Electrospray Ionization Triple Quadrupole Research Articles

Discriminating Benign from Malignant Lung Diseases Using Plasma Glycosaminoglycans and Cell-Free DNA.

We aimed to investigate the use of free glycosaminoglycan profiles (GAGomes) and cfDNA in plasma to differentiate between lung cancer and benign lung disease, in a cohort of 113 patients initially suspected of lung cancer. GAGomes were analyzed in all samples using the MIRAM® Free Glycosaminoglycan Kit with ultra-high-performance liquid chromatography and electrospray ionization triple quadrupole mass spectrometry. In a subset of samples, cfDNA concentration and NGS-data was available. We detected two GAGome features, 0S chondroitin sulfate (CS), and 4S CS, with cancer-specific changes. Based on the observed GAGome changes, we devised a model to predict lung cancer. The model, named the GAGome score, could detect lung cancer with 41.2% sensitivity (95% CI: 9.2-54.2%) at 96.4% specificity (95% CI: 95.2-100.0%, n = 113). When we combined the GAGome score with a cfDNA-based model, the sensitivity increased from 42.6% (95% CI: 31.7-60.6%, cfDNA alone) to 70.5% (95% CI: 57.4-81.5%) at 95% specificity (95% CI: 75.1-100%, n = 74). Notably, the combined GAGome and cfDNA testing improved the sensitivity, compared to cfDNA alone, especially in ASCL stage I (55.6% vs 11.1%). Our findings show that plasma GAGome profiles can enhance cfDNA testing performance, highlighting the applicability of a multiomics approach in lung cancer diagnostics.

Metabolomics Analysis of Phenolic Composition and Content in Five Pear Cultivars Leaves.

Phenolic compounds are the predominant chemical constituents in the secondary metabolites of plants and are commonly found in pears. In this study, we focused on the analysis of the phenolic composition and antioxidant activity of leaves from five pear cultivars (Cuiguan, Chaohong, Kuerle, Nanguoli, and Yali) and tea leaves (Fudingdabai as the control) using ultra-performance liquid chromatography coupled with electrospray ionization triple quadrupole mass spectrometry. The results indicated significant differences in the amount and composition of phenolic metabolites between tea and pear leaves, as well as among the five pear varieties. Only approximately one-third of the metabolites exhibited higher levels in pear leaves compared to that in tea leaves. The total phenol content in the Yali cultivar was higher than that in the other pear cultivars. Furthermore, specific phenolic metabolites with high expression were identified in the leaves of different groups. The levels of delphinidin 3-glucoside, aesculin, prunin, cosmosiin, quercetin 3-galactoside, isorhamnetin-3-O-glucoside, nicotiflorin, narcissin, chlorogenic acid, and cryptochlorogenic acid were relatively high among the five pear cultivars. (-)-Gallocatechin gallate, 6-methylcoumarin, aesculetin, hesperidin, kaempferol, and caftaric acid were identified as specific metabolic substances unique to each type of pear leaf. Most of the differential metabolites showed positive correlations and were primarily enriched in the flavonoid biosynthesis, flavone and flavonol biosynthesis, and phenylpropanoid biosynthesis pathways. DPPH (1,1-Diphenyl-2-picrylhydrazyl radical) analysis indicated that the Yali cultivar exhibited the highest antioxidant activity compared to other varieties. This systematic analysis of the differences in phenolic metabolite composition and antioxidant activity between pear and tea leaves provides a theoretical foundation for the development and utilization of pear leaf resources.

Longitudinal characterization of the muscle metabolome in dairy cows during the transition from lactation cessation to lactation resumption

