[Research progress on intestinal microecology regulating mechanism and biological activities of polysaccharides].

Intestinal microecology is an important defense system in the human body. The intestinal flora is the core micro-ecosystem in the human intestine. It has a symbiotic relationship with the overall functions of the body. It has strong metabolic activity to maintain the normal functioning of the body and resist the invasion of various viral antigens in the body. Playing a protective function,the imbalanced intestinal microecology can cause various diseases. Polysaccharides can be extracted from a wide range of sources and have low toxicity and side effects. They have attracted wide attention because of their anti-tumor, anti-oxidant, anti-inflammatory and other biological activities. Studies have demonstrated that polysaccharides can regulate intestinal microecological disorders. According to the studies in recent years, this review summarizes that polysaccharides mainly modulate intestinal microecological disorders through regulating the composition of intestinal flora, improving the metabolism of the flora, and repairing the intestinal tract barrier. On the basis of these mechanisms of action, this paper elaborates the anti-tumor, immunomodulatory, and anti-inflammatory activities of polysaccharides. This paper can provide reference for the future research on the intestinal microecology-regulating mechanism and biological activities of polysaccharides.

[Research progress on anti-hepatocellular carcinoma mechanism of active ingredients of traditional Chinese medicine].

According to the latest data, the annual mortality rate of liver cancer in China ranks the second among malignant tumors, and it has become one of the most fatal cancers in urban and rural areas. This article starts with the active ingredients of traditional Chinese medicines for liver cancer to summarize the relevant literature at home and abroad, with the hope to provide theoretical references for the development of traditional Chinese medicines against liver cancer. The results show that the active ingredients of traditional Chinese medicines such as Chinese herbal extracts, flavonoids, polysaccharides, alkaloids, saponins, volatile oils, terpenes, quinones, phenols, bioenzymes, and protein components are the entry points, mainly by inhibiting the proliferation of cancer cells, blocking the cancer cell cycle, inhibiting the invasion and metastasis of cancer cells, regulating signal pathways, regulating vascular endothelial growth factor(VEGF) and reactive oxygen species(ROS), etc. to play the effects against liver cancer. It can be seen that the active ingredients of traditional Chinese medicine with coordinated intervention effects at multiple levels and multiple targets are expected to become ideal medicines for the treatment of liver cancer.

[Research progress on mechanism of active ingredients in traditional Chinese medicine for gastric cancer].

Gastric cancer is a disease with high mortality, which threatens the health of people for a long time. At present, the main treatment methods are surgery and chemotherapy, but these methods have great harm to the human body. However, it is found that the active ingredients of traditional Chinese medicine have an obvious therapeutic effect in the adjuvant treatment of the tumor. Therefore, the active ingredients of traditional Chinese medicine have become a research hotspot in the anti-tumor field. In recent years, many related researchers have been particularly active in studying the in vitro activity and mechanism of active ingredients of traditional Chinese medicine on human gastric cancer cells. In this paper, the Chinese herbal medicine extracts, polysaccharides, alkaloids, saponins, flavones, terpenes, quinones, volatile oils, esters, phenols, protein components and other active ingredients of Chinese medicine were used as the starting points to investigate the anti-gastric cancer mechanism, such as inhibiting cell proliferation and inducing apoptosis of cancer cells, inhibiting cell invasion and migration; inhibiting over-expression of vascular endothelial growth factor(VEGF); interfering with cell mitosis; and regulating cell signaling pathways. Their in vitro inhibitory activity and mechanism for gastric cancer cells were described in this study, providing a theoretical reference for the development and application of anti-gastric cancer drugs.

[Research progress in mechanism of traditional Chinese medicine active ingredients against cervical cancer].

