Profiling alkaloids in Aconitum pendulum N. Busch collected from different elevations of Qinghai province using widely targeted metabolomics.

Aconitum pendulum N. Busch (Ranunculaceae) is rich in alkaloids with anti-inflammatory and analgesic activities. Many studies have focused on the identification or quantification of alkaloid components using low-throughput tests. However, the metabolic differences of plants from environmentally distinct regions remain unclear. The present study profiled alkaloid chemical compounds in the rhizomes of A. pendulum from different regions. A total of 80 chemical compounds were identified using a widely targeted ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) approach. Principal component, hierarchical clustering, and orthogonal partial least squares-discriminant analyses of the chemical compounds indicated that the plants from 6 regions clearly separated into distinct groups. A total of 19 compounds contributed the most to the metabolite differences between collection areas and were identified as potential metabolic markers. The anti-inflammatory activities of the A. pendulum extracts were also evaluated and the potential environmental effects on the regulation of metabolite composition and bioactivity were explored. These results improve our understanding of the variation in chemical composition of plants from different regions and will serve as a reference for evaluating the medicinal value of A. pendulum in different environments.

Bioactive halogenated dihydroisocoumarins produced by the endophytic fungus Lachnum palmae isolated from Przewalskia tangutica.

Guided by the UPLC-ESIMS profile, seven previously undescribed halogenated dihydroisocoumarins, palmaerones A-G, along with eleven known dihydroisocoumarins, were isolated from Lachnum palmae, an endophytic fungus from Przewalskia tangutica by exposure to a histone deacetylase inhibitor SAHA. Structures of the isolates were elucidated by analysis of their NMR, MS and optical rotation values. The antimicrobial, anti-inflammatory and cytotoxic activities of palmaerones A-G were evaluated. Palmaerones A-G showed antimicrobial activities against the strains (C. neoformans, Penicillium sp., C. albicans, B. subtilis and S. aureus), and palmaerone E exhibited potential antimicrobial activities against all the test strains with the MIC value in the range of 10-55 µg/mL. Palmaerones A and E exhibited moderate inhibitory effects on NO production in LPS-induced RAW 264.7 cells, with the IC50 values of 26.3 and 38.7 µM, respectively and no obvious toxicities were observed at 50 µM. Palmaerone E showed weak cytotoxicity against HepG2 with the IC50 value of 42.8 µM. This work provides an effective strategy for expanding natural product resource.

Analysis of the lithiated leucosceptroids from Leucosceptrum canum to facilitate their identification and differentiation by electrospray ionization tandem mass spectrometry.

RATIONALE: Leucosceptroids, isolated from Leucosceptrum canum (L. canum), are sesterterpenoids with novel molecular scaffolds which possess potent antifeedant activities. Their molecular scaffolds comprise a 5,6,5-framework with a great diversity due to different hydroxylation and substituent positions. The biological activities of leucosceptroids are affected by the subtle structural differences. The structural characterization and differentiation of the leucosceptroid isomers are of great importance. METHODS: Firstly, different kinds of cation adducts of leucosceptroids were examined by adding alkali metal ions to the solution, and the lithiated adducts of leucosceptroid were found to be readily formed and yielded characteristic fragment ions under collision-induced dissociation (CID). Then, a systematic mass spectrometric investigation of the [M + Li](+) ions was carried out to clarify their characteristic fragment pathways by electrospray ionization quadrupole time-of-flight-type tandem mass spectrometry (ESI-QTOF-MS/MS). The proposed fragmentation pathways were confirmed through ion trap ESI-MS(n) (n ≥ 3) spectra. Finally, the proposed MS/MS method was applied to investigate the extracts of L. canum. RESULTS: A specific fragmentation pathway of the lithiated adduct, which leads to the production of diagnostic ions of leucosceptroids, was observed. This fragmentation is initiated by cleavage of the C-ring and leads to formation of two types of ions by further dissociation. Both pathways could yield characteristic fragment ions, which could be used to define the substituents at the skeletal structure and at the C-ring. The representative characteristic fragmentations of [M + Li](+) ions and the proposed fragmentation pathways were successfully adopted to investigate the L. canum extracts, and a total of eleven leucosceptroids were identified or tentatively characterized. CONCLUSIONS: The characteristic fragmentation pathways of [M + Li](+) species of leucosceptroid isomers were proposed. Three types of leucosceptroid isomers were successfully differentiated. Eleven leucosceptroids were characterized from L. canum extracts. The fragmentation knowledge will facilitate the analysis of leucosceptroids and other sesterterpenoids in future research. Copyright © 2016 John Wiley & Sons, Ltd.

Structural characterization and antiviral effect of a novel polysaccharide PSP-2B from Prunellae Spica.

In the present study, a novel polysaccharide, PSP-2B, was isolated from aqueous extracts of Prunellae Spica by direct ultrafiltration membrane separation and gel chromatography purification. PSP-2B is a partially sulphated polysaccharide with a molecular weight of approximately 32kDa. Its sulfate content is 10.59% by elemental analysis. The major sugars comprising PSP-2B are arabinose, galactose and mannose, in addition to small amounts of glucose and uronic acids. The framework of PSP-2B is speculated to be a branched arabinogalactomannan, and the side chains are terminated primarily by the Araf residues. PSP-2B also contains 2.98% protein. PSP-2B exhibits activity against herpes simplex virus (HSV), with a half maximal inhibitory concentration (IC50) of approximately 69µg/mL for HSV-1 and 49µg/mL for HSV-2. However, PSP-2B demonstrated no cytotoxicity even when its concentration was increased to 1600µg/mL, suggesting that it has potential as an anti-HSV drug candidate.