Skeletal muscle is vital in maintaining metabolic homeostasis and adapting to the physiological needs of pregnancy and lactation. Despite advancements in understanding metabolic changes in dairy cows around calving and early lactation, there are still gaps in our knowledge, especially concerning muscle metabolism and the changes associated with drying off. This study aimed to characterize the skeletal muscle metabolome in the context of the dietary and metabolic changes occurring during the transition from the cessation of lactation to the resumption of lactation in dairy cows. Twelve Holstein dairy cows housed in tie stalls were dried off 6 weeks (wk) before the expected calving date. Cows were individually fed ad libitum total mixed rations composed of grass silage, corn silage, and concentrate during lactation and of corn silage, barley straw, and concentrate during the dry period. The metabolome was characterized in skeletal muscle samples (M. longissimus dorsi) collected on wk -7 (9 d before dry-off), -5 (6 d after dry-off), and wk -1, and 1 relative to calving. The targeted metabolomics approach was conducted using the MxP Quant 500 kit (Biocrates Life Sciences AG) with liquid chromatography, flow injection, and electrospray ionization triple quadrupole mass spectrometry. Statistical analysis on the muscle metabolite data was performed using MetaboAnalyst 5.0, which allowed us to conduct various multivariate analyses such as principal component analysis (PCA), partial least squares discriminant analysis (PLS-DA), informative heat map generation, and hierarchical clustering. The statistical analysis revealed a clear separation between pregnancy (wk -7, -5, and -1) and post-calving (wk 1). Starting 5 wk before calving and continuing through the first wk thereafter, the concentration of 3-methylhistidine (3-MH) in the muscle increased. This coincided with an increase in the concentrations of 11 AA (Phe, His, Tyr, Trp, Arg, Asn, Leu, Ile, Gly, Ser, and Thr) in the first wk after calving, whereas Gln decreased. l-arginine pathway metabolites (homoarginine, ornithine, citrulline, and asymmetric dimethylarginine), betaine, and sarcosine followed a similar pattern, increasing from wk -7 to -5, but decreasing from wk -1 to 1. The transition from pregnancy to lactation was associated with an increase in concentrations of the long-chain acylcarnitine species C16, C16:1, C18, and C18:1 in the muscle, whereas the concentrations of phosphatidylcholine and sphingomyelin in the muscle remained stable. The significant changes observed in the metabolome mainly concerned the AA and AA-related metabolites, indicating muscle protein breakdown in the first wk after calving. The metabolites produced by the L-Arg pathway might contribute to regulating skeletal muscle mass and function in periparturient dairy cows. The elevated concentrations of long-chain acylcarnitine species in the muscle in the first wk after calving suggest incomplete fatty acid oxidation, likely due to insufficient metabolic adaptation in response to the fatty acid load around the time of calving.

Rapid determination of four flavonoids from hawthorn leaves in plasma by ultra‐high performance liquid chromatography‐tandem mass spectrometry and study on its pharmacokinetics

AbstractIn this study, a rapid and sensitive ultra‐high performance liquid chromatography‐tandem mass spectrometry method was established for the simultaneous determination of four Vitexin‐4“‐O‐glucoside (VOG), Vitexin‐2”‐O‐rhamnoside (VOR), vitexin (VIT), and hyperoside (HP) in plasma, which are representative flavonoids constituents of hawthorn leaves. The four components were separated by CORTECS UPLC C18 column (100 × 3.0 mm, 1.6 µm), using acetonitrile‐water containing 0.02% formic acid and 2 mmol/L ammonium formate as the mobile phase, the gradient elution flow rate was 0.30 mL/min. The analyte was quantified using an electrospray ionization triple quadrupole mass spectrometer. The analytes and baicalin internal standard were performed at m/z 593.2/413.2 for VOG, m/z 577.2/413.1 for VOR, m/z 431.1/310.9 for VIT, m/z 463.2/300 for HP and m/z 445.1/269 for internal standard. Among the four flavonoids, the correlation coefficient of the standard curve of each component is between 0.9939 and 0.9987. After oral administration of hawthorn leaf extract, VOG and VOR showed similar pharmacokinetic characteristics; VIT was eliminated in the body more slowly than the other three flavonoids and distributed more in the tissues; HP can be quickly absorbed in the blood. The developed analytical method has high sensitivity, precision, and accuracy with no interference from plasma components and has been successfully used in pharmacokinetic studies.

