Quality evaluation of Lonicerae Japonicae Flos and Lonicerae Flos based on simultaneous determination of multiple bioactive constituents combined with multivariate statistical analysis.

INTRODUCTION: Lonicerae Japonicae Flos (LJF) and Lonicerae Flos (LF) belong to different genera of Caprifoliaceae. They have been historically utilised as herbal medicine to treat various diseases. However, the comprehensive assessment of them still remains a challenge. OBJECTIVE: To develop a comprehensive method of ultra-fast liquid chromatography-tandem triple quadrupole mass spectrometry (UFLC-QTRAP-MS/MS) coupled with multivariate statistical analysis for the quality evaluation and reveal differential components of LJF and LF. METHODOLOGY: A validated UFLC-QTRAP-MS/MS method was established for simultaneous determination of 50 constituents, including 12 organic acids, 12 flavonoids, 6 iridoids, 3 saponins, 13 amino acids and 4 nucleosides. The obtained data were employed to multivariate statistical analysis. Principal component anlysis (PCA) and partial least squares determinant analysis (PLS-DA) were performed to classify and reveal differential components of samples; grey relational analysis (GRA) was introduced to assess the samples according to the contents of 50 constituents by calculating the relative correlation degree of each sample. RESULTS: Fifty constituents were simultaneously determined of LJF and LF. Based on obtained data, PCA and PLS-DA were easy to distinguish samples and the classification of the samples was related to 11 chemical constituents. GRA implied the quality of LJF was better, and that the flower buds were superior to the flowers. Moreover, organic acids are the main components of samples. CONCLUSION: This study not only established a method of simultaneous determination of multiple bioactive constituents in LJF and LF, but provided comprehensive information on the quality control of them. The developed method is conducive to distinguish orthologues or paralogues of them, and supply the support for "heterologous effects".

Simultaneous quantitation of glycyrrhetic acid and puerarin in plasma using ultra flow liquid chromatography tandem mass spectrometry and application in a pharmacokinetics study in healthy and alcoholic liver injury rats.

A rapid, sensitive, and accurate ultra flow liquid chromatography tandem mass spectrometry (UFLC-MS/MS ) method was developed and validated for simultaneous quantitation of glycyrrhetic acid and puerarin in plasma derived from healthy and alcoholic liver injury rats. Plasma samples from healthy and model rats were deproteinated with methanol using liquiritin as an internal standard. Chromatography separation was performed by a Waters BEH (ethylene-bridged hybrid) C18 column (2.1 × 50 mm; 1.7 µm) using a gradient elution from acetonitrile and water (containing 0.1% formic acid) and at a flow rate of 0.4 mL/min. Quantitation was performed on a Triple Quad 4500 tandem mass spectrometer coupled with an electrospray ionization source in negative multiple reaction monitoring mode. Specificity, carryover, dilution integrity, recovery, linearity, precision and accuracy, matrix effect, and stability were within acceptable limits. The newly established method was successfully applied to a pharmacokinetics study to investigate glycyrrhetic acid and puerarin in healthy and alcoholic liver injury rats.

Distribution Patterns for Bioactive Constituents in Pericarp, Stalk and Seed of Forsythiae Fructus.

Forsythiae Fructus (FF) is a widely used folk medicine in China, Japan, and Korea. The distribution of bioactive constituents throughout the fruit segments has rarely been addressed, although mounting evidence suggests that plant secondary metabolites are synthesized and distributed regularly. The phytochemical profiles of three segments of FF (pericarp, stalk and seed) were firstly revealed by liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based quantitative analysis of twenty-one bioactive constituents, including three phenylethanoid glycosides, five lignans, eight flavonoids, and five phenolic acids to explore the spatial distribution of bioactive constituents. Furthermore, the hierarchical clustering analysis (HCA) and one-way analysis of variance (one-way ANOVA) were conducted to visualize and verify the distribution regularity of twenty-one analytes among three segments. The results showed that phytochemical profiles of the three segments were similar, i.e., phenylethanoid glycosides covering the most part were the predominant compounds, followed by lignans, flavonoids and phenolic acids. Nevertheless, the abundance of twenty-one bioactive constituents among three segments was different. Specifically, phenylethanoid glycosides were highly expressed in the seed; lignans were primarily enriched in the stalk; flavonoids were largely concentrated in the pericarp, while the contents of phenolic acids showed no much difference among various segments. The research improves our understanding of distribution patterns for bioactive constituents in FF, and also complements some scientific data for further exploring the quality formation mechanism of FF.

