Ultra-high-performance Liquid Chromatography Coupled With Quadrupole Time-of-flight Mass Spectrometry Research Articles

Analysis and evaluation on Paeoniae Radix Alba from different cultivars by UPLC-Q-TOF-MS and HPLC

In this study, an established ultra-high performance liquid chromatography coupled with quadrupole-time-of-flight mass spectrometry(UPLC-Q-TOF-MS) method was combined with multivariate statistical analysis to investigate the commonality and difference of main chemical components in the medicinal parts of Paeonia lactiflora from different cultivars; in addition, a high performance liquid chromatography(HPLC) method was established to simultaneously determine the content of eight active components in Paeoniae Radix Alba. Non-targeted analysis was carried out by UPLC-Q-TOF-MS on a Waters ACQUITY UPLC BEH C_(18)(2.1 mm×100 mm, 1.7 μm) column with a gradient elution of 0.1% aqueous formic acid(A)-acetonitrile(B) as the mobile phase at a flow rate of 0.2 mL·min~(-1). The column temperature was 30 ℃, and an electrospray ionization source was used to acquire mass spectrometry data in positive and negative ion modes. According to the accurate molecular weight and fragment ion information provided by multi-stage mass spectrometry and by comparison with reference substances and literature reports, thirty-six identical components were identified in Paeoniae Radix Alba from different cultivars with positive and negative ion modes. In the negative ion mode, two groups of samples were well separated; specifically, seventeen components with significant differences in content were screened and identified, and one component unique in "Bobaishao" was obtained. Quantitative analysis was conducted by high-performance liquid chromatography(HPLC) on an Agilent HC-C_(18)(4.6 mm×250 mm, 5 μm) column with a gradient elution of 0.1% aqueous phosphoric acid(A)-acetonitrile(B) as the mobile phase at a flow rate of 1.0 mL·min~(-1). The column temperature was 30 ℃ and the detection wavelength was at 230 nm. An HPLC method was developed for the simultaneous determination of eight active components(gallic acid, oxypaeoniflorin, catechin, albiflorin, paeoniflorin, galloylpaeoniflorin, 1,2,3,4,6-O-pentagalloylglucose, benzoyl-paeoniflorin) in Paeoniae Radix Albaa from different cultivars. Satisfactory linearity was achieved within the investigated linear ranges and with fine coefficients(r>0.999 0), and the methodological investigation showed that the method had good precision, repeatability and stability. The mean recoveries were 90.61% to 101.7% with RSD of 0.12% to 3.6%(n=6). UPLC-Q-OF-MS provided a rapid and efficient qualitative analytical method for the identification of the chemical components in Paeoniae Radix Alba, and the developed HPLC method was simple, rapid and accurate, which could provide a scientific basis for the evaluation of the germplasm resources and herbal quality of Paeoniae Radix Alba from different cultivars.

Rapid flavonoid-focused sub-chemome characterization of Draconis Sanguis using UPLC-Q-TOF-MS in combination with molecular weight imprinting and mass defect filtering

Draconis Sanguis is a precious Chinese medicinal material for activating blood and resolving stasis, and its effective components are flavonoids. However, the structural diversity of flavonoids in Draconis Sanguis brings great challenges to the in-depth chara-cterization of its chemical composition profiles. To clarify the substance basis of Draconis Sanguis, ultra-high performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry(UPLC-Q-TOF-MS) was used in this study to acquire MS data of Draconis Sanguis. The molecular weight imprinting(MWI) and mass defect filtering(MDF) were developed for rapid screening of flavonoids in Draconis Sanguis. Full-scan MS and MS~2 were recorded within the mass range m/z 100-1 000 in positive ion mode. Accor-ding to previous literature, MWI was employed to hunt for reported flavonoids in Draconis Sanguis, and the mass tolerance range of [M+H]~+ was set as ±10×10~(-3). A five-point MDF screening frame was further constructed to narrow the screening range of flavonoids from Draconis Sanguis. Combined with diagnostic fragment ions(DFI) and neutral loss(NL) as well as mass fragmentation pathways, 70 compounds were preliminarily identified from the extract of Draconis Sanguis, including 5 flavan oxidized congeners, 12 flavans, 1 dihydrochalcones, 49 flavonoids dimers, 1 flavonoids trimer and 2 flavonoid derivatives. This study clarified the chemical composition of flavonoids in Draconis Sanguis. Moreover, it also showed that high-resolution MS combined with data post-processing methods such as MWI and MDF could achieve rapid characterization of the chemical composition in Chinese medicinal materials.