Longitudinal characterization of the metabolome of dairy cows transitioning from one lactation to the next: Investigations in the liver

This study aimed to investigate the metabolic changes in the livers of dairy cows from 1 wk before dry-off to 1 wk after calving. Twelve high-yielding Holstein cows were included in a longitudinal study and housed in a tie-stall barn. The cows were dried off at 6 wk before the expected calving date (dry period length = 42 d). During the entire lactation, the cows were milked twice daily at 0600 and 1700 h. Liver biopsies were taken from each cow at 4 different times: wk -7 (before drying off), -5 (after drying off), -1 and +1 relative to calving. A targeted metabolomics approach was performed by liquid chromatography and flow injection with electrospray ionization triple quadrupole mass spectrometry using the MxP® Quant 500 kit. A total of 185 metabolites in the liver were used for the final data analysis. Principal component analysis (PCA) revealed a clear separation by days of sampling, indicating a notable shift in metabolic phenotype from late lactation to dry period and further changes after calving. Changes were observed in several classes of compounds, including amino acids and biogenic amines. In particular, the changes in acylcarnitines (AcylCN), phosphatidylcholines (PC), sphingomyelins (SM) and bile acids (BA) indicated extensive remodeling of the hepatic lipidome. The changes in AcylCN concentrations in early lactation suggest incomplete fatty acid oxidation in the liver, possibly indicating mitochondrial dysfunction or enzymatic imbalance. In addition, the changes in PC and SM species in early lactation indicate altered cell membrane composition, which may affect cell signaling and functionality. In addition, changes in BA concentrations and profiles indicate dynamic adaptations in BA synthesis, lipid digestion and absorption during the observation period. In particular, PCA analysis showed an overlapping distribution of liver metabolites in primiparous and multiparous cows, indicating no significant difference between these groups. In addition, Volcano plots showed similar liver metabolism between primiparous and multiparous cows, with no significant fold changes (>1.5) in any metabolite at significant P-values (false discovery rate <0.05). These results provide valuable insight into the physiological ranges of liver metabolites during dry period and calving in healthy dairy cows and should contribute to the design and interpretation of future metabolite-based studies of the transition dairy cow.

A Qualitative Method to Detect Paraproteins from Serum Using Ultra Performance Liquid Chromatography Electrospray Triple Quadrupole Mass Spectrometry.

Mass spectrometry-based techniques are increasingly reported in the literature for identifying paraproteins due to their improved specificity and sensitivity. The present study demonstrates the capability of ultra performance liquid chromatography (UPLC) electrospray ionization triple quadrupole mass spectrometry for the qualitative analysis of paraproteins. Paraproteins from patient serum (n = 40) were immunopurified using agarose beads coated with camelid antibodies that are specific for various subtypes of immunoglobulins (Igs; G, A, M, and light chains κ, λ). The extracted Igs are reduced to separate light chains from heavy chains in solution. The reduced sample was subjected to UPLC and mass measured using electrospray ionization-mass spectrometry. The mass spectral peaks at specific retention times were deconvoluted after clean-up to obtain the mass of light chains. The interpretation of liquid chromatography peaks and LC-MS data was validated by comparing them with immunofixation electrophoresis (IFE) results. The interpretation from the chromatographic pattern had a 92.5% (37/40) agreement when compared with mass information. The correlation of mass spectrometry data to IFE was 90% (36/40). The high mass of light chains (>25 kDa) was suggestive of glycosylation. Patient sera positive for IgGκ on IFE (n = 15) were analyzed for the interference of tAbs. The mass of Daratumumab observed in a sample was confirmed by the treating physician. A biclonal of same isotype (IgGκ) was identified. The feasibility of using liquid chromatography triple quadrupole mass spectrometry for the identification of the subtype of paraproteins has been demonstrated. The method's applicability to screen for interference from tAbs and identification of biclonals of the same isotype has been highlighted.

Development and Validation of a Method for Hydrolysis and Analysis of Amino Acids in Ruminant Feeds, Tissue, and Milk Using Isotope Dilution Z-HILIC Coupled with Electrospray Ionization Triple Quadrupole LC-MS/MS.

