As a treasure resource of novel drug lead compounds, how to rapidly and high-efficiently screen and isolate active components from natural products is critical. Thanks to its high resolution, high automation and flexible integration, online two-dimensional liquid chromatography has great potential for screening active ingredients from complex matrices by integrating a highly specific bio-recognition process into a two-dimensional liquid chromatography system before, on or after the column separation. This review comprehensively summarized recent developments, applications and shortcomings of online two-dimensional liquid chromatography for natural product screening from different integration modes, including pre-column, on-column and post-column screening methods.

As one kind of v-myb avian myeloblastosis viral oncogene homolog (MYB) transcription factors, R1-MYB (MYB-related) family plays an important role in plant growth and development, as well as environmental stress and hormone signal transduction. In this study, R1-MYB family genes in Rheum palmatum L. were systematically screened based on full-length transcriptome sequencing analysis. Firstly, the physicochemical, protein domain and molecular evolution characteristics of the coding proteins were analyzed. Furthermore, the tissue expression levels of R1-MYB genes were analyzed by RNA-seq. We also investigated the expression pattern of RpMYB24 in response to various hormones and abiotic stresses. The results showed that a total of 49 R1-MYB genes were identified, which mainly encoded thermally stable hydrophilic proteins. Most of the deduced proteins were predicted to locate in nucleus. Each protein had a large proportion of random curl and α helix, and also had the W-type conserved amino acids which were the signature of MYB. R1-MYB family members were distributed in five subgroups, including circadian clock associated 1 (CCA1)-like, I-box (GATAAG)-like, CAPRICE (CPC)-like, telomere repeat binding factor (TRF)-like and TATA binding protein (TBP)-like, and the number of CCA1-like was the majority. RNA-seq revealed that 49 R1-MYB genes were differentially expressed in roots, rhizomes and leaves of R. palmatum, and the expression levels of 15 and 23 genes in roots and rhizomes were higher than those in leaves, respectively. RpMYB24 transcript was induced by abscisic acid (ABA), salicylic acid (SA), and methyl jasmonate (MeJA) treatment, and could also significantly respond to injury, low temperature and high temperature stresses except drought stress. This study systematically identified the R1-MYB family genes and their molecular characteristics, better for further gene functional validation, and then provide a scientific basis for the transcriptional regulation mechanism research into rhubarb quality formation.

The goal of this work was to investigate the antidepressant fraction from Radix Paeoniae Alba and identify its major chemical constituents. Corticosterone injured rat phaeochromocytoma (PC12) cells and behavioral despair depression models of mice were used to evaluate the antidepressant effects of Radix Paeoniae Alba (Bai-Shao) ethanol extract (BS-E) and its three fractions (BS-10E, BS-60E, BS-95E) isolated by macroporous resin column chromatography. Animal experimental procedures were approved by the Animal Ethics Committee of the Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences and Peking Union Medical College (approval No.: SLXD-20210618051). The results showed that BS-E, BS-10E and BS-60E had protective effects against PC12 cells injury induced by corticosterone, among which BS-60E had the strongest protective effect. BS-60E could significantly shorten the time of forced swimming and tail suspension in despair depression models of mice, and was identified as the antidepressant fraction of Radix Paeoniae Alba. The major chemical constituents in the antidepressant fraction were identified by ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS), and their proposed fragmentation pathways in MS spectra were deduced. A total of 79 chemical constituents were identified from BS-60E, including 36 monoterpenes, 34 polyphenols, 6 oligosaccharides, and 3 other constituents, and monoterpenes and polyphenols may be major effective constituents of BS-60E.

Based our previous work, twelve purine derivatives were designed and synthesized as dual modulators of GPR119 and DPP-4by conjugating the GPR119 activating and DPP-4 inhibiting fragments with the position 6 and 9 of purine core via an approach of merged pharmacophores. Compound 11, bearing 2-fluoro-4-methylsulphonyl anilide and cyanopyrrolidine moieties, exhibited the most potent GPR119 agonistic activities (EC₅₀ = 0.33 μmol·L^-1, IA = 71.1%) and DPP-4 inhibitory (58.4% inhibition at 10 μmol·L^-1, 21.2% inhibition at 1 μmol·L^-1) activities in the in vitro antidiabetic study. Subsequently, we performed studies on structure activity relationships and molecular docking to guide the further drug design.