Serum and urine metabolomics analyses reveal metabolic pathways and biomarkers in relation to nasopharyngeal carcinoma.

Nasopharyngeal carcinoma (NPC) is a malignant tumor that is endemic in Southeast Asia, North Africa, and southern China. There is an urgent need for effective early diagnosis and treatment of this disease since NPC is currently often detected at advanced stages. To reveal the underlying metabolic mechanisms and discover potential diagnostic biomarkers of NPC, we employed ultrahigh-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS) and UHPLC-Q-Exactive Orbitrap MS, respectively, to analyze 54 serum samples and 54 urine samples from 27 patients with NPC and 27 healthy control individuals. A total of 1230 metabolites were determined in serum samples, and 181 of the 1230 metabolites were significantly changed in NPC patients. The 181 metabolites were enriched in 16 pathways, including biosynthesis of unsaturated fatty acids, cholesterol metabolism, and ferroptosis. A total of 2509 metabolites were detected in the urine samples. Among them, 179 metabolites were significantly altered in NPC patients, and these metabolites were enriched in eight pathways, including the tricarboxylic acid (TCA) cycle and caffeine metabolism. Seven metabolites, including creatinine and paraxanthine, were found to be significantly changed in both NPC serum and urine samples. Based on them, further biomarker analysis revealed that the panel of three serum metabolites, octanoylcarnitine, creatinine, and decanoyl-l-carnitine, displayed a perfect diagnostic performance (area under the curve [AUC] = 0.973) to distinguish NPC patients from controls, while the other three-metabolite biomarker panel, consisting of stachydrine, decanoyl-l-carnitine, and paraxanthine, had an AUC = 0.809 to distinguish NPC and control in urine samples. This work highlights the key metabolites and metabolic pathways disturbed in NPC and presents potential biomarkers for effective diagnosis of this disease.

Chai-Gui Decoction and its representative components ameliorate spontaneous hypertension rats by modulating lipid metabolism and gut microbiota

Ethnopharmacological relevanceHypertension coincides with the category of “vertigo” and/or “headache” on the basis clinical manifestations and traditional Chinese medicine (TCM) theory. Chai-Gui Decoction (CGD), which is in usage for relieving “vertigo” and/or “headache”, had been demonstrated to be useful in ameliorating hypertension. Aim of studyThis study was planned to investigate the mechanism of CGD and its components in hypertension by using spontaneous hypertension rat (SHR). Materials and methodsCGD extract and its classification component samples (compounds in plasma, CP; compounds in gut, CG; compounds in plasma and gut, CPG) were prepared for animal experiment. SHR rats were induced with CGD extract (3 g/kg/d BW, 5 g/kg/d BW, 15 g/kg/d BW) and CGD-component classes (CP = 19.501 mg/kg/d, CG = 5.240 mg/kg/d, CPG = 24.741 mg/kg/d) for 4 weeks. Blood pressure (BP) and indexes of renin-angiotensin-aldosterone system (RAAS system) were measured. Histopathology was carried out to assess the efficacy of CGD and its components on aorta tissues. Untargeted metabolomics of lipid from rat serum samples were applied by Ultra-High performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS) and chemometric analysis to explore the relationship between metabolic pathways and hypertension. 16S rRNA gene sequencing of rat colon content and bioinformatics analysis were used to characterize the effects of CGD and its components on the gut microbiota composition of SHR rats. ResultsCGD and its component mixtures showed antihypertensive effect on SHR rats, decreased the blood pressure and reduced the aortic wall thickness in SHR rats. CGD and its component mixtures could improve the RAAS in SHR rats, including increase the percentage of angiotensin 1-7 (Ang 1–7), decrease the percentage of angiotensin II (Ang II), and decrease the Ang Ⅱ/Ang 1–7 ratio. CGD and its component mixtures could regulate the metabolome in SHR rats, mainly as decreasing the higher serum levels of Lysophosphatidylcholine (LPC) 16: 0, LPC 20: 4, and LPC 22: 6. In addition, bacteria from family S24-7 were negatively correlated with levels of LPE 16:0, LPE 18:0, LPE 18:1, and LPE 18:2. ConclusionCGD and its component mixtures exhibited antihypertensive effect on SHR rats. The underlying mechanism could be related to modulation on RAAS, LPC metabolism and the bacterial abundance of family S24-7 in gut.