Current analytical methods for amino acid (AA) analysis in ruminant nutrition are time-consuming and expensive. This study aimed to develop a method for AA analysis that is faster, more efficient, rugged, and accessible. Four representative matrixes were selected for method development and validation: milk, tissue, feed, and soy flour standard reference material from National Institute of Standards and Technology. Acid and alkaline hydrolysis were used to analyze 18 AA. Separation of AA was performed using a Z-HILIC column in an 18-min run coupled to a triple quadrupole LC/MS system in positive and negative electrospray ionization for identification and quantitation. The method was evaluated for recovery, precision, calibration curve linearity, and limits of detection (LODs) and limits of quantitation (LOQs) and applied to other feed samples. Good quantitation results were achieved for all AA, with coefficients of determination (R2) over 0.995; LODs at 0.2-28.2 and LOQs at 0.7-94.1 ng/mL; intraday and interday precision <14.9% relative standard deviation; blank recovery between 75.6 and 116.2%; and sample recovery between 75.6 and 118.0%. Overall, AA concentrations were similar to literature values, and there was a tendency for higher N recovery as AA. In conclusion, an efficient and robust method was validated to routinely analyze AA for appropriate characterization in diet formulation for dairy cattle.

Determination of 27 bovine plasma amino acids and metabolites using zwitterionic-hydrophilic interaction liquid chromatography coupled with isotope dilution electrospray ionization triple quadrupole liquid chromatography-mass spectrometry and the effect of deproteinization timing

The use of zwitterionic-hydrophilic interaction liquid chromatography (Z-HILIC) columns for analysis of underivatized analytes has allowed simpler sample preparation of bovine plasma for sensitive and selective analysis, when coupled with mass spectrometry (MS). The objective of this study was to evaluate and validate this analytical technique to measure AA and metabolites in bovine plasma at 2 deproteinization times. A robust method using Z-HILIC coupled to a triple quadrupole MS (TQMS) was evaluated and validated to quantitatively analyze 19 AA using isotope dilution and 8 AA metabolites qualitatively in bovine deproteinized plasma. The timing of deproteinization was investigated to determine if plasma should be deproteinized upon collection (on-site) or immediately before analysis (in-lab). Analytes were separated using a Z-HILIC column in a 21 min run and analyzed with a TQMS in positive electrospray ionization for identification and quantification. The method was validated for standard curve linearity, limits of detection (LOD) and quantification (LOQ), intra- and interday precision (% coefficient of variation; CV), recovery (%), and freeze-thaw stability (% CV) after 1 mo. Coefficients of determination (R2) were over 0.993, and LOD and LOQ were below measured values for all AA. The CV for the intraday and interday precision were below 18%, except for cystine (Cys2) and Orn in-lab. Recoveries on-site and in-lab ranged from 75% to 120% for all analytes except Cys2 in-lab. Most analytes were stable after 1 mo of freezing regardless of deproteinization timing, CV <25%, except for hydroxyproline (Hyp). The concentration of Cys2 was affected by deproteinization in-lab compared with on-site, and even though Glu and Hyp were different between the 2 deproteinization timings, the concentrations between the 2 timings were within the standard deviation.

Longitudinal characterization of the metabolome of dairy cows transitioning from one lactation to the next: Investigations in blood serum