A hydrophilic interaction chromatography tandem mass spectrometry method was developed for simultaneous quantification of 35 components in gualoupi injection. The analytes were separated with an ACQUITY XBridge Amide column using 20 mmol·L^-1 ammonium formate aqueous solution (pH 3.0) as mobile phase A and 20 mmol·L^-1 ammonium formate (pH 3.0)∶acetonitrile (1∶9) as mobile phase B for gradient elution. Mass spectrometry with dynamic multiple reaction monitoring and external standard method were used for quantitative analysis. A total of 35 components were determined in 10 batches of gualoupi injection. The results showed that the 35 compounds had a good linear relationship within their respective concentration ranges with the correlation coefficients (R² > 0.998 0), the recoveries ranged from 76.6% to 118.5%. The results showed that γ-aminobutyric acid, trigonelline, alanine, threonine, homoserine, citrulline, and leucine were abundant in gualoupi injection, while nicotinamide, methylsuccinic acid, cytosine and choline account for a low percentege. The present study provides an important reference for elucidation of the effective material basis and the improvement of quality standard of gualoupi injection.

A BBB co-culture cell model consisting of rat brain microvascular endothelial cells (BMEC) and astrocytes (AS) was established to study the effect of Angelica dahurica coumarins on the transport behavior of puerarin across blood-brain barrier (BBB) in vitro and in vivo. The barrier function of this model was evaluated by measuring the transendothelial resistance, phenol red permeability and BBB related protein expression. The permeability assay and western blot methods were performed to study the effects of Angelica dahurica coumarins on the BBB permeability and the expression of BBB related protein. The animal experiment protocols in this study were approved by the Animal Ethics Committee of Xi'an Jiaotong University (Animal Ethics No.: 2021-1329). The results showed that the established BMEC/AS co-culture model could be used to evaluate drug transport across BBB in vitro. After combined with Angelica dahurica coumarins, the transport capacity of puerarin was significantly increased in vitro and in vivo. Additionally, Angelica dahurica coumarins enhanced BBB permeability and inhibited the protein expression of P-glycoprotein (P-gp), zonula occludens-1 (ZO-1) and occludin. Angelica dahurica coumarins might increase BBB permeability by inhibiting the expression of P-gp and tight junction protein, thereby increasing the content of puerarin in brain tissue.

O-methyltransferases (OMTs) are one of the key tailoring enzymes in the biosynthesis of many natural products, O-methylation can not only reduce the reactivity of natural products, but also alter their solubility, stability and biological activities. Based on the transcriptome data of Ardisia japonica, a full-length cDNA sequence of candidate OMT (termed as AjOMT1) was cloned by reverse transcription-polymerase chain reaction (RT-PCR) and expressed in Escherichia coli (E. coli) for the first time. In vitro enzyme catalytic activity assay showed that the recombinant AjOMT1 could effectively catalyze quercetin to form O-methylated products. Most importantly, AjOMT1 showed unprecedented substrate promiscuity towards structurally various compounds including flavonoids, stilbenes, coumarins, alkaloids and phenylpropanoids, especially preferring to the compounds with adjacent phenolic hydroxyl groups, and showed regio-selectivity. These results suggested that AjOMT1 could be used as the tool enzyme to conduct O-methylation of different types of compounds to produce diverse active methylated products, and provide a new method for drug discovery, even universal part for synthetic biology of natural products.