Characterization of seven new related impurities and forced degradation products of tetracaine hydrochloride and proposal of degradation pathway by UHPLC-Q-TOF-MS

A rapid and highly sensitive method was developed for separation and identification of the related impurities and degradation products in tetracaine hydrochloride by ultra-high performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS). The chromatographic separation was achieved on an Agilent Infinity Lab Poroshell 120 EC-C18 column (4.6 ×100 mm, 2.7 µm) using gradient elution with mobiles phase of A (10 mM ammonium acetate buffer containing 0.1% formic acid) and B (acetonitrile) at a flow rate of 1.0 mL/min. Forced degradation experiments were also performed under acidic, alkaline, thermal, photolytic, and oxidative stress conditions following ICH guidance. The result revealed that tetracaine hydrochloride is extremely sensitive to oxidation condition and highly sensitive to alkaline/acidic hydrolysis, and susceptible to light condition. In total, five related impurities and seven degradation products were successfully detected in the positive mode of electrospray ionization. The structures of all these impurities were characterized based on the high-resolution MS data and manufacture process, and the fragmentation pathways of tetracaine and these impurities were constructed and discussed. Seven of them have not been reported before, and two of them were specified impurities described in various pharmacopoeias. The fragmentation pathways and plausible mechanisms for the formation of these impurities were proposed.

Monoacylglycerol lipase from marine Geobacillus sp. showing lysophospholipase activity and its application in efficient soybean oil degumming

Enzymatic degumming is an essential refining process to improve oil quality. In this study, a monoacylglycerol lipase GMGL was derived from marine Geobacillus sp., and was found that not only took monoacylglycerol (MAG) as substrate, but also had activity toward lysophosphatidylcholine (LPC), lysophosphatidylethanolamine (LPE) and glycerolphosphatidylcholine (GPC). Binding free energy showed LPC and LPE could bind with enzyme stably as MAG. It presented great potential in the field of enzymatic degumming. The phosphorus content in crude soybean oil decreased from 680.50 to 2.01 mg/kg and the yield of oil reached to 98.80 % after treating with phospholipase A1 (Lecitase Ultra) combined with lipase GMGL. An ultrahigh-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) was developed to identify 21 differential phospholipids between crude soybean oil and enzymatic treatment. This work might shed some light on understanding the catalytic mechanism of monoacylglycerol lipase and provide an effective strategy for enzymatic degumming.

Multiclass target analysis of contaminants of emerging concern including transformation products, soil bioavailability assessment and retrospective screening as tools to evaluate risks associated with reclaimed water reuse

The occurrence of 200 multiclass contaminants of emerging concern (CECs) encompassing 168 medicinal products and transformation products (TPs), 5 artificial sweeteners, 12 industrial chemicals, and 15 other compounds was investigated in influent and effluent wastewater samples collected during 7 consecutive days from 5 wastewater treatment plants (WWTPs) located in Cyprus. The methodology included a generic solid-phase extraction protocol using mixed-bed cartridges followed by Ultra-High Performance Liquid Chromatography coupled with Quadrupole-Time of Flight Mass Spectrometry (UHPLC-QTOF-MS) analysis. A total of 63 CECs were detected at least in one sample, with 52 and 55 out of the 200 compounds detected in influents and effluents, respectively. Ten (10) out of the 24 families of parent compounds and associated TPs were found in the wastewater samples (influent or effluent).1-H-benzotriazole, carbamazepine, citalopram, lamotrigine, sucralose, tramadol, and venlafaxine (>80 % frequency of appearance in effluents) were assessed with respect to their bioavailability in soil as part of different scenarios of irrigation with reclaimed water following a qualitative approach. A high score of 12 (high probability) was predicted for 2 scenarios, a low score of 3 (rare occasions) for 2 scenarios, while the rest 28 scenarios had scores 5–8 (unlikely or limited possibility) and 9–11 (possibly).Retrospective screening was performed with the use of a target database of 2466 compounds and led to the detection of 158 additional compounds (medicinal products (65), medicinal products TPs (15), illicit drugs (7), illicit drugs TPs (3), industrial chemicals (11), plant protection products (25), plant protection products TPs (10), and various other compounds (22).This work aspires to showcase how the presence of CECs in wastewater could be investigated and assessed at WWTP level, including an expert-based methodology for assessing the soil bioavailability of CECs, with the aim to develop sustainable practices and enhance reclaimed water reuse.