Cervical cancer is the second cancer that threatens women' s health,and has attracted the attention of researchers at home and abroad because of its extremely high mortality rate. At present,most of the radiotherapy methods and chemical drugs for cancer treatment have serious side effects,and the active ingredients of traditional Chinese medicine have become the key research and development targets of anti-cancer drugs due to many advantages,such as multi-channel,multi-link,multi-target,and less toxicity. In recent years,researchers have been particularly active in researching the inhibitory activity and mechanism of active ingredients of traditional Chinese medicine for human cervical cancer cells. In this paper,the inhibitory activity and mechanism of traditional Chinese medicine against human cervical cancer cells were investigated from crude extract of traditional Chinese medicine,polysaccharides,alkaloids,saponins,flavonoids,terpenoids,quinones,volatile oils,esters,phenols,arsenical,protein components as the starting point; anti-cervical cancer mechanism was investigated,such as inhibiting cell proliferation inducing apoptosis of cancer cells,inhibiting cell invasion,migration and focal adhesion kinase( FAK) phosphorylation,inhibiting vascular endothelial growth factor( VEGF)over-expression interfering with cell mitosis,inhibiting Granzyme activity,regulating cellular signaling pathway,down-regulating HPV E6 gene expression,and regulating immune function. Its in vitro inhibitory activity and mechanism of action on cervical cancer cells were reviewed,in order to provide a theoretical basis for the development and utilization of anti-cervical cancer drugs.

Optimized microwave-assistant extraction combined ultrasonic pretreatment of flavonoids from Periploca forrestii Schltr. and evaluation of its anti-allergic activity.

Microwave extraction combined ultrasonic pretreatment of flavonoids from Periploca forrestii Schltr. was investigated in this study, extraction process was first performed in an ultrasonic cleaner, then treated by microwave irradiation. The optimum ultrasonic time of 25 min was selected by single-factor experiments. A response surface methodology has been used to obtain a mathematical model that describes the process and analyzes the significant parameters ethanol concentration 59.92%, liquid to raw materials ratio 21.24 mL/g, microwave radiation time 209.53 s, and microwave power 274.14 w. In these optimum conditions, the yield of flavonoids from P. forrestii (TFPF) could be up to 9.11 ± 0.08%, which was increased by 14.30 and 19.86% compared microwave extraction and ultrasonic extraction, respectively. In vitro suppress hyaluronidase experimentation showed that TFPF purified using polyamide exhibited good anti-hyaluronidase ability with IC50 value of 1.033 mg/mL, possessing certain anti-antiallergic and potential application prospect in pharmaceutical production of treating inflammation and other related fields.

UHPLC-ESI-MS/MS determination and pharmacokinetics of pinoresinol glucoside and chlorogenic acid in rat plasma after oral administration of Eucommia ulmoides Oliv extract.

This study aimed to develop a specific UHPLC-ESI-MS/MS method for simultaneous determination and pharmacokinetics of pinoresinol glucoside and chlorogenic acid in rat plasma after oral administration of Eucommia ulmoides. The chromatographic separation was achieved on a Hypersil GOLD column with gradient elution by using a mixture of 0.1% formic acid aqueous solution and acetonitrile as the mobile phase at a flow rate of 200 µL/min. A tandem mass spectrometric detection was conducted using multiple-reaction monitoring via an electrospray ionization source in negative ionization mode. Samples were pre-treated by a single-step protein precipitation with acetonitrile, and bergenin was used as internal standard. After oral administration of 3 mL/kg E. ulmoides extract in rats, the maximum plasma concentrations of pinoresinol glucoside and chlorogenic acid were 57.44 and 61.04 ng/mL, respectively. The times to reach the maximum plasma concentration were 40.00 and 23.33 min for pinoresinol glucoside and chlorogenic acid, respectively. The intra- and inter-day precision (RSD) values for the two analytes were <2.46 and 5.15%, respectively, and the accuracy (RE) values ranged from -12.76 to 0.00. This is the first study on pharmacokinetics of bioactive compounds in rat plasma after oral administration of E. ulmoides extract.

Relative tissue distribution and excretion studies of gastrodin and parishin from powder and extract of Gastrodiae Rhizoma in rat by UHPLC-ESI-MS/MS.