Beta-elemene inhibits breast cancer metastasis through blocking pyruvate kinase M2 dimerization and nuclear translocation.

Pyruvate kinase M2 (PKM2), playing a central role in regulating aerobic glycolysis, was considered as a promising target for cancer therapy. However, its role in cancer metastasis is rarely known. Here, we found a tight relationship between PKM2 and breast cancer metastasis, demonstrated by the findings that beta-elemene (ß-elemene), an approved drug for complementary cancer therapy, exerted distinct anti-metastatic activity dependent on PKM2. The results indicated that ß-elemene inhibited breast cancer cell migration, invasion in vitro as well as metastases in vivo. ß-Elemene further inhibited the process of aerobic glycolysis and decreased the utilization of glucose and the production of pyruvate and lactate through suppressing pyruvate kinase activity by modulating the transformation of dimeric and tetrameric forms of PKM2. Further analysis revealed that ß-elemene suppressed aerobic glycolysis by blocking PKM2 nuclear translocation and the expression of EGFR, GLUT1 and LDHA by influencing the expression of importin α5. Furthermore, the effect of ß-elemene on migration, invasion, PKM2 transformation, and nuclear translocation could be reversed in part by fructose-1,6-bisphosphate (FBP) and L-cysteine. Taken together, tetrameric transformation and nuclear translocation of PKM2 are essential for cancer metastasis, and ß-elemene inhibited breast cancer metastasis via blocking aerobic glycolysis mediated by dimeric PKM2 transformation and nuclear translocation, being a promising anti-metastatic agent from natural compounds.

Synthesis and biological evaluation of a novel series of curcumin-peptide derivatives as PepT1-mediated transport drugs.

Curcumin (CUR) is a natural yellow pigment from turmeric with extensive bioactivities. However its relatively poor solubility limited its absorption and bioavailability. In this study, a novel series of CUR-peptide conjugates were designed and synthesized as PepT1-mediated transport drugs and their solubility, cellular uptakes and anti-tumor activities were evaluated. Ten compounds showed better water solubility than CUR due to the dipeptide moiety. Compared with CUR, compound 5e exhibited the slightly better activity and 5d showed the similar activity with CUR. Besides, compounds 5d and 5e performed higher cellular uptakes in Caco-2 cell and dose-dependently inhibited by the addition of PepT1 typical substrate glycylsarcosine (Gly-Sar). Compound 5d and 5e have improved the absorption of CUR by PepT1-mediated without affected the activity. These new dipeptide conjugates of CUR may serve as promising lead compounds for future drug development.

Simultaneous Characterization and Quantification of Varied Ingredients from Sojae semen praeparatum in Fermentation Using UFLC⁻TripleTOF MS.

Systematic comparison of active ingredients in Sojae semen praeparatum (SSP) during fermentation was performed using ultra-fast liquid chromatography (UFLC)-TripleTOF MS and principal component analysis (PCA). By using this strategy, a total of 25 varied compounds from various biosynthetic groups were assigned and relatively quantified in the positive or negative ion mode, including two oligosaccharides, twelve isoflavones, eight fatty acids, N-(3-Indolylacetyl)-dl-aspartic acid, methylarginine, and sorbitol. Additionally, as the representative constituents, six targeted isoflavones were sought in a targeted manner and accurately quantified using extracted ion chromatograms (XIC) manager (AB SCIEX, Los Angeles, CA, USA) combined with MultiQuant software (AB SCIEX, Los Angeles, CA, USA). During the fermentation process, the relative contents of oligoses decreased gradually, while the fatty acids increased. Furthermore, the accurate contents of isoflavone glycosides decreased, while aglycones increased and reached a maximum in eight days, which indicated that the ingredients converted obviously and regularly throughout the SSP fermentation. In combination with the morphological changes, which meet the requirements of China Pharmacopoeia, this work suggested that eight days is the optimal time for fermentation of SSP from the aspects of morphology and content.