The objective of this study was to characterize changes in the serum metabolome and various indicators of oxidative balance in dairy cows starting 2 wk before dry-off and continuing until wk 16 of lactation. Twelve Holstein dairy cows (body weight 745 ± 71 kg, body condition score 3.43 ± 0.66; mean ± SD) were housed in a tiestall barn from 10 wk before to 16 wk after parturition. Cows were dried off 6 wk before the expected calving date (mean dry period length = 42 d). From 8 wk before calving to 16 wk after calving, blood samples were taken weekly to study redox metabolism by determining antioxidant capacity, measured as the ferric-reducing ability of plasma, reactive oxidative metabolites, oxidative stress index, oxidative damage of lipids, measured as thiobarbituric acid reactive substances, and glutathione peroxidase activity. According to these results, dairy cows had the lowest serum antioxidant capacity and greater levels of oxidative stress during the dry-off period and the early postpartum period. For metabolomics, a subset of serum samples including wk -7 (before dry-off), -5 (after dry-off), -1, 1, 5, 10, and 15 relative to calving were used. A targeted metabolomics approach was performed using liquid chromatography and flow injection with electrospray ionization triple quadrupole mass spectrometry using the MxP Quant 500 kit (Biocrates Life Sciences AG). A total of 240 metabolites in serum were used in the final data analysis. Principal component analysis revealed a clear separation by days of sampling, indicating a remarkable shift in metabolic phenotype between the dry period and late and early lactation. Changes in many non-lipid metabolites associated with one-carbon metabolism, the tricarboxylic acid cycle, the urea cycle, and AA catabolism were observed in the study, with changes in AA serum concentrations likely related to factors such as energy and nitrogen balance, digestive efficiency, and changing diets. The study confirmed an extensive remodeling of the serum lipidome in peripartum dairy cows, highlighting the importance of changes in acylcarnitine (acylCN), phosphatidylcholines (PC), and triacylglycerols (TG), as they play a crucial role in lipid metabolism. Results showed that short-chain acylCN increased after dry-off and decreased thereafter, whereas lipid-derived acylCN increased around parturition, suggesting that more fatty acids could enter mitochondria. Phospholipids and sphingolipids in serum showed changes during lactation. In particular, concentrations of sphingomyelins, PC, and lysoPC decreased around calving but increased in mid- and late lactation. In contrast, concentrations of TG remained consistently low after parturition. The serum concentrations of bile acids fluctuated during the dry period and lactation, with glycocholic acid, cholic acid, glycodeoxycholic acid, and taurocholic acid showing the greatest concentrations. These changes are likely due to the interplay of diet, liver function, and the ability of the gut microbiota to convert primary to secondary bile acids. Overall, these descriptive results may aid in hypothesis generation and in the design and interpretation of future metabolite-based studies in dairy cows. Furthermore, they contribute to our understanding of the physiological ranges in serum metabolites relative to the lactation cycle of the dairy cow.

Coumarins of Ferulopsis hystrix: LC–MS Profiling and Gastroprotective and Antioxidant Activities of Skimmin and Peucenidin

Ferulopsis hystrix is a perennial plant of the Apiaceae family. In Buryat and Mongolian medicine, it is used as a substitute for Costus speciosus roots (ru rta), and in Tibetan medicine, it is used to cure digestive system diseases and used as a wound-healing agent. However, its metabolites and their bioactivities are still poorly understood. High-performance liquid chromatography with photodiode array detection and electrospray ionization triple quadrupole mass-spectrometric detection (HPLC–PDA–ESI–tQ–MS/MS) were used to identify approximately 42 phenolic compounds in F. hystrix, and 30 coumarins were characterized and quantified. The major compounds in F. hystrix roots were skimmin (umbelliferone 7-O-glucoside) and peucenidin (vaginidiol 3′-O-acetyl-4′-O-senecioyl ester), of which, gastroprotective and antioxidant effects were found for the first time. The abovementioned compounds exhibit a gastroprotective effect against indomethacin and steroid gastropathy by reducing the amount of damage (point, large, and strip-like erosions) in the gastric mucosa and lowering the corresponding Paul’s index. The most pronounced gastroprotective effect was exhibited by skimmin at a dose of 1 mg/kg and by peucenidin at doses of 16 and 48 mg/kg; these compounds help to limit the development of pronounced erosive–necrotic processes in the gastric mucosa. In pathological conditions, these compounds reduce malondialdehyde, increase the activity of catalase, and increase the content of reduced glutathione in the blood. Thus, this study demonstrates that F. hystrix roots are a good source of bioactive coumarins with gastroprotective potential, which supports earlier ethnopharmacological studies.

Metabolites of Geum aleppicum and Sibbaldianthe bifurca: Diversity and α-Glucosidase Inhibitory Potential.

