPLC-HRMS/MS leaf extracts for non-volatile compounds. Interestingly, molecular and biochemical approaches provided separate classifications while highlighting several common clusters. The profiling of secondary metabolites was proved most suitable for the classification of hundreds of extracts. The combination of phylogenetic and chemotaxonomic approaches, allowed the classification of 96% of the entire collection. A correlation network revealed specific chemotaxonomic biomarkers, in particular flavonoids, coumarins and cyclotides. Overall, our pioneering approach could be useful to solve misclassification issues within collections of close plant species.

BBX31 promotes hypocotyl growth, primary root elongation and UV-B tolerance in Arabidopsis

ABSTRACTPhotomorphogenesis is an important developmental process that helps the seedlings adapt to external light conditions. B-Box proteins are a family of transcription factors that regulate photomorphogenic responses. BBX31 negatively regulates photomorphogenesis under visible light. In contrast, it promotes photomorphogenesis under UV-B and enhances tolerance to high doses of UV-B radiation. BBX31 and HY5 independently and oppositely regulate the ability of seedlings to adapt to varying light intensities. BBX31 also regulates primary root elongation under low intensities of white light. GC-MS and HPLC-based metabolite profiling identified differential accumulation of multiple primary and secondary metabolites in 35S:BBX31 that might enhance tolerance to UV-B.

LC-ESI/LTQOrbitrap/MS/MS and GC–MS profiling of Stachys parviflora L. and evaluation of its biological activities

The use of some Stachys genus as herbal remedies is known and the aerial parts have a pharmaceutical interest, being used in Anatolia and Iran as wild tea. In this study, chemical composition, antimicrobial, antioxidant, and antiproliferative activities of the methanolic extract and essential oil (EO) of Stachys parviflora L. (S. parviflora) were evaluated. Qualitative analysis of metabolites of S. parviflora methanolic extract was studied using liquid chromatography coupled to high resolution mass spectrometry (LC-ESI/LTQOrbitrap/MS), evidencing the presence of phenolic acids and flavonoids derivatives. The EO was analyzed using gas chromatography coupled to mass spectrometry (GC/MS). Eighty-seven compounds were characterized in the EO of S. parviflora, of which α-terpenyl acetate (23.6%), β-caryophyllene (16.8%), bicyclogermacrene (9.3%), spathulenol (4.9%) and α-pinene (4.2%) were found to be the major components. The highest antimicrobial effect of EO was found to S. aureus and B. cereus (MIC = 0.01 μg/ml), while the highest activity of extract was against B. cereus (MIC = 125 μg/ml). The methanolic extract exhibited strong antioxidant activity in DPPH (IC50 = 76.87 μg/ml) and β-carotene/linoleic acid assay (BCB, IC50 = 188.47 μg/ml) methods. Furthermore, in vitro cytotoxicity evaluation against three cell lines namely human ovarian carcinoma (A2780), human colon carcinoma (HCT), and mouse melanoma cell line (B16F10), showed an anti-proliferative activity of the EO ranging from IC50 value 30.95 μg/ml to 16.55 μg/ml. The results from this study have demonstrated the promising cytotoxic, antibacterial, and antifungal properties of S. parviflora, which could have wide potential applications in food and pharmaceutical industries.

Alpha Amylase, Alpha Glucosidase Inhibition and Profiling of Volatile Compounds of Biologically Active Extracts from Momordica cymbalaria (Hook, Fenzl) Skin and Seeds

Momordica cymbalaria (Hook, Fenzl) has been used in traditional medicine for diabetes in India. This study was designed to examine, total phenolic, total flavonoid contents and in vitro anti-oxidant activity by using seven different types of solvent extracts. The biologically active extracts, ethanol extract of fruit skin (EESK) and methanol extract of seed (MESE) were evaluated for in vitro anti-diabetic activity followed by detection of bioactive metabolites by GC-MS analysis. EESK and MESE exhibited IC50 values of 185.1 μg/ml against α – amylase and 224.4 μg/ml against α – glucosidase, respectively. GC-MS profiling of skin and seed portion of the vegetable revealed the presence of 40 compounds from EESK and 29 compounds from MESE. Thus, this study reveals that EESK and MESE exhibited in vitro anti-diabetic activity and GC-MC profiling confirms the presence of anti-diabetic phytocompounds such as L-Cysteine and cis- Vaccenic acid.