A Method to Study the Distribution Patterns for Metabolites in Xylem and Phloem of Spatholobi Caulis.

Spatholobi Caulis (SC), the vine stem of Spatholobus suberectus Dunn, is a widely used traditional Chinese medicine (TCM) for the treatment of blood stasis syndrome and related diseases. Xylem and phloem are the main structures of SC and the color of xylem in SC is red brown or brown while the phloem with resin secretions is reddish brown to dark brown. They are alternately arranged in a plurality of concentric or eccentric rings. In order to investigate the distribution patterns of metabolites in xylem and phloem of SC, an analytical method based on UFLC-QTRAP-MS/MS was established for simultaneous determination of 22 constituents including four flavanols, nine isoflavones, two flavonols, two dihydroflavones, one flavanonol, one chalcone, one pterocarpan, one anthocyanidin and one phenolic acid in the samples (xylem and phloem) from Laos. Furthermore, according to the contents of 22 constituents, heat map, principal components analysis (PCA), orthogonal partial least squares discriminant analysis (OPLS-DA) and t-test were used to evaluate the samples and discover the differences between xylem and phloem of SC. The results indicated that the measured ingredients in xylem and phloem were significantly different. To be specific, the contents of flavonoids in xylem were higher than that in phloem, while the content of protocatechuic acid showed a contrary tendency. This study will not only reveal the distribution patterns of metabolites in xylem and phloem of SC but also facilitate further study on their quality formation.

Quality evaluation of Eucommiae Cortex processed by different methods and "sweating" conditions based on simultaneous determination of multiple bioactive constituents combined with gray relational analysis.

Eucommiae Cortex is a classical traditional Chinese medicine, which needs to be processed by "sweating" methods. To select the suitable processing method and "sweating" processing condition for Eucommiae Cortex, in this study, the quality of Eucommiae Cortex was evaluated based on simultaneous determination of multiple bioactive constituents combined with gray relational analysis. The contents of lignans, iridoids, penylpropanoids, flavonoids, and phenols in samples were simultaneously determined using ultra-fast performance liquid chromatography coupled with triple quadrupole-linear ion trap tandem mass spectrometry. The chromatographic separation was performed on a Synergiۛ Hydro-RP 100 Å column (100 mm × 2.0 mm, 2.5 µm) at 30°C with a gradient elution of acetonitrile with 0.1% formic acid/0.1% aqueous formic acid as the mobile phase. Furthermore, gray relational analysis was performed to evaluate and sort the samples according to the contents of 14 constituents by calculating the relative correlation degree of each sample. The results demonstrated that the quality of Eucommiae Cortex "sweating" at source area was better and the better "sweating" condition was to scrape off the cork layer before "sweating" with straw covering and sun drying. The developed method could provide the foundation and support for "sweating" processing method of Eucommiae Cortex in normalization and standardization.

Comparison of Multiple Bioactive Constituents in Different Parts of Eucommia ulmoides Based on UFLC-QTRAP-MS/MS Combined with PCA.

Eucommia ulmoides Oilv. (EU), also called Du-zhong, is a classical traditional Chinese medicine. Its bark, leaf, and male flower are all used for medicinal purposes, called Eucommiae Cortex (EC), Eucommiae Folium (EF), and Eucommiae Flos Male (EFM). In order to study the difference in synthesis and the accumulation of metabolites in different parts of EU, a reliable method based on ultra-fast liquid chromatography tandem triple quadrupole mass spectrometry (UFLC-QTRAP-MS/MS) was developed for the simultaneous determination of a total of 21 constituents, including two lignans, 6 iridoids, 6 penylpropanoids, 6 flavonoids, and one phenol in the samples (EC, EF, and EFM). Furthermore, principal component analysis (PCA) was performed to evaluate and classify the samples according to the contents of these 21 constituents. All of the results demonstrated that the chemical compositions in EC, EF, and EFM were significantly different and the differential constituents (i.e., aucubin, geniposidic acid, chlorogenic acid, pinoresinol-di-O-ß-d-glucopyranoside, geniposide, cryptochlorogenic acid, rutin, and quercetin) were remarkably associated with sample classifications. The research will provide the basic information for revealing the laws of metabolite accumulation in EC, EF, and EFM from the same origin.