Study on the mechanism of anti-hepatic fibrosis of Glycyrrhiza Uralensis-Salvia miltiorrhiza prescription based on serum and urine metabolomics and network pharmacology

Hepatic fibrosis (HF) is a kind of chronic epidemic liver disease. Glycyrrhiza Uralensis and Salvia Miltiorrhiza (GUSM), traditional Chinese medicine, has the obvious clinical treatment of liver fibrosis. This study aimed to investigate the mechanisms of GUSM against HF by an integrated strategy combining untargeted metabolomics with network pharmacology. The results showed that GUSM prescription can improve the morphology and structure of liver tissue, inhibit the proliferation of collagen fibers and reducing the inflammatory response of the liver and so on. Endogenous metabolites and HF-related potential biomarkers in serum and urine were detected by ultra-high-performance liquid chromatography coupled with quadrupole-time-of-flight mass spectrometry (UPLC-Q-TOF/MS). The metabolic pathways were enriched by MetaboAnalyst. GUSM prescription showed an antifibrotic effect on rats by regulating metabolic pathways, mainly pentose and glucuronate interconversions and arachidonic acid metabolism. Network pharmacology was then applied to find 42 overlapping targets of GUSM-HF. Quercetin was found to be the main active component and STAT3 was the main active target in GUSM prescription. Molecular docking showed high affinities between quercetin and STAT3. Therefore, GUSM has protective effects on HF by regulating the metabolism and different signaling pathways. The work also shows that the metabolomic and network pharmacology methods are promising tools to gain insight into the efficacy and mechanism research of traditional Chinese medicines.

Identification of biomarkers and the mechanisms of multiple trauma complicated with sepsis using metabolomics.

Sepsis after trauma increases the risk of mortality rate for patients in intensive care unit (ICUs). Currently, it is difficult to predict outcomes in individual patients with sepsis due to the complexity of causative pathogens and the lack of specific treatment. This study aimed to identify metabolomic biomarkers in patients with multiple trauma and those with multiple trauma accompanied with sepsis. Therefore, the metabolic profiles of healthy persons designated as normal controls (NC), multiple trauma patients (MT), and multiple trauma complicated with sepsis (MTS) (30 cases in each group) were analyzed with ultra-high performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF/MS)-based untargeted plasma metabolomics using collected plasma samples. The differential metabolites were enriched in amino acid metabolism, lipid metabolism, glycometabolism and nucleotide metabolism. Then, nine potential biomarkers, namely, acrylic acid, 5-amino-3-oxohexanoate, 3b-hydroxy-5-cholenoic acid, cytidine, succinic acid semialdehyde, PE [P-18:1(9Z)/16:1(9Z)], sphinganine, uracil, and uridine, were found to be correlated with clinical variables and validated using receiver operating characteristic (ROC) curves. Finally, the three potential biomarkers succinic acid semialdehyde, uracil and uridine were validated and can be applied in the clinical diagnosis of multiple traumas complicated with sepsis.

Anti-Inflammatory Effects and Molecular Mechanisms of Shenmai Injection in Treating Acute Pancreatitis: Network Pharmacology Analysis and Experimental Verification.

BackgroundAcute pancreatitis (AP) is an inflammatory disorder of the exocrine pancreas without specific treatment. Shenmai injection (SMI) was reported to eliminate the severity of experimental AP. This study aimed to explore the mechanisms underlying the synergistic protective effects of SMI on AP based on network pharmacology and experimental validation.MethodsNetwork pharmacology analysis and molecular docking based on identified components were performed to construct the potential therapeutic targets and pathways. The principal components of SMI were detected via ultra-high-performance liquid chromatography-coupled with quadrupole time-of-flight mass spectrometry (UHPLC-QTOF/MS). Effect of SMI and the identified components on cellular injury and IL6/STAT3 signaling was assessed on mouse pancreatic acinar cell line 266–6 cells. Finally, 4% sodium taurocholate (NaT) was used to induce AP model to assess the effects of SMI in treating AP and validate the potential molecular mechanisms.ResultsBy searching the TCMSP and ETCM databases, 119 candidate components of SMI were obtained. UHPLC-QTOF/MS analysis successfully determined the representative components of SMI: ginsenoside Rb1, ginsenoside Rg1, ginsenoside Re, and ophiopogonin D. Fifteen hub targets and eight related pathways were obtained to establish the main pharmacology network. Subnetwork analysis and molecular docking indicated that the effects of these four main SMI components were mostly related to the interleukin (IL) 6/STAT3 pathway. In vitro, SMI, ginsenoside Rb1, ginsenoside Rg1, ginsenoside Re, and ophiopogonin D increased the cell viability of NaT-stimulated mouse pancreatic acinar 266–6 cells and decreased IL6 and STAT3 expression. In vivo, 10 mL/kg SMI significantly alleviated the pancreatic histopathological changes and the expression of IL6 and STAT3 in the AP mice.ConclusionThis study demonstrated SMI may exert anti-inflammatory effects against AP by suppressing IL6/STAT3 activation, thus providing a basis for its potential use in clinical practice and further study in treating AP.