Cichorium intybus from India: GC-MS Profiling, Phenolic Content and in vitro Antioxidant Capacity of Sequential Soxhlet Extracted Roasted Roots

Abstract Research on the bio-activities and chemical composition of roasted C. intybus roots from India is very little. In present studies GC-MS analysis of volatile components of roasted C. intybus roots, phenolics and flavonoid content estimation and antioxidant potential of roasted C. intybus roots was carried out. Antioxidant potential was also evaluated using FRAP, DPPH, hydroxyl radical, nitric oxide and superoxide free radical scavenging method. Extracts were prepared by sequential Soxhlet extraction. GC-MS analysis of volatile components of roasted C. intybus root extracts revealed that 5-hydroxymethyl furfural was major volatile component in dichloromethane and methanol extract whereas lupeol and its derivative compounds were major constituents of hexane extract. Quantitative estimation for total phenols and flavonoids showed that the methanol extract of C. intybus roots contained highest phenolic and flavonoid content as compared to other extracts and also showed strong radical scavenging activities which were comparable with ascorbic acid used as standard. All extracts showed IC50 values less than 0.6 mg/mL furthermore, extracts of roasted C. intybus showed the high total antioxidant potential for the reduction of Fe3+ to Fe2+. The C. intybus roots possess good antioxidant capacity even after roasting and all the extracts showed good activities.

Phytochemical profile, anti-oxidant, anti-inflammatory, and anti-proliferative activities of Pogostemon deccanensis essential oils.

Essential oils (EOs) obtained from aerial parts of Pogostemon deccanensis were analyzed for GC-MS profiling, and evaluated for antioxidant, anti-inflammatory, and anti-proliferative activities. GC-MS analysis revealed a total of 47 constituents, establishing the EOs rich in sesquiterpene with > 20 sesquiterpenes constituting around 77% of the total EO yield. Major constituents included Curzerene (Benzofuran, 6-ethenyl-4,5,6,7-tetrahydro-3,6-dimethyl-5-isopropenyl-, trans-) (26.39%) and epi-Cadinol (22.68%), Ethanone, 1-(2,4,6-trihydroxyphenyl) (6.83%, Acetophenones), and Boldenone (3.47%, anabolic steroid). EOs found to be rich in phytochemicals attributed for antioxidant potentials of aromatic/medicinal plants, viz., flavonoids (2.71µg quercetin equivalents g-1 EO), total phenols (3.94µg gallic acid equivalents (GAE) g-1 EO), carotenoids (14.3µg β-carotene equivalents g-1 EO), and ascorbic acid (2.21µg ascorbic acid equivalents g-1 EO). P. deccanensis EOs exhibited striking antioxidant activities assessed by wide range of assays including ferric reducing antioxidant potential (FRAP, 255.3 GAE at 2µg mL-1 EO), total antioxidant activity (TAA, 264.3 GAE at 2µg ml-1) of EO, DPPH (65% inhibition at 2µg mL-1), and OH (58% inhibition at 2µg mL-1) scavenging. Interestingly, EOs showed considerably higher anti-lipid peroxidation activity than the standard antioxidant molecule ascorbic acid, with 50% protection by 1.29µg mL-1 EO against 20.0µg mL-1 standard. EOs showed strong anti-inflammatory activity with 50% inhibition at 1.95µg mL-1 EO. The anti-proliferative activity of EOs was tested against mouse cancer cell line and the EOs proved a potent anti-proliferative agent with only 2.1% cell survival at 2µg mL-1 EO, whereas the EOs were largely non-toxic-to-normal (non-cancerous) cells with approximately 80% cell survival at the 2µg mL-1 EOs. This being the first attempt of phytochemical profiling and wide array of biological activities of P. deccanensis EOs holds significance as the striking activities were observed at very low concentrations, in some cases at lower than the commercial standards, and has, therefore, great potential for pharmaceutical or commercial exploration.

Cold Acclimation Favors Metabolic Stability in Drosophila suzukii.