Quality Evaluation of Apocyni Veneti Folium from Different Habitats and Commercial Herbs Based on Simultaneous Determination of Multiple Bioactive Constituents Combined with Multivariate Statistical Analysis.

Apocyni Veneti Folium (AVF) is a kind of staple traditional Chinese medicine with vast clinical consumption because of its positive effects. However, due to the habitats and adulterants, its quality is uneven. To control the quality of this medicinal herb, in this study, the quality of AVF was evaluated based on simultaneous determination of multiple bioactive constituents combined with multivariate statistical analysis. A reliable method based on ultra-fast liquid chromatography tandem triple quadrupole mass spectrometry (UFLC-QTRAP-MS/MS) was developed for the simultaneous determination of a total of 43 constituents, including 15 flavonoids, 6 organic acids, 13 amino acids, and 9 nucleosides in 41 Luobumaye samples from different habitats and commercial herbs. Furthermore, according to the contents of these 43 constituents, principal component analysis (PCA) was employed to classify and distinguish between AVF and its adulterants, leaves of Poacynum hendersonii (PHF), and gray relational analysis (GRA) was performed to evaluate the quality of the samples. The proposed method was successfully applied to the comprehensive quality evaluation of AVF, and all results demonstrated that the quality of AVF was higher than the PHF. This study will provide comprehensive information necessary for the quality control of AVF.

Contents variation analysis of free amino acids, nucleosides and nucleobases in semen sojae praeparatum fermentation using UFLC-QTRAP MS.

UFLC-QTRAP MS was used to develop a sensitive and rapid method of evaluating content variation during Semen sojae praeparatum (SSP) fermentation. It did this through the simultaneous quantification of 22 free amino acids and 16 nucleosides and nucleobases in the raw materials and processed products of SSP. The method was shown to be reproducible and accurate. The limits of detection and quantity values were 0.09-168.75 and 0.31-562.50 ng/mL for the 38 analytes, respectively. The data were examined through principal components analysis to compare the content variations. The quantitative results showed that the ingredients were properly determined in most of the samples and were converted regularly throughout the SSP fermentation process. These results correspond to the morphologic changes and principal components analysis results.

Determination of a natural DNMT1 inhibitor, peperomin E, in rat plasma by UFLC-MS/MS and method application in a pharmacokinetic study.

Peperomin E (PepE), a naturally occurring secolignan isolated from Peperomia dindygulensis, has drawn much attention recently owing to its anticancer and DNA methyltransferase 1 (DNMT1) inhibitory activity. Here, a simple and sensitive ultra-fast liquid chromatography-tandem mass spectrometry method was developed and validated for the determination of PepE in rat plasma for the first time. Samples were prepared by simple protein precipitation. Separation was performed on an XBridge™ C18 column using a mobile phase of acetonitrile and 0.1% (v/v) aqueous formic acid. PepE and the internal standard arctigenin were detected in a positive-ion mode using multiple reaction monitoring of the transitions at m/z 413.2 → 261.0 and 373.2 → 137.2, respectively. The calibration curve for PepE was linear over the range of concentrations of 1.46-6000 ng/mL, with a lower limit of quantitation of 1.46 ng/mL. Both intra- and interday precisions were within 11.05%, and the accuracy ranged from -11.5 to 5.51%. The extraction recovery and matrix effect were within acceptable limits. Stability tests showed that PepE remained stable throughout the analytical procedure. The validated method was then used to analyze the pharmacokinetics of PepE administered to rats orally (12.5 and 25 mg/kg) or intravenously (6.25 and 12.5 mg/kg).

[Difference of chemical constituents in Eucommiae Cortex from different habitats by LC-QTOF MS/MS].