Main components in different varieties of Xihuangcao by UPLC-Q-TOF-MS and UPLC-DAD

This study established an ultra-high performance liquid chromatography coupled with quadrupole-time-of-flight mass spectrometry(UPLC-Q-TOF-MS) method to analyze the main components in different varieties of Xihuangcao and established a UPLC-DAD method to simultaneously determine the five active components(caffeic acid, rosmarinic acid, schaftoside, isoschaftoside, and oridonin).The chromatographic separation was performed on a Waters ACQUITY UPLC BEH C_(18) column(2.1 mm×100 mm, 1.7 μm) with a gradient elution of methanol(B)-water containing 0.1% formic acid(A) at a flow rate of 0.3 mL·min~(-1).The column temperature was 30 ℃.The Q-TOF-MS discriminant analysis was performed under positive electrospray ion mode and the split ratio was 1∶1. Quantitative analysis was carried out by UPLC-DAD.The determination wavelength was set at 245 nm.Thirty-two main components of Xihuangcao were separated and identified by UPLC-Q-TOF-MS, where 19 were identified in Rabdosia serra, nine in R.nervosa, 10 in R.lophanthoides, 15 in R.lophanthoides var.graciliflora, 10 in R.lophanthoides var.gerardianus, and seven in R.stracheyi.The UPLC-DVD method was developed for simultaneously determining five active components in different varieties of Xihuangcao.The standard curves for five compounds showed good linearity with correlation coefficients higher than 0.999 0.The precision, repeatability, and stability were good.The average recoveries(n=6) were between 97.01% and 102.7% with RSD<3.0%.The results of UPLC-Q-TOF-MS analysis provided a scientific basis for the use of R.stracheyi as a medicinal material of Xihuangcao and the equivalent use of R.lophanthoides var.gerardianus with R.lophanthoides var.graciliflora to some extent.The UPLC-DAD method for simultaneously determining five active components is simple, rapid, and accurate.This study can provide the basis for the quality control of different varieties of Xihuangcao.

Metabolomics analysis reveals the renal protective effect of Panax ginseng C. A. Mey in type 1 diabetic rats.

The dry root and rhizome of Panax ginseng C. A. Mey has garnered much interest owing to its medicinal properties against diabetes and cardiovascular diseases. In this study, an ultra-high performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS)-based metabolomics approach was used to illustrate the therapeutic mechanisms of ginseng extract on the serum and urinary metabolic profiles in streptozotocin-induced type 1 diabetes mellitus (T1DM) rats. Pharmacological and renal parameters in response to the administration of ginseng were also evaluated. In total, 16 serum endogenous metabolites and 14 urine endogenous metabolites, including pyruvic acid, indoleacetic acid, and phenylacetylglycine, were identified as potential biomarkers for diabetes. Pathway enrichment and network analysis revealed that the biomarkers modulated by ginseng were primarily involved in phenylalanine and pyruvate metabolism, as well as in arginine biosynthesis. Moreover, the levels of several renal injury-related biomarkers in T1DM rats were significantly restored following treatment with ginseng. The administration of the extract helped maintain tissue structure integrity and ameliorated renal injury. The findings suggest that the regulatory effect of ginseng extract on T1DM involves metabolic management of diabetic rats, which subsequently attenuates T1DM-induced early renal dysfunction.

A Comprehensive Strategy Based on UPLC-Q/TOF-MS for the Identification of Compounds in a Chinese Patent Medicine, Xiao'er Chiqiao Qingre Granules.