α-Glucosidase inhibitors are essential in the treatment of diabetes mellitus. Plant-derived drugs are promising sources of new compounds with glucosidase-inhibiting ability. The Geum aleppicum Jacq. and Sibbaldianthe bifurca (L.) Kurtto & T.Erikss. herbs are used in many traditional medical systems to treat diabetes. In this study, metabolites of the G. aleppicum and S. bifurca herbs in active growth, flowering, and fruiting stages were investigated using high-performance liquid chromatography with photodiode array and electrospray ionization triple quadrupole mass spectrometric detection (HPLC-PDA-ESI-tQ-MS/MS). In total, 29 compounds in G. aleppicum and 41 components in S. bifurca were identified including carbohydrates, organic acids, benzoic and ellagic acid derivatives, ellagitannins, flavonoids, and triterpenoids. Gemin A, miquelianin, niga-ichigoside F1, and 3,4-dihydroxybenzoic acid 4-O-glucoside were the dominant compounds in the G. aleppicum herb, while guaiaverin, miquelianin, tellimagrandin II2, casuarictin, and glucose were prevailing compounds in the S. bifurca herb. On the basis of HPLC activity-based profiling of the G. aleppicum herb extract, the most pronounced inhibition of α-glucosidase was observed for gemin A and quercetin-3-O-glucuronide. The latter compound and quercetin-3-O-arabinoside demonstrated maximal inhibition of α-glucosidase in the S. bifurca herb extract. The obtained results confirm the prospects of using these plant compounds as possible sources of hypoglycemic nutraceuticals.

Hogweed Seed Oil: Physico-Chemical Characterization, LC-MS Profile, and Neuroprotective Activity of Heracleum dissectum Nanosuspension.

The seeds of dissected hogweed (Heracleum dissectum Ledeb., Apiaceae) are the source of hogweed oil (HSO), which is still underexplored and requires careful chemical and biological studies. The performed physico-chemical analysis of HSO elucidated basic physical characteristics and revealed the presence of fatty acids, essential oil components, pigments, and coumarins. High-performance liquid chromatography with photodiode array detection and electrospray ionization triple quadrupole mass spectrometric detection (HPLC-PDA-ESI-tQ-MS/MS) identified 38 coumarins that were characterized and quantified. Various furanocoumarins were the major components of HSO polyphenolics, including imperatorin, phellopterin, and isoimperatorin, and the total coumarin content in HSO varied from 181.14 to 238.42 mg/mL. The analysis of storage stability of the selected compounds in HSO indicated their good preservation after 3-year storage at cold and freezing temperatures. The application of the CO2-assisted effervescence method allowed the production of an HSO nanosuspension, which was used in a brain ischemia model of rats. The HSO nanosuspension enhanced cerebral hemodynamics and decreased the frequency of necrotic processes in the brain tissue. Thus, H. dissectum seeds are a good source of coumarins, and HSO nanosuspension promotes neuroprotection of the brain after lesions, which supports earlier ethnopharmacological data.

A passive membrane system for on-line mass spectrometry reagent addition.

Post-separation addition of chemical modifiers in liquid chromatography-mass spectrometry is widely used for improving ionization sensitivity and selectivity. This is typically accomplished using a post-column T-junction, which can result in sample dilution and imperfect mixing. We present a passive semi-permeable hollow fiber membrane approach for the addition of chemical modifiers that avoids these issues. Model compounds were directly infused by flow injection to an electrospray ionization triple quadrupole mass spectrometer after passing through a polydimethylsiloxane hollow fiber membrane. Ionization enhancement reagents were introduced into the flowing stream by membrane permeation from aqueous solutions. Ionization enhancement from volatile acids and bases in positive and negative electrospray ionization was evaluated to assess the feasibility of this approach. The membrane-based apparatus resulted in relative ionization enhancement factors of up to 14×, depending upon the analyte, reagent, and ionization mode used. Ionization enhancement signal stability is reasonable (relative standard deviation of 5-7%) for extended periods from the same reagent solution, and minimal analyte dilution is observed. A proof-of-concept demonstration of the chromatographic "trifluoroacetic acid fix" strategy is presented. An on-line mass spectrometry ionization reagent addition method with potential post-chromatography reagent addition applications was developed using a hollow fiber polydimethylsiloxane membrane. This approach offers a promising alternative to traditional methods requiring additional hardware such as pumps and T-junctions that can result in sample dilution and imperfect reagent mixing.

Comprehensive Evaluation of the Quality of Tripterygium Glycosides Tablets Based on Multi-Component Quantification Combined with an In Vitro Biological Assay.