The invasive fruit fly pest, Drosophila suzukii, is a chill susceptible species, yet it is capable of overwintering in rather cold climates, such as North America and North Europe, probably thanks to a high cold tolerance plasticity. Little is known about the mechanisms underlying cold tolerance acquisition in D. suzukii. In this study, we compared the effect of different forms of cold acclimation (at juvenile or at adult stage) on subsequent cold tolerance. Combining developmental and adult cold acclimation resulted in a particularly high expression of cold tolerance. As found in other species, we expected that cold-acclimated flies would accumulate cryoprotectants and would be able to maintain metabolic homeostasis following cold stress. We used quantitative target GC-MS profiling to explore metabolic changes in four different phenotypes: control, cold acclimated during development or at adult stage or during both phases. We also performed a time-series GC-MS analysis to monitor metabolic homeostasis status during stress and recovery. The different thermal treatments resulted in highly distinct metabolic phenotypes. Flies submitted to both developmental and adult acclimation were characterized by accumulation of cryoprotectants (carbohydrates and amino acids), although concentrations changes remained of low magnitude. After cold shock, non-acclimated chill-susceptible phenotype displayed a symptomatic loss of metabolic homeostasis, correlated with erratic changes in the amino acids pool. On the other hand, the most cold-tolerant phenotype was able to maintain metabolic homeostasis after cold stress. These results indicate that cold tolerance acquisition of D. suzukii depends on physiological strategies similar to other drosophilids: moderate changes in cryoprotective substances and metabolic robustness. In addition, the results add to the body of evidence supporting that mechanisms underlying the different forms of acclimation are distinct.

Antioxidant and Antimutagenic Activities of Different Fractions from the Leaves of Rhododendron arboreum Sm. and Their GC-MS Profiling

In this era of urbanization and environmental pollution, antioxidants and antimutagens derived from plants are promising safeguards for human health. In the current investigation, we analyzed the antioxidant and antimutagenic effects of the hexane, chloroform, and ethyl acetate fractions of Rhododendron arboreum Sm. leaves and determined their chemical composition. The different fractions inhibited lipid peroxidation, repressed the production of nitric oxide radicals, and prevented deoxyribose degradation. The antimutagenic activity of the leaf fractions was analyzed against 4-nitro-O-phenylenediamine, sodium azide and 2-aminofluorene mutagens in two test strains (TA-98 and TA-100) of Salmonella typhimurium. The experiment was conducted using pre- and co-incubation modes. The best results were obtained in the pre-incubation mode, and against indirect acting mutagen. The presence of a number of bioactive constituents was confirmed in the different fractions by GC-MS analysis. The study reveals the strong antioxidant and antimutagenic activity of R. arboreum leaves. We propose that those activities of R. arboreum might correspond to the combined effect of the phytochemicals identified by GC-MS analysis. To the best of our knowledge, this is the first report on the antimutagenic activity of R. arboreum leaves.

Metabolic Phenotyping of Anks3 Depletion in mIMCD-3 cells - a Putative Nephronophthisis Candidate

Nephronophthisis (NPH) is an autosomal recessive form of cystic kidney disease and the leading cause of hereditary kidney failure in children and young adults. Like other NPH proteins, the NPHP16/Anks6-interacting protein Anks3 has been identified to cause laterality defects in humans. However, the cellular functions of Anks3 remain enigmatic. We investigated the metabolic impact of Anks3 depletion in cultured murine inner medullary collecting duct cells via GC-MS profiling and LC-MS/MS analysis. Combined metabolomics successfully identified 155 metabolites; 48 metabolites were identified to be significantly altered by decreasing Anks3 levels. Especially, amino acid and purine/pyrimidine metabolism were affected by loss of Anks3. Branched-chain amino acids were identified to be significantly downregulated suggesting disrupted nutrient signalling. Tryptophan and 1-ribosyl-imidazolenicotinamide accumulated whereas NAD+ and NADP+ concentrations were diminished indicating disturbances within the tryptophan-niacin pathway. Most strikingly, nucleosides were reduced upon Anks3 depletion, while 5-methyluridine and 6-methyladenosine accumulated over time. Hence, elevated PARP1 and cleaved PARP1 levels could be detected. Furthermore, living cell number and viability was significantly declined. In combination, these results suggest that Anks3 may be involved in DNA damage responses by balancing the intracellular nucleoside pool.