In order to study the influence of ecological environment regarding the synthesis and accumulation of metabolites in Eucommiae Cortex, LC-QTOF MS/MS method combined with multivariate statistical analysis was used to analyze the differences of chemical constituents in Eucommiae Cortex from different habitats. Through the analysis of the multistage tandem mass spectrometry, the characteristic peaks were extracted with mass spectrometry data peak matching, peak alignment, and noise filtering. Principal component analysis (PCA) and partial least-squares discriminant analysis (PLS-DA) were used for data processing. The chemical constituents were identified or tentative presumed according to MS accurate mass and MS/MS spectrometry fragmentation information, combined with the software of database search, comparison with reference standards and literature. The results show the differences among samples of Eucommiae Cortex from different habitats are distinguishable. A total of 23 chemical constituents in Eucommiae Cortex were identified or tentative presumed. Among of them, 14 kinds of common differential chemical constituents (aucubin, geniposidic acid, neochlorogenic acid, syringin, olivil-4',4'-di-O-ß-D-glucopyranoside, chlorogenic acid, cryptochlorogenic acid, 1-hydroxypinoresinol- 4',4'-di-O-ß-D-glucopyranoside, caffeic acid, pinoresinol-di-O-ß-D-glucopyranoside, syringaresionl-di-O-ß-D-glucopyranoside, pinoresinol-4'-O-ß-D-glucopyranoside, eucommiol, isochlorogenic acid C and asiatic acid) presented different changing laws. This study provides basic information for revealing the influence law of ecological environment on the biosynthesis of metabolites in Eucommiae Cortex.

Peperomin E reactivates silenced tumor suppressor genes in lung cancer cells by inhibition of DNA methyltransferase.

Advanced lung cancer has poor prognosis owing to its low sensitivity to current chemotherapy agents. Therefore, discovery of new therapeutic agents is urgently needed. In this study, we investigated the antitumor effects of peperomin E, a secolignan isolated from Peperomia dindygulensis, a frequently used Chinese folk medicine for lung cancer treatment. The results indicate that peperomin E has antiproliferative effects, promoting apoptosis and cell cycle arrest in non-small-cell lung cancer (NSCLC) cell lines in a dose-dependent manner, while showing lower toxicity against normal human lung epidermal cells. Peperomin E inhibited tumor growth in A549 xenograft BALB/c nude mice without significant secondary adverse effects, indicating that it may be safely used to treat NSCLC. Furthermore, the mechanisms underlying the anticancer effects of peperomin E have been investigated. Using an in silico target fishing method, we observed that peperomin E directly interacts with the active domain of DNA methyltransferase 1 (DNMT1), potentially affecting its genome methylation activity. Subsequent experiments verified that peperomin E decreased DNMT1 activity and expression, thereby decreasing global methylation and reactivating the epigenetically silenced tumor suppressor genes including RASSF1A, APC, RUNX3, and p16INK4, which in turn activates their mediated pro-apoptotic and cell cycle regulatory signaling pathways in lung cancer cells. The observations herein report for the first time that peperomin E is a potential chemotherapeutic agent for NSCLC. The anticancer effects of peperomin E may be partly attributable to its ability to demethylate and reactivate methylation-silenced tumor suppressor genes through direct inhibition of the activity and expression of DNMT1.

Quality Evaluation of Pseudostellariae Radix Based on Simultaneous Determination of Multiple Bioactive Components Combined with Grey Relational Analysis.

Pseudostellariae Radix (PR) is an important traditional Chinese herbal medicine (TCM) with vast clinical consumption because of its positive effects. However, little attention has been devoted to simultaneous analysis of its bioactive components for quality control of PR based on its different harvesting times, different growing habitats, and different processing methods. In this research, the quality of PR was evaluated based on simultaneous determination of multiple bioactive components combined with grey relational analysis (GRA). A reliable method based on ultra-fast liquid chromatography tandem triple quadrupole mass spectrometry (UFLC-QTRAP-MS/MS) was established to simultaneously determine the contents of 30 components in PR, including two cyclopeptides, 12 nucleosides, and 16 amino acids. Furthermore, grey relational analysis was performed to evaluate the quality of PR samples according to the contents of these 30 components. The results showed that the quality of PR harvested in 6 August 2013, cultivated in Jurong, Jiangsu, and treated by oven drying 60 °C was better than that of other PR samples. The proposed method is useful for the overall assessment on the quality of PR, and this study provides valuable information for revealing the dynamic change laws of metabolite accumulation in PR and choosing the most suitable harvesting time and reasonable processing method of PR to obtain the best quality.