New research has indicated that Gastrodiae Rhizome (GR) has potential anti-diabetic and anti-asthmatic effects in mouse models. On the basis of our previous study of the relative bioavailability of gastrodin (GAS) and parishin (PA) from extract and powder of GR, we performed further research on the tissue distribution and excretion of the two analytes. A reliable bioanalytical method for the quantification of GAS and PA in rat tissues and excretion is required. Chromatographic separation was carried out on a gradient mobile phase of acetonitrile-water with 0.1% formic acid. Calibration curves (1/x2 weighted) offered satisfactory linearity (r2 > 0.9835) within 100-3000 ng mL-1 for GAS and (r2 > 0.9862) within 10-1000 ng mL-1 for PA. The relative standard deviations of the intra-day and inter-day precision were all <14.98%, whilst the relative errors of the intra-day and inter-day accuracy were all within ±14.71%. The matrix effect and recovery values were satisfactory in all of the biological matrices examination. The data of relative differences in tissue distribution and excretion of GAS and PA from powder and extract of GR indicated that higher bioavailabilities for GAS and PA were obtained when a dosage of 4 g kg-1 GR powder was used.

Crystal structures of Apo and GMP bound hypoxanthine-guanine phosphoribosyltransferase from Legionella pneumophila and the implications in gouty arthritis.

Hypoxanthine-guanine phosphoribosyltransferase (HGPRT) (EC 2.4.2.8) reversibly catalyzes the transfer of the 5-phophoribosyl group from 5-phosphoribosyl-alpha-1-pyrophosphate (PRPP) to hypoxanthine or guanine to form inosine monophosphate (IMP) or guanosine monophosphate (GMP) in the purine salvage pathway. To investigate the catalytic mechanism of this enzyme in the intracellular pathogen Legionella pneumophila, we determined the crystal structures of the L. pneumophila HGPRT (LpHGPRT) both in its apo-form and in complex with GMP. The structures reveal that LpHGPRT comprises a core domain and a hood domain which are packed together to create a cavity for GMP-binding and the enzymatic catalysis. The binding of GMP induces conformational changes of the stable loop II. This new binding site is closely related to the Gout arthritis-linked human HGPRT mutation site (Ser103Arg). Finally, these structures of LpHGPRT provide insights into the catalytic mechanism of HGPRT.

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.

NZ suppresses TLR4/NF-κB signalings and NLRP3 inflammasome activation in LPS-induced RAW264.7 macrophages.

OBJECTIVE: The purpose of the present study was to evaluate the potential therapeutic effects of NZ on lipopolysaccharide (LPS)-induced RAW264.7 cells and explore its underlying mechanisms. METHODS: The effect of NZ on NO generation in LPS-activated macrophage was measured by Griess assay. The concentrations of TNF-α, IL-18, IL-1ß were analyzed with ELISA kits. The LPS-induced production of reactive oxygen species (ROS) was determined by flow cytometry. The protein expressions of TLR4, NF-κB and NLRP3 signaling pathway were investigated with Western blot analysis. RESULTS: It was shown that NZ significantly reduced the production of NO and the generation of pro-inflammatory cytokines in LPS-induced RAW264.7 cells. In addition, NZ markedly inhibited the up-regulation of toll-like receptor 4 (TLR4), myeloid differentiation factor 88 (MyD88) and the activation of nuclear factor kappa B (NF-κB) in LPS-stimulated RAW 264.7 macrophages. Of note, NZ suppressed the expression of the inflammasome component such as NOD-like receptor 3(NLRP3), apoptosis-associated speck-like protein containing CARD(ASC), as well as the levels of cytokines including Interleukin-18(IL-18) and Interleukin-1ß(IL-1ß). CONCLUSION: These results indicated that NZ inhibited the generations of NO and pro-inflammatory cytokines by suppressing TLR4/MyD88/NF-κB pathway, suggesting that NZ could be an effective candidate for ameliorating LPS-induced inflammatory responses.