Gas chromatography–mass spectrometry and high-performance thin-layer chromatography quantifications of some physiologically active secondary metabolites in Averrhoa carambola L. fruits

The extraction of crude drugs by using different solvents provides polarity-based fractions containing specific types of secondary metabolites. Averrhoa carambola L. fruits were extracted and fractionated, and petroleum ether extract was processed by fatty acid methyl ester (FAME) technique for characterization by gas chromatography–mass spectrometry (GC–MS) analysis. The remaining part was extracted with methanol for high-performance thin-layer chromatography (HPTLC) analysis, for simultaneous quantitative determination of gallic acid, protocatechuic acid, and quercetin in methanolic fractions. Petroleum ether and methanol fractions were found to be the best for the highest possible recovery of target analytes. The chromatographic elutions of FAME compounds generated from ether extract were evaluated by GC–MS profiling. Ten fatty acid compounds were separated with the highest quantity of oleic acid methyl ester (42.88%). On other hand, polar fraction was processed by HPTLC profiling. For achieving good separation, a mobile phase of toluene–ethyl acetate–formic acid (5:4:1, v/v) was used. The densitometric determination was carried out at 310 nm in reflection–absorption mode. The calibration curves were linear in the range of 100–600 ng per spot for gallic acid, protocatechuic acid, and quercetin. During the analysis, the dried raw material from A. carambola L. fruits showed the presence of gallic acid (0.96%), protocatechuic acid (0.05%), and quercetin (0.40%). The proposed method is simple, precise, specific, and accurate. The statistical analysis of the data obtained proves that the method is reproducible and selective and can be used for the routine analysis of the reported phenolic compounds in crude drug and extracts. The results indicated that the methanolic extracts of the plant contained a considerable amount of bioactive compounds. The presence of phytochemicals especially phenolics and flavonoids explains its use in various diseases. It may be concluded that the results obtained from the quantitative evaluation of quercetin by HPTLC fingerprinting could be useful in its authentication, the quality control of the drug, and in ensuring therapeutic efficacy.

Chemical and genetic differences between Hawaiian lineages of the alga Asparagopsis taxiformis

Asparagopsis taxiformis is a widely distributed red alga, known locally in Hawai‘i as limu kohu. Over the past decade, it was discovered that A. taxiformis is a multiphyletic species, with analysis of mitochondrial DNA revealing several genetically distinct lineages present around the world. A. taxiformis samples from around O‘ahu, belonging to the three different lineages found in Hawaiian waters, were collected and subjected to parallel genetic and chemical profiling. Sequencing of the mitochondrial cytochrome oxidase I gene clearly separated the three lineages. In parallel, GC-MS profiling of the samples was conducted in order to determine whether there were any accompanying chemical differences. Analysis of the metabolite profiles using a range of multivariate statistical techniques showed that lineage 4 could be readily differentiated from the other two lineages, lineage 1 and lineage 2, by its chemical profile. Several halogenated metabolites characteristic of lineage 4 were also identified, including methyl bromoiodoacetate, methyl tribromoacetate, carbon tetrabromide, and dibromoacetone.

GC-MS profiling of leukemia cells: an optimized preparation protocol for the intracellular metabolome

A preparation protocol for the robust and reproducible metabolome profiling of leukemia cells with relatively high coverage has been developed.

Arabidopsis ACYL-COA-BINDING PROTEIN1 interacts with STEROL C4-METHYL OXIDASE1-2 to modulate gene expression of homeodomain-leucine zipper IV transcription factors.