Tripterygium glycosides tablets (TGTs) are widely used in clinical practice to treat rheumatoid arthritis and other autoimmune diseases, with significant beneficial effects but also high toxicity, necessitating rigorous quality evaluation and control. In current study, a rapid resolution liquid chromatography tandem electrospray ionization triple quadrupole mass spectrometry (RRLC–ESI–MS/MS) method was developed and validated for the quantitative analysis of 14 components of ten batches of TGTs produced by different manufacturers, including four diterpenoids, three triterpenoids, and seven sesquiterpene alkaloids. Meanwhile, the NO inhibition effects of these TGTs were evaluated in LPS-induced RAW264.7 cells for their downstream anti-inflammatory activities, as well as their cytotoxicity. The results indicate that the TGTs from different manufacturers showed poor quality consistency, as evidenced by large variations in chemical profiles and biological effects, which may increase the risks associated with clinical use. To improve the quality status of TGTs, it is crucial to identify indicator components whose characterization can accurately reflect the efficacy and toxicity of TGTs from which they were derived. Our study reveals that triptolide, triptoquinone B, celastrol, and demethylzelaysteral considerably contributed to the anti-inflammatory activity and/or cytotoxicity of TGTs, implying that they should be further investigated as candidate indicator components for TGT quality control.

Influence of ripening stage and meteorological parameters on the accumulation pattern of polyphenols in greengages (Prunus mume Sieb. Et Zucc) by widely targeted metabolomic

Greengage is a Chinese traditional medicine food homology plant that contains abundant polyphenols. Greengages with different levels of maturity have different medicinal functions and applications. Therefore, this study comprehensively analysed the phenolic compounds in greengage of whole mature stage. Notably, the influence of climate conditions on greengage polyphenol synthesis was deeply explored. The polyphenols of greengages were profiled based on the widely targeted metabolomic technology using ultra-performance liquid chromatography–electrospray ionisation triple quadrupole mass spectrometry (UPLC–QTRAP–MS/MS). A total of 214 polyphenols were detected, covering 11 subclasses. During the ripening of greengages, these polyphenols first increased and then decreased, peaking during the swell stage. Multivariate statistical methods, including redundancy analysis (RDA), random forest analysis, Mantel test, and Spearman's correlation indicated that temperature, sunshine hours, humidity, and radiation were important factors driving the formation and changes in the polyphenols of greengages. In particular, flavanones and flavonols, showed a structure-dependent response to temperature and radiation.

A Simple, Fast, and Reliable LC-MS/MS Method for the Measurement of Homovanillic Acid and Vanillylmandelic Acid in Urine Specimens.

Homovanillic acid (HVA) and vanillylmandelic acid (VMA) are catecholamine metabolites used in the diagnostic workup of neuroendocrine tumors. Here we describe a simple dilute-and-shoot method for simultaneously quantitating HVA and VMA in human urine specimens. The method employs analyte separation on a reverse-phase liquid chromatography column followed by detection using electrospray ionization triple quadrupole mass spectrometry (ESI-MS/MS), wherein qualifier and quantifier ion transitions are monitored. This is a simple and fast analytical method with an injection-to-injection time of 4min.

Phytohormones and Elicitors Enhanced the Ecdysteroid and Glycosylflavone Content and Antioxidant Activity of Silene repens

In the course of the ongoing chemical study of species of Silene genus, S. repens Patrin as a common species of the genus, was selected as the object of this study. Using high-performance liquid chromatography with photodiode array detection and electrospray ionization triple quadrupole mass spectrometric detection (HPLC-PDA-ESI-tQ-MS), the presence of 12 ecdysteroids and 6 glycosylflavones was established in S. repens introduced seedlings. 20-Hydroxyecdysone and polypodine B, as well as sileneside E and schaftoside, were the dominant compounds in introduced seedlings of S. repens. The effect of exogenous phytohormones and elicitors on the productivity and accumulation of ecdysteroids and glycosylflavones in introduced seedlings of S. repens was investigated for the first time. It was found that the use of ethyl arachidonate (100 mg/L) to increase the productivity of S. repens is justified. To obtain S. repens with a high content of ecdysteroids and glycosylflavones, it is recommended to apply epibrassinolide (100 mg/L) and 4-chlorophenoxyacetic acid (100 mg/L), respectively. Antioxidant activity of S. repens against 2,2-diphenyl-1-picrylhydrazyl radical (DPPH·) was determined, and it was revealed that sileneside E and schaftoside, as well as meloside A and isovitexin had the highest antioxidant activity among the studied compounds of S. repens.