Enantioselective determination of (R)- and (S)-lansoprazole in human plasma by chiral liquid chromatography with mass spectrometry and its application to a stereoselective pharmacokinetic study.

A simple and enantioselective method was developed and validated for the simultaneous determination of (R)- and (S)-lansoprazole in human plasma by chiral liquid chromatography with tandem mass spectrometry. Lansoprazole enantiomers and internal standard (esomeprazole) were extracted from plasma using acetonitrile as protein precipitating agent. Baseline chiral separation was achieved within 9.0 min on a Chiralpak IC column (150 mm × 4.6 mm, 5 µm) with the column temperature of 30°C. The mobile phase consisted of 10 mM ammonium acetate solution containing 0.05% acetic acid/acetonitrile (50:50, v/v). The mass spectrometric analysis was performed using a QTrap 5500 mass spectrometer coupled with an electrospray ionization source in positive ion mode. The multiple reactions monitoring transitions of m/z 370.1→252.1 and 346.1→198.1 were used to quantify lansoprazole enantiomers and esomeprazole, respectively. For each enantiomer, no apparent matrix effect was found, the calibration curve was linear over 5.00-3000 ng/mL, the intra- and inter-day precisions were below 10.0%, and the accuracy was -3.8 to 3.3%. Analytes were stable during the study. No chiral inversion was observed during sample storage, preparation procedure and analysis. The method was applied to the stereoselective pharmacokinetic studies in human after intravenous administration of dexlansoprazole or racemic lansoprazole.

A study of the geo-herbalism of evodiae fructus based on a flow-injection mass spectrometric fingerprinting method combined with chemometrics.

A flow-injection mass spectrometric (FIMS) fingerprinting method in combination with principal component analysis (PCA) was used to study the geo-herbalism of Evodiae Fructus (EF) samples. Twenty four EF samples from different regions in China were collected and analyzed. The PCA scores plot showed that the samples from Guizhou Province were scattered in different groups, however, most of the samples from other provinces were basically scattered in the same group. Nine characteristic compounds responsible for the classification of the samples were tentatively characterized. These nine compounds might help differentiating EF samples from different regions.

Studies on Chromatographic Fingerprint and Fingerprinting Profile-Efficacy Relationship of Saxifraga stolonifera Meerb.

This work investigated the spectrum-effect relationships between high performance liquid chromatography (HPLC) fingerprints and the anti-benign prostatic hyperplasia activities of aqueous extracts from Saxifraga stolonifera. The fingerprints of S. stolonifera from various sources were established by HPLC and evaluated by similarity analysis (SA), hierarchical clustering analysis (HCA) and principal component analysis (PCA). Nine samples were obtained from these 24 batches of different origins, according to the results of SA, HCA and the common chromatographic peaks area. A testosterone-induced mouse model of benign prostatic hyperplasia (BPH) was used to establish the anti-benign prostatic hyperplasia activities of these nine S. stolonifera samples. The model was evaluated by analyzing prostatic index (PI), serum acid phosphatase (ACP) activity, concentrations of serum dihydrotestosterone (DHT), prostatic acid phosphatase (PACP) and type II 5α-reductase (SRD5A2). The spectrum-effect relationships between HPLC fingerprints and anti-benign prostatic hyperplasia activities were investigated using Grey Correlation Analysis (GRA) and partial least squares regression (PLSR). The results showed that a close correlation existed between the fingerprints and anti-benign prostatic hyperplasia activities, and peak 14 (chlorogenic acid), peak 17 (quercetin 5-O-ß-d-glucopyranoside) and peak 18 (quercetin 3-O-ß-l-rhamno-pyranoside) in the HPLC fingerprints might be the main active components against anti-benign prostatic hyperplasia. This work provides a general model for the study of spectrum-effect relationships of S. stolonifera by combing HPLC fingerprints with a testosterone-induced mouse model of BPH, which can be employed to discover the principle components of anti-benign prostatic hyperplasia bioactivity.