Multiple Differential Networks Strategy Reveals Carboplatin and Melphalan-Induced Dynamic Module Changes in Retinoblastoma.

BACKGROUND Retinoblastoma (RB) is the most common malignant tumor of the eye in childhood. The objective of this paper was to investigate carboplatin (CAR)- and melphalan (MEL)-induced dynamic module changes in RB based on multiple (M) differential networks, and to generate systems-level insights into RB progression. MATERIAL AND METHODS To achieve this goal, we constructed M-differential co-expression networks (DCNs), assigned a weight to each edge, and identified seed genes in M DCNs by ranking genes based on their topological features. Starting with seed genes, a module search was performed to explore candidate modules in CAR and MEL condition. M-DMs were detected according to significance evaluations of M-modules, which originated from refinement of candidate modules. Further, we revealed dynamic changes in M-DM activity and connectivity on the basis of significance of Module Connectivity Dynamic Score (MCDS). RESULTS In the present study, M=2, a total of 21 seed genes were obtained. By assessing module search, refinement, and evaluation, we gained 18 2-DMs. Moreover, 3 significant 2-DMs (Module 1, Module 2, and Module 3) with dynamic changes across CAR and MEL condition were determined, and we denoted them as dynamic modules. Module 1 had 27 nodes of which 6 were seed genes and 56 edges. Module 2 was composed of 28 nodes and 54 edges. A total of 28 nodes interacted with 45 edges presented in Module 3. CONCLUSIONS We have identified 3 dynamic modules with changes induced by CAR and MEL in RB, which might give insights in revealing molecular mechanism for RB therapy.

The Tissue Distribution and Urinary Excretion Study of Gallic Acid and Protocatechuic Acid after Oral Administration of Polygonum Capitatum Extract in Rats.

In the present study, we investigated the tissue distribution and urinary excretion of gallic acid (GA) and protocatechuic acid (PCA) after rat oral administration of aqueous extract of Polygonum capitatum (P. capitatum, named Herba Polygoni Capitati in China). An UHPLC-MS/MS analytical method was developed and adopted for quantification of GA and PCA in different tissue homogenate and urine samples. Interestingly, we found that GA and PCA showed a relatively targeted distribution in kidney tissue after dosing 60 mg/kg P. capitatum extract (equivalent to 12 mg/kg of GA and 0.9 mg/kg of PCA). The concentrations of GA and PCA in the kidney tissue reached 1218.62 ng/g and 43.98 ng/g, respectively, at one hour after oral administration. The results helped explain the empirical use of P. capitatum for kidney diseases in folk medicine. Further studies on urinary excretion of P. capitatum extract indicated that GA and PCA followed a concentrated elimination over a 4-h period. The predominant metabolites were putatively identified to be 4-methylgallic acid (4-OMeGA) and 4-methylprotocatechuic acid (4-OMePCA) by analyzing their precursor ions and characteristic fragment ions using tandem mass spectrometry. However, the amount of unchanged GA and PCA that survived the metabolism were about 14.60% and 15.72% of the total intake, respectively, which is reported for the first time in this study.

HPLC-DAD-ELSD Combined Pharmacodynamics and Serum Medicinal Chemistry for Quality Assessment of Huangqi Granule.

OBJECTIVE: To more scientifically and reasonably control the quality of Huangqi Granules, preliminary studies on the pharmacodynamics and serum pharmacochemistry of this medicine were performed. DPPH and MTT experiments showed that water extracts of Huangqi Granules had good antioxidant activity and increased immunity. Timed blood samples collected 5 min, 15 min, and 30 min after oral administration of a set amount of Huangqi Granules were collected and tested using UPLC-ESI-MS/MS. As a result, calycosin-7-O-ß-D-glucoside, ononin, calycosin, astragaloside IV, and formononetin were found to exist in rat blood after dosing, indicating that the five chemical compounds might have pharmacological activity, and based on this result, they were designated biomarkers for quality control of Huangqi Granules. Consequently, a simple, rapid and efficient method was developed in the present study for the simultaneous determination of the five characteristic compounds in Huangqi Granules using HPLC-DAD-ELSD. MATERIALS AND METHODS: The separation was performed using an Agilent Hypersil ODS column (4.6 × 250 mm, 5 µm) at 30 ℃. The mobile phase was composed of water (solvent A) and acetonitrile (solvent B) with a flow rate of 1 mL/min. The drift tube temperature of the ELSD system was set to 85 ℃, and the nitrogen pressure was 3.5 bar. RESULTS: All five characteristic compounds had good linear behavior with r2 values greater than 0.9972. The recoveries varied from 96.31% to 101.22%. Subsequently, the developed method was applied to evaluate the quality of Huangqi Granules from different batches, and hierarchical clustering analysis (HCA) was used to analyze the classification of the samples based on the values of the five compounds. CONCLUSION: The established HPLC method combined with HCA proved to be effective to evaluate the quality of Huangqi Granules.