Predicting Deleterious Non-Synonymous Single Nucleotide Polymorphisms (nsSNPs) of HRAS Gene and In Silico Evaluation of Their Structural and Functional Consequences towards Diagnosis and Prognosis of Cancer.

The Harvey rat sarcoma (HRAS) proto-oncogene belongs to the RAS family and is one of the pathogenic genes that cause cancer. Deleterious nsSNPs might have adverse consequences at the protein level. This study aimed to investigate deleterious nsSNPs in the HRAS gene in predicting structural alterations associated with mutants that disrupt normal protein-protein interactions. Functional and structural analysis was employed in analyzing the HRAS nsSNPs. Putative post-translational modification sites and the changes in protein-protein interactions, which included a variety of signal cascades, were also investigated. Five different bioinformatics tools predicted 33 nsSNPs as "pathogenic" or "harmful". Stability analysis predicted rs1554885139, rs770492627, rs1589792804, rs730880460, rs104894227, rs104894227, and rs121917759 as unstable. Protein-protein interaction analysis revealed that HRAS has a hub connecting three clusters consisting of 11 proteins, and changes in HRAS might cause signal cascades to dissociate. Furthermore, Kaplan-Meier bioinformatics analyses indicated that the HRAS gene deregulation affected the overall survival rate of patients with breast cancer, leading to prognostic significance. Thus, based on these analyses, our study suggests that the reported nsSNPs of HRAS may serve as potential targets for different proteomic studies, diagnoses, and therapeutic interventions focusing on cancer.

Anti-depressant effect of Zhi-zi-chi decoction on CUMS mice and elucidation of its signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Zhi-zi-chi decoction (ZZCD) is used for treating depression as an effectively traditional Chinese medicine. Until now, studies on pharmacological research of ZZCD have mostly been centered in pharmacokinetic level. Little was known about its pharmacological mechanism of relieving depression. AIM OF THE STUDY: This study was to evaluate the effect of ZZCD on relieving depression via behavioral tests, serum metabolomics and signaling target expression analysis on chronic unpredictable mild stress (CUMS) model mice. MATERIALS AND METHODS: The CUMS exposure lasted 7 consecutive weeks. The mice were administrated with ZZCD for the last 3 weeks. Behavioral tests were applied and a serum metabolomics method based on UFLC/Q-TOF-MS with multivariate statistical and global metabolic network analysis was performed to identify relevant metabolites and pathways. Finally, the protein expressions in mouse hippocampi were determined by western blot to verify the metabolomics deduction. RESULTS: Behavioral parameters were visibly changed after modeling, while high and medium dosage groups showed status improvement compared to the model group. Seventy six metabolites were identified as potential biomarkers from the metabolomics profiles in C18 and HILIC systems. In addition, 9 significant pathways related to changed biomarkers were conducted. The pathways were closely connected by some key targets, which were significantly reduced in the model group compared with those in control group, while ZZCD treated groups showed corrections after 3-week administration. The results revealed that the anti-depression efficacy of ZZCD might be associated with PKA-CREB-BDNF-TrkB-PSD-95 pathway influenced by metabolic changes, verifying the pathway annotation speculation. CONCLUSION: This study demonstrated that ZZCD had a positive treatment effect on CUMS depression model mice. Metabolomics results revealed the holistic and interconnected metabolic changes of ZZCD in CUMS mice. The metabolic pathway annotation suggested that the anti-depression mechanism of ZZCD might be related to signaling pathway in brain. PKA-CREB-BDNF-TrkB-PSD-95 signaling expression was a verification and complement to the metabolomics results.

Evaluation of insulated isothermal PCR devices for the detection of avian influenza virus.