The aim of this study was to establish a comprehensive strategy based on liquid chromatography coupled with mass spectrometry to potently identify as many compounds of Chinese patent medicine as possible. Ultrahigh performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q/TOF-MS) was used to qualitatively analyze the Chinese patent medicine Xiao'er Chiqiao Qingre Granules (XCQG), which is recorded in the Chinese Pharmacopoeia. A novel strategy, including targeted, semi-targeted and non-targeted identification, was built to explore the compounds based on accurate mass, characteristic fragments, retention time of standard substances, databases or literature. Based on the integrated identification, 250 compounds were identified in total, including 7 alcohols, 3 aldehydes, 17 alkaloids, 9 amino acids, 10 coumarins, 30 flavonoids, 29 glycosides, 12 ketones, 7 lignans, 20 organic acids, 12 phenols, 11 phenylpropanoids, 9 quinones, 3 steroids, 26 terpenes, 14 volatile oils and 31 other compounds. A novel strategy for the identification of compounds in traditional Chinese medicine (TCM) was developed with Ultrahigh performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q/TOF-MS). It is also the first systematic study of compounds in XCQG, laying a foundation for further mechanism research of XCQG. More importantly, the strategy shows good application prospect in identifying compounds of TCM.

Effect of traditional chinese medicine (TCM) and its fermentation using Lactobacillus plantarum on ceftriaxone sodium-induced dysbacteriotic diarrhea in mice

BackgroundBuzhongyiqi decoction (BD), Sijunzi decoction (SD), and Shenlingbaizhu decoction (SHD) have been extensively used clinically for the treatment of diseases caused by spleen-Qi deficiency and microbial fermentation has historically been utilized in traditional Chinese medicine (TCM). This study aimed to investigate the mitigative effect of TCM and fermented TCM (FTCM) with Lactobacillus plantarum (LP) on antibiotic-associated diarrhea, and to select an optimal formula and then identify its compounds.MethodsDysbacteriosis in mice was induced by ceftriaxone sodium (CS). The mice were then treated with LP, BD, SD, SHD, fermented BD, fermented SD (FSD), and fermented SHD. Diarrhea indexes, the abundances of gut bacteria, intestinal morphometrics, and mRNA expressions of genes related to intestinal barrier function were assessed. Then, ultra-high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF/MS) were employed to identify and relatively quantify the compounds in the selected decoctions.ResultsCS significantly increased the fecal output weight, the total number of fecal output, and fecal water content, indicating the occurrence of diarrhea. Bacterial culture tests showed that the above symptoms were accompanied by the disruption of specific intestinal flora. TCM, LP, and FTCM alleviated the diarrhea index and recovered the intestinal microbiota. FTCM showed more advantageous than TCM or LP alone. The mRNA expressions of aquaporins (AQPs) and tight junctions (TJs) decreased by CS were enhanced by TCM, LP, and FTCM. In addition, through UHPLC-Q-TOF/MS, (S)-(-)-2-hydroxyisocaproic acid, L-methionine, 4-guanidinobutyric acid (4GBA), and phenyllactate (PLA) in SD and FSD were identified and relatively quantified.ConclusionsTCM, LP, and TCM fermented with LP alleviated CS-induced diarrhea symptoms, and improved the intestinal flora and barrier function. Four compounds including (S)-(-)-2-hydroxyisocaproic acid, L-methionine, 4GBA, and PLA in FSD, which were identified by UHPLC-Q-TOF/MS, might function in modulating intestinal flora and improving villi structure.

Schisandrol A, the main active ingredient of Schisandrae Chinensis Fructus, inhibits pulmonary fibrosis through suppression of the TGF-β signaling pathway as revealed by UPLC-Q-TOF/MS, network pharmacology and experimental verification