Stable Isotope Phosphate Labelling of Diverse Metabolites is Enabled by a Family of 18 O-Phosphoramidites.

Stable isotope labelling is state‐of‐the‐art in quantitative mass spectrometry, yet often accessing the required standards is cumbersome and very expensive. Here, a unifying synthetic concept for 18O‐labelled phosphates is presented, based on a family of modified 18O2‐phosphoramidite reagents. This toolbox offers access to major classes of biologically highly relevant phosphorylated metabolites as their isotopologues including nucleotides, inositol phosphates, ‐pyrophosphates, and inorganic polyphosphates. 18O‐enrichment ratios >95 % and good yields are obtained consistently in gram‐scale reactions, while enabling late‐stage labelling. We demonstrate the utility of the 18O‐labelled inositol phosphates and pyrophosphates by assignment of these metabolites from different biological matrices. We demonstrate that phosphate neutral loss is negligible in an analytical setup employing capillary electrophoresis electrospray ionisation triple quadrupole mass spectrometry.

Stable Isotope Phosphate Labelling of Diverse Metabolites is Enabled by a Family of 18O-Phosphoramidites.

Stable isotope labelling is state-of-the-art in quantitative mass spectrometry, yet often accessing the required standards is cumbersome and very expensive. Here, a unifying synthetic concept for 18O-labelled phosphates is presented, based on a family of modified 18O2-phosphoramidite reagents. This toolbox offers access to major classes of biologically highly relevant phosphorylated metabolites as their isotopologues including nucleotides, inositol phosphates, -pyrophosphates, and inorganic polyphosphates. 18O-enrichment ratios >95 % and good yields are obtained consistently in gram-scale reactions, while enabling late-stage labelling. We demonstrate the utility of the 18O-labelled inositol phosphates and pyrophosphates by assignment of these metabolites from different biological matrices. We demonstrate that phosphate neutral loss is negligible in an analytical setup employing capillary electrophoresis electrospray ionisation triple quadrupole mass spectrometry.

Metabolites of Prickly Rose: Chemodiversity and Digestive-Enzyme-Inhibiting Potential of Rosa acicularis and the Main Ellagitannin Rugosin D.

Prickly rose (Rosa acicularis Lindl.) is the most distributed rose species in the Northern Hemisphere, used by indigenous people for various food purposes. The lack of detailed information about the chemical composition of R. acicularis has led us to study the phytochemical composition and metabolic profile of prickly rose extracts using chromatographic techniques. Many groups of phenolic and non-phenolic compounds were quantified in the leaves, flowers, roots and fruits of R. acicularis. Phenolic compounds were the dominant phytochemicals in the aerial parts and roots of R. acicularis. A precise study by high-performance liquid chromatography with photodiode array detection and electrospray ionization triple quadrupole mass spectrometric detection showed the presence of 123 compounds, among which ellagic acid derivatives, ellagitannins, gallotannins, catechins, catechin oligomers, hydroxycinnamates and flavonoid glycosides of kaempferol, quercetin and dihydroquercetin were all identified for the first time. The most abundant phenolic compounds were ellagitannins and flavonoid glycosides, with a maximal content of 70.04 mg/g in leaves and 66.72 mg/g in flowers, respectively, indicating the great ability of R. acicularis organs to accumulate phenolic compounds. By applying a standardized static, simulated gastrointestinal digestion method, we found the inhibitory potential of the leaf extract against digestive α-amylases. A pancreatic α-amylase activity-inhibiting assay coupled with HPLC microfractionation demonstrated high inhibition of enzyme activity by ellagitannin rugosin D, which was later confirmed by a microplate reaction with mammalian α-amylases and the simulated digestion method. This study clearly demonstrates that R. acicularis leaf extract and its main component, ellagitannin rugosin D, strongly inhibit digestive α-amylase, and may be a prospective antidiabetic agent.