Human hormones at trace levels play a vital role in the regulation of a variety of functions and systems in the body, and an imbalance in hormone levels can lead to the emergence and development of diverse diseases. Therefore, the development of reliable sample pretreatment methods and sensitive and accurate analytical techniques for human hormone detection could contribute to the prevention, diagnosis and treatment of diseases, providing significant improvement for human health. Human samples which are usually used to detecting hormones, such as blood, saliva, urine and other matrix are more complex, so sample pretreatment is an important step to ensure the accuracy and reliability in the detection of hormones. In this review three common sample pretreatment methods including solid phase extraction (SPE), liquid-liquid extraction (LLE) and protein precipitation (PP) methods are discussed. Then, recent research progress in conventional techniques like liquid/gas chromatography and liquid/gas chromatography-mass spectrometry (LC/GC-MS/MS), as well as some novel strategies, such as immunoassay including chemiluminescence immunoassay (CLIA), lateral-flow immunoassay (LFIA) and time-resolved fluoroimmunoassay (TRFIA), and sensor technology including electrochemical (EC), fluorescent (FL) and surface-enhanced Raman scattering (SERS) sensors, and microfluidic chip analysis are discussed for human hormone detection. Finally, the future perspective on the use of these methods for hormone detection is considered. It is hoped to provide powerful insights to researchers for the relevant researches.

The complete chloroplast genome of medicinal plant Asarum caudigerum Hance and its close relative A. cardiophyllum Franchet were sequenced using Illumina Hiseq technology, and assembled, annotated, and characterized by bioinformatic methods in this study. Then phylogenetic analysis of the complete chloroplast genomes of A. caudigerum, A. cardiophyllum, and twelve published species was conducted. The results indicated that the chloroplast genomes ranged from 186 215-186 985 bp in length, with a large single copy (LSC, 89 445-90 169 bp) and two inverted repeats (IRa/IRb, 48 387-48 408 bp). The overall GC content was 37.4%-37.5%. A total of 144 chloroplast genes were annotated, including 98 protein coding genes, 38 tRNA genes and 8 rRNA genes. In addition, complex genomic rearrangements were detected in the chloroplast genome of Asarum. Meanwhile, visual evaluation of the discrete type of the sequence indicated that the variation level of non-coding region was higher than that of coding region. Phylogenetic analyses suggested that A. caudigerum and A. cardiophyllum were clustered into a single clade and A. cardiophyllum, A. sieboldii var. seoulense, A. misandrum and A. maculatum were clustered into another single branch. These two clade were sister species. This study provides a scientific basis for the identification, phylogenetic relationship, molecular breeding of Asarum species.

This study started from the effect of baicalin (BC), the main active component of the labiaceae plant Scutellaria baicalensis, on collagen-induced arthritis (CIA) in rats, to explore the mechanism of glucose metabolism reprogramming in fibroblast like synoviocytes (FLSs), a key effector cell of synovial inflammation in rheumatoid arthritis (RA). First of all, CIA rats and tumor necrosis factor-α (TNF-α)-induced RASFs in vitro and in vivo models were established, the arthritis index (AI) score and histopathological changes of CIA rats after BC administration were observed, and the levels of inflammatory factors in serum and cell supernatant were quantified by ELISA, immunocytochemistry and Western blot were used to detect the expression of G-protein-coupled receptor 81 (GPR81) and pyruvate dehydrogenase kinase 1 (PDK1) proteins. In addition, the kit was used to measure the levels of key products and enzyme activities in glucose metabolism reprogramming. The results showed that BC (50, 100 and 200 mg·kg^-1) could alleviate the symptoms of arthritis in CIA rats in a dose-dependent manner, inhibit synovial hyperplasia, alleviate the infiltration of inflammatory cells, down-regulate the levels of pro-inflammatory factors TNF-α and interleukin (IL)-1β, and up-regulate the levels of anti-inflammatory factor IL-10 in CIA rats. At the same time, the secretion levels of lactate, pyruvate, acetyl-CoA, citrate and the activity of lactate dehydrogenase B (LDH-B) were decreased, and the expressions of GRP81 and PDK1 were down-regulated, suggesting that BC mediated the reprogramming process of glucose metabolism. However, when GPR81 inhibitor 3-OBA inhibited lactate uptake, the activity of LDH-B was significantly increased, suggesting that BC inhibited the expression of PDK1, a key enzyme in the reprogramming metabolism from glycolysis to oxidative phosphorylation. All animal experiments in this study were conducted in accordance with the ethical standards of the Laboratory Animal Care Center of Anhui University of Chinese Medicine (approval number: AHUCM-rats-2021049). These studies revealed that baicalin mediated metabolic reprogramming of RASFs from glycolysis to oxidative phosphorylation by inhibiting PDK1 protein expression, and alleviated joint inflammation in CIA rats.