Ethnopharmacological relevanceSchisandra chinensis decoction derived from the book of Waitai Miyao (Tao Wang, Tang dynasty) is often used in the treatment of idiopathic pulmonary fibrosis (IPF), which is included in the Grand Ceremony of Chinese formulae (Huairen Peng, 1994). Schisandrae Chinensis Fructus (Sch) is one of the most important herbs in this formula. According to the “Shennong's Herbal Classicherbal” of the Han Dynasty, Sch has sour taste, warm nature, which has the effect of tonifying qi and curing cough. In addition, according to the “Compendium of Materia Medica” of the Ming Dynasty, Sch is used to treat cough and asthma, which has the effect of moistening the lung and tonifying the kidney. However, the active ingredients of Sch absorption into the plasma and its pharmacological mechanism of treatment for IPF still remained unclear. Aim of the studyOur research aimed at identifying the absorbed active ingredients and metabolized of Sch in rat plasma and the mechanism of anti-IPF based on serum pharmacochemistry. Materials and methodsFirst, the rats were divided into control group and Sch group. Sch sample was orally administrated to the rats for seven days. The blood samples were drawn into an Eppendorf tube after the last dosing. The ultrahigh performance liquid chromatography coupled with quadrupole-time of flight mass spectrometry (UPLC-Q-TOF/MS) was applied to identify the absorption components and metabolites of Sch in rat plasma. Second, the network pharmacology combined with molecular docking analysis was further investigated to illuminate its potential mechanism of treatment for IPF by the biological targets regulating related pathways. Finally, the mechanism of action was verified by experimental in vitro and in vivo. ResultsA total of 78 compounds, consist of 13 prototype lignans and 65 metabolites (including isomers) were identified. Network pharmacology study and molecular docking analysis indicated that schisandrol A (L1) play an anti-fibrosis role by regulating the TGF-β signaling pathway. Experimental in vitro and in vivo verified that the schisandrol A could inhibiting pulmonary fibrosis through TGF-β signaling pathway. The effect and mechanism of schisandrol A inhibiting pulmonary fibrosis were reported for the first time. ConclusionsIn this study, the absorption active ingredients of Sch in rat plasma were combined with the network pharmacology investigation and experimental in vitro and in vivo to elucidate its biological mechanism of treatment for IPF. The results provided a theoretical support for understanding the bioactive compounds and the pharmacological mechanism of Sch.

Structural Characterization and Discrimination of Morinda officinalis and Processing Morinda officinalis Based on Metabolite Profiling Analysis.

Morindae officinalis Radix (MOR) is a famous traditional Chinese medicine (TCM) and functional food material for invigorating kidneys and tonifying yang. Processed Morindae officinalis Radix (PMOR) is obtained by steaming MOR. Traditionally, the clinical effects are discrepant between processing and nonprocessing herbal medicines. MOR and PMOR are commonly used in both clinical practice and dietary supplements, and the effect of invigorating kidneys and tonifying yang of PMOR is stronger than MOR. To clarify the overall chemical composition and the difference of MOR and PMOR, a method was developed with an ultrahigh-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF/MS). Among the 110 identified components shared by MOR and PMOR, 55 compounds showed significant differences in contents. Among them, the contents of 29 components, including fructooligosaccharides, monotropein, deacetylasperulosidic acid, geniposide, and anthraquinone glycosides, were higher in MOR than in PMOR; the contents of 26 components, including difructose anhydride sucrose, and iridoid glycoside derivatives, were higher in PMOR than in MOR. Difructose anhydrides and iridoid glycoside derivatives were first discovered in PMOR. These results provided a scientific basis for research on the therapeutic material basis of MOR. It would provide a method for the comparison of processing and nonprocessing in Chinese medicines.

Metabolomics analysis revealed acupuncture treatment target the upstream for dry eye disease

AIM: To find out the pathogenesis of dry eye disease (DED) and the possible mechanisms of available effective treatments. METHODS: A non-targeted technology, ultra-high performance liquid chromatography coupled with quadrupole-time-of-flight mass spectrometry (UHPLC-Q-TOF-MS), to investigate metabolic characterizations for tear samples from 18 patients with DED upon drug or acupuncture treatment. RESULTS: A total of 190 named metabolites were identified, which presented so far the broadest tear metabolome. Further analysis indicated a significantly distinct metabolomics profile among all patients, but very subtle metabolic differences upon drug or acupuncture treatment. On one hand, only six significantly changed metabolites were determined after drug treatment, five of which including inosine, monopalmitin, urate, propionylcarnitine, and nicotinamide were all increased and involved in inflammatory responses. On the other hand, merely four metabolites including alanine, serine, and homoserine were found to be significantly different. Further pathway analysis of those six and four significantly changed metabolites revealed that only one pathway, aminoacyl-tRNA biosynthesis was significantly influenced in acupuncture-treated patients, which were highly associated with the cause of the disease. The results here indicated acupuncture treatment may address the cause rather than the symptoms for dry eye disease, displaying partially better compared with drug treatment. CONCLUSION: Collectively, this work extended our understanding on the key regulatory elements or pathways involved in the potential mechanisms of available effective treatments, and would be useful for providing novel potential targets and therapeutic strategies for DED.