Highly pathogenic avian influenza viruses (HPAIVs) and low pathogenic avian influenza viruses (LPAIVs) represent important threats to the poultry industry and global human health. Due to the high rates of avian influenza virus (AIV) transmission, controlling AIV outbreaks is challenging. HPAIV is known to be transmitted from wild birds to domestic ducks, from which it can be transmitted to layer and broiler chickens. Therefore, surveillance of AIV in domestic ducks and chickens in advance of outbreaks can prevent its spread and enable timely implementation of disease control measures. Certain molecular diagnostic tools can be applied in the field for faster AIV detection. In this study, we evaluated the AIV-detection ability of two insulated isothermal PCR (iiPCR) devices, POCKIT™Micro DUO Nucleic Acid Analyzer (POCKIT DUO) and POCKIT™ Central Nucleic Acid Analyzer (POCKIT Central). We found that the analytical, in vivo and clinical performances of the two POCKIT devices were comparable to those of real-time reverse transcription PCR. Due to their brief protocols and short detection times, POCKIT DUO and POCKIT Central represent promising molecular diagnostic devices for the reliable detection of AIV.

Comprehensive Comparison of Two Color Varieties of Perillae Folium Using Rapid Resolution Liquid Chromatography Coupled with Quadruple-Time-of-Flight Mass Spectrometry (RRLC-Q/TOF-MS)-Based Metabolic Profile and in Vivo/in Vitro Anti-Oxidative Activity.

Perillae Folium (PF), which is extensively used as a dietary vegetable and medicinal herb, contains two varietal forms corresponding to purple perilla leaf (Perilla frutescens var. crispa) and green perilla leaf (Perilla frutescens var. frutescens). However, the components and efficacy of different PF varieties remain underexplored so far. In the present work, a nontargeted rapid resolution liquid chromatography coupled with quadruple-time-of-flight mass spectrometry (RRLC-Q/TOF-MS)-based metabolomics approach was developed to investigate the difference in the chemical compositions between green PF and purple PF. A total of 71 compounds were identified or tentatively identified, among which 7 phenolic acids, 10 flavonoids, and 9 anthocyanins were characterized as differential metabolites. In addition, heatmap visualization and ultraperformance liquid chromatography coupled with triple-quadrupole tandem mass spectrometry (UPLC-TQ-MS/MS)-based quantitative analysis revealed that flavonoids and anthocyanins especially had higher contents in purple PF. Furthermore, the anti-oxidative activities of two varietal PFs were evaluated in vivo zebrafish and in vitro human umbilical vein endothelial cells (HUVECs). The results showed that the purple PF had more pronounced anti-oxidative activities than did the green PF, which may be due to the presence of anthocyanins and a higher concentration of flavonoids in its phytochemical profile. The outcome of the present study is expected to provide useful insight on the comprehensive utilization of a PF resource.

LC-Q-TOF/MS-oriented systemic metabolism study of pedunculoside with in vitro and in vivo biotransformation.

As a triterpene saponin, pedunculoside is one of the most abundant, representative and active components in plants of genus Ilex (Aquifoliaceae). Pedunculoside has been used to treat myocardial ischemia, ameliorate hyperlipidemia and prevent liver injury. In this paper, a systemic in vitro liver microsomes / S9 and intestinal bacteria incubation, and in vivo animal experiment were performed, using LC-Q-TOF/MS analysis and a three-step data processing protocol. As a result, Bifidobacterium adolescentis and Bifidobacterium breve were identified to potentially metabolize pedunculoside among the intestinal bacteria tested. A total of 11 metabolites were found and tentatively identified, with 6 in both microsomal and bacterial incubation systems, and 9 after rats orally administered with pedunculoside. The metabolites detected involving both phase I and phase II metabolism, mainly through deglycosylation (hydrolyzation), dehydrogenation, hydroxylation and conjugation, and some of them underwent more than one-step metabolic reactions. Most of the metabolites have not been reported before. In vitro, liver microsome and intestinal bacteria prefer to metabolize pedunculoside in totally different ways; while in vivo, intestinal tract is the most important site for the metabolism and excretion of pedunculoside, where both intestinal bacteria and the host metabolic enzymes participate in its metabolism and disposition. The importance of intestinal bacteria should be highlighted. This study would contribute to a better understanding of pedunculoside metabolism, which can provide scientific evidence for its pharmacodynamic mechanism research and prove its clinical application.