Fatty acids (FAs) and sterols constitute building blocks of eukaryotic membranes and lipid signals. Co-regulation of FA and sterol synthesis is mediated by sterol regulatory element-binding proteins in animals but remains elusive in plants. We reported recently that Arabidopsis ACYL-COA-BINDING PROTEIN1 (ACBP1) modulates sterol synthesis via protein-protein interaction with STEROL C4-METHYL OXIDASE1-1 (SMO1-1). Herein, ACBP1 was demonstrated to co-express and interact with SMO1-2 by yeast two-hybrid, co-localization, pull-down, co-immunoprecipitation and β-glucuronidase assays. SMO1-2 silenced in acbp1 was used in phenotyping, GC-MS and expression profiling. ACBP1 co-expressed with SMO1-2 in embryo sacs, pollen and trichomes, corroborating with cooperative tissue-specific functions unseen with SMO1-1. SMO1-2 silencing in acbp1 impaired seed development, male and female gamete transmission, and pollen function. Genes encoding homeodomain-leucine zipper IV transcription factors (HDG5, HDG10, HDG11 and GLABRA2), which potentially bind phospholipids/sterols, were transcribed aberrantly. GLABRA2 targets (MYB23, MUM4 and PLDα1) were misregulated, causing glabra2-resembling trichome, seed coat mucilage and oil-accumulating phenotypes. Together with altered sterol and FA compositions upon ACBP1 mutation and/or SMO1-2 silencing, ACBP1-SMO1 interaction appears to mediate homeostatic co-regulation of FAs and sterols, which serve as lipid modulators for gene expression of homeodomain-leucine zipper IV transcription factors.

GC–MS profiling, descriptive sensory analysis, and consumer acceptance of Costa Rican papaya (Carica papaya L.) fruit purees

Volatiles of papaya purees from four Costa Rican cultivars were analysed by solid-phase microextraction and gas chromatography-mass spectrometry. A total of 83 volatiles was assigned in the purees, of which 19 were detected for the first time as papaya constituents. As revealed by multivariate statistics, i.e., principal component analysis (PCA) and partial least squares discriminant analysis (PLS-DA), the purees may be allocated to terpene- and lactone-rich (‘SH-5’), ester-containing (‘Criolla’), and intermediate chemotypes (‘Pococí’ and ‘Silvestre’). Descriptive sensory analysis and a consumer acceptance test were additionally performed. Floral, fruity, and honey-like notes were perceived at significantly higher intensities in ‘SH-5’ puree. The latter descriptors strongly correlated with volatiles discriminating ‘SH-5’ in the PCA and PLS-DA, respectively. Consumer acceptance of the papaya purees differed significantly. ‘Pococí’ and ‘SH-5’ purees appear to be suitable for improving the nutritional value of blended fruit juices without impairing their sensory quality.

Ymr210wp leads to the accumulation of phospholipids and steryl esters in yeast.

Ymr210w was identified as a MAG (Monoacylglycerol) lipase. The accumulation of the phospholipids in the ymr210wΔ was not clearly understood. It was expressed in S. cerevisiae using pYES2/CT vector and His-tag purified recombinant protein confirmed TAG lipase activity. To further evaluate the role of YMR210w, ester hydrolase activity was also confirmed with pNP-acetate, pNP-butyrate and pNP - palmitate. GC-MS lipid profiling of ymr210wΔ showed an increase in the 15:0 Pentadecanoic acid by 76% among the total lipids. Phospholipid, Erucic acid 22:1 (Δ13) showed 43% increase while steryl esters showed significant changes with 16:0 hexadecanoic acid augmentations by 80% and 18:0 Octadecanoic acid by 165% when compared to wild type (WT). Increase in the steryl ester and TAG content supports the accumulation of lipid bodies in ymr210wΔ strain when compared with WT cells.

Making It Last: Storage Time and Temperature Have Differential Impacts on Metabolite Profiles of Airway Samples from Cystic Fibrosis Patients.