Fast Screening and Identification of Illegal Adulterated Glucocorticoids in Dietary Supplements and Herbal Products Using UHPLC-QTOF-MS With All-Ion Fragmentation Acquisition Combined With Characteristic Fragment Ion List Classification.

Ultra-high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UHPLC-QTOF-MS) with all-ion fragmentation (AIF) acquisition was established for an identification and quantification of illegal adulterated glucocorticoids in dietary supplements and herbal products. Next, a novel method called characteristic fragment ion list classification (CFILC) was developed for a fast screening of adulterated compounds. CFILC could provide the characteristic ions comprehensively and completely through direct extract from the MS2 library instead of finding them manually. This is time-saving and provides fast screening results with a high confidence level by filtering of a pre-calculated threshold of similarity scores for illegal adulterants that are not included in the library as well as for new emerging structural analogs. The obtained results demonstrated the great qualitative and quantitative strength of this approach, providing a promising and powerful method for a routine fast screening of illegal adulterated glucocorticoids.

Analysis of verteporfin liposomal formulations for phospholipids and phospholipid degradation products by liquid chromatography-mass spectrometry (LC-MS)

Lipid composition and lipid degradation are critical to the stability of liposomal formulations which can impact the safety and efficacy of the drug. Herein we developed and validated an ultrahigh performance liquid chromatography coupled with quadrupole time of flight mass spectrometry (UPLC-QTOF-MS) method for determining phospholipid composition and phospholipid degradation products in a verteporfin liposomal formulation (Visudyne). The high mass accuracy (<5 ppm) of the QTOF method coupled with database searching (SimLipid) and comparison with known standards accurately identified and quantified the phospholipid compositions and lipid degradation products. The analysis of Visudyne indicated that more than 50% (w/w) of the total phospholipids are composed of phosphatidylcholine (PC) 14:0–14:0 and major phosphatidylglycerol (PG) species found are PG 16:0–18:2, PG 16:0–18:1, PG 18:0–18:2, and PG 18:0–18:1. The LC-MS method developed is capable of separating structural isomers such as PG 18:1–18:1 versus PG 18:0–18:2 and the separation of PG stereoisomers, such as PG 18:1–18:1 cis and PG 18:1–18:1 trans. The major lipid degradation products in Visudyne includes lysophosphatidylcholine and a few saturated and unsaturated lysophosphatidylglycerols, and free fatty acids (FFA). Each degradation product is less than 1% of the total phospholipids (w/w). In addition, the lipid profiles of naturally sourced egg PG from six different vendors were compared with the PG composition in Visudyne. Differences in lipid composition in egg PGs from different vendors were observed and the PG composition in Visudyne is matched with the lipid profile of the some of the egg PGs from different vendors. Drug developers can utilize this method to assess raw materials and lipid-based drug product quality and regulatory scientists can monitor the quality of the drug available in the market using this validated method.

Protective mechanisms of Agrimonia pilosa Ledeb in dextran sodium sulfate-induced colitis as determined by a network pharmacology approach.

Previous studies reported that Agrimonia pilosa (AP) Ledeb possessed diverse biological activities, including anti-inflammatory, antioxidant, and anti-tumor activities. However, the effect of AP on ulcerative colitis (UC) remains unclear. In this study, we investigated the therapeutic effect and mechanisms of AP on dextran sodium sulfate (DSS)-induced colitis. The potential constituents of AP were investigated by ultra-high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS). A total of 13 compounds were recognized by UPLC-Q-TOF/MS chromatogram. Furthermore, a network pharmacology approach revealed that there are 297 candidate targets of UC and 549 common targets for the 13 active ingredients of AP. GO enrichment and KEGG pathway analysis indicated that AP might have a protective effect on UC through the nuclear factor κB (NF-κB) and nucleotide-binding oligomerization domain (NOD)-like receptor signaling pathways. Subsequent experimental validation in a DSS-induced colitis model revealed that AP alleviated the severity of DSS-induced colitis, reduced the production of proinflammatory factors, and protected against the loss of intestinal integrity. Moreover, AP inhibited the phosphorylation of NF-κB p65 and the activation of the NLRP3 inflammasome. In conclusion, AP ameliorated DSS-induced colitis through suppressing the activation of the NLRP3 inflammasome and NF-κB signaling pathways.