[Study on quality standard of Eucommia ulmoides based on absorbed active components in rat plasma].

The method was established for determination of geniposidic acid, chlorogenic acid, pinoresinoldiglucoside, which are three kinds of constituents of Eucommia ulmoides absorbed into the blood components. LC-MS/MS technique was applied to determine the blood components of the bloodstream after administration of E. ulmoides extract. At the same time, HPLC was used for detection of the ingredients content of the blood sample from 23 batches of E. ulmoides. The results showed that geniposidic acid, chlorogenic acid and pinoresinoldiglucoside are prototype into the blood in rats after oral administration of E. ulmoides extract, The linear range of geniposidic acid, chlorogenic acid and pinoresinoldiglucoside was good, and the average recoveries geniposidic acid, chlorogenic acid and pinoresinoldiglucoside were 98.69%, 100.8% and 98.39%, respectively. The method is simple and feasible with good reproducibility.

Crystal structure of Legionella pneumophila dephospho-CoA kinase reveals a non-canonical conformation of P-loop.

Dephospho-CoA kinase (DPCK; EC 2.7.1.24) catalyzes the final step in the coenzyme A biosynthetic pathway. DPCK transfers a phosphate group from ATP to the 3-hydroxyl group of the ribose of dephosphocoenzyme A (dCoA) to yield CoA and ADP. Upon the binding of ligands, large conformational changes is induced in DPCKs, as well as in many other kinases, to shield the bound ATP in their catalytic site from the futile hydrolysis by bulk water molecules. To investigate the molecular mechanisms underlying the phosphoryl transfer during DPCK catalytic cycle, we determined the crystal structures of the Legionellapneumophila DPCK (LpDPCK) both in its apo-form and in complex with ATP. The structures reveal that LpDPCK comprises of three domains, the classical core domain, the CoA domain, and the LID domain, which are packed together to create a central cavity for substrate-binding and enzymatic catalysis. The binding of ATP induces large conformational changes, including a hinge-bending motion of the CoA binding domain and the "helix to loop" conformational change of the P-loop. Finally, modeling of a dCoA molecule to the enzyme provides insights into the catalytic mechanism of DPCK.

The complete chloroplast genome of Pilea notata C. H. Wright, 1899 (Urticaceae).

Pilea notata (Pilea notata C. H. Wright_C. H. Wright, 1899) is Pilea Lindl. of Urticaceae, which is a commonly used Miao medicine in Guizhou province. The P. notata chloroplast genome is 150,979 bp, contains a pair of inverted repeats (IRs 25,743bp), and is separated by a large single-copy region (81,446bp) and a small single-copy region (18,047bp). A total of 131 genes, including 86 protein-coding genes, 37 tRNA genes, and eight rRNA genes. Phylogenetic analysis showed that P. notata, P. verrucosa and P. monilifera united as a single branch, while Pilea cadierei was defined as a sister group of this branch.

Inhibitory effect of r-hirudin variant III on streptozotocin-induced diabetic cataracts in rats.

The in vivo inhibitory effect of r-hirudin variant III (rHV3) on streptozotocin (STZ)-induced diabetic cataracts in rats was investigated. SD-rats were firstly made diabetic by a single intraperitoneal injection of 2% (W/V) STZ (65 mg/kg). Two weeks later, cataract formation was examined by slit lamp microscope, and the cataracted animals were randomly grouped. The animals in the treated groups received rHV3 drops administration to the eyes with various doses. After 4 weeks treatment, the animals were sacrificed to evaluate the biochemical changes of aldose reductase (AR), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and malondialdehyde (MDA) levels in the eye lens. Meanwhile, the cataract progression was monitored by slit lamp microscope. As a result, rHV3 drops treatment significantly increased the activities of SOD and GSH-Px in the lens in a dose-dependent manner, whereas AR activity and MDA level in the lens were dramatically decreased. Also, the morphological observation further confirmed the inhibition of the development of STZ-induced diabetic cataracts