Metabolites of human or microbial origin have the potential to be important biomarkers of the disease state in cystic fibrosis (CF). Clinical sample collection and storage conditions may impact metabolite abundances with clinical relevance. We measured the change in metabolite composition based on untargeted gas chromatography-mass spectrometry (GC-MS) when CF sputum samples were stored at 4°C, -20°C, or -80°C with one or two freeze-thaw cycles. Daily measurements were taken for 1week and then weekly for 4weeks (4°C) and 8weeks (-20°C). The metabolites in samples stored at -20°C maintained abundances similar to those found at-80°C over the course of 8weeks (average change in Bray-Curtis distance, 0.06 ± 0.04) and were also stable after one or two freeze-thaw cycles. However, the metabolite profiles of samples stored at 4°C shifted after 1 day and continued to change over the course of 4weeks (average change in Bray-Curtis distance, 0.31 ± 0.12). The abundances of several amino acids and other metabolites increased with time of storage at 4°C but remained constant at -20°C. Storage temperature was a significant factor driving the metabolite composition (permutational multivariate analysis of variance: r2 = 0.32 to 0.49, P < 0.001). CF sputum samples stored at -20°C at the time of sampling maintain a relatively stable untargeted GC-MS profile. Samples should be frozen on the day of collection, as more than 1 day at 4°C impacts the global composition of the metabolites in the sample. IMPORTANCE Metabolomics has great potential for uncovering biomarkers of the disease state in CF and many other contexts. However, sample storage timing and temperature may alter the abundance of clinically relevant metabolites. To assess whether existing samples are stable and to direct future study design, we conducted untargeted GC-MS metabolomic analysis of CF sputum samples after one or two freeze-thaw cycles and storage at 4°C and -20°C for 4 to 8weeks. Overall, storage at -20°C and freeze-thaw cycles had little impact on metabolite profiles; however, storage at 4°C shifted metabolite abundances significantly. GC-MS profiling will aid in our understanding of the CF lung, but care should be taken in studies using sputum samples to ensure that samples are properly stored.

Identification of physiological changes and key metabolites coincident with postharvest internal browning of pineapple (Ananas comosus L.) fruit

Chilling causes biochemical and physiological dysfunction in pineapple fruit, evidently as internal browning (IB). Here, putative biomarkers of the early responses to chilling stress, prior to the appearance of IB, were investigated using a combination of GC–MS profiling and physiological and biochemical assay. Two cultivars – one tolerant and one susceptible – were used, and attention was made to tissue sampling from the early reversible to late irreversible phase of chilling injury. At these stages, electrolyte leakage and the levels of phenolic compounds were not altered even after prolonged chilling stress; however, changes in ascorbic acid levels and antioxidant activity coincided with IB occurrence. These data suggest that antioxidant capacity and ascorbic acid are the measurable and ‘early’ responses to IB. The GC–MS metabolite profiling data of chilled pineapple fruit presented here is the first reported. The tolerant pineapple showed greater shifts in metabolism and accumulated higher levels of amino acids and organic acids, especially, valine, cysteine, aspartate and galacturonate. In contrast, the lactic acid content of susceptible cultivar was higher upon chilled storage.

GC-MS profiling and antidepressant-like effect of the extracts of Hamelia patens in animal model

&lt;p class="Abstract"&gt;Hamelia patens is used in folk medicine in the treatment of nervous shock. The present study deals to evaluate the antidepressant-like effects of chloroform and methanol extracts of H. patens on the performance of male mice and GC-MS profiling of bioactive extract. Mice were given extracts orally in acute doses of 100 and 200 mg/kg daily for 7 days and then subjected to forced swim test, tail suspension test and open field test. Imipramine (10 mg/kg/day, p.o.) and fluxetamine (10 mg/kg/day, p.o.) were used as the standard in forced swim test and tail suspension test respectively. GC-MS profiling of chloroform extract was performed to find out the chemical constituents in bioactive fraction. After one-week treatment, the chloroform extract (100 and 200 mg/kg/day, p.o.) significantly reduced immobility time in forced swim test and tail suspension test (p&amp;lt;0.05). All extracts did not show any significant change in the locomotor activity in open field test. These data indicate that the extract of H. patens possesses antidepressant-like properties in mice without any significant effect on locomotor activity.&lt;/p&gt;&lt;p&gt;&lt;strong&gt;Video Clip of Methodology&lt;/strong&gt;:&lt;/p&gt;&lt;p&gt;4 min 06 sec: &lt;a href="https://www.youtube.com/v/DDJzDqiwCKU"&gt;Full Screen&lt;/a&gt; &lt;a href="https://www.youtube.com/watch?v=DDJzDqiwCKU"&gt;Alternate&lt;/a&gt;&lt;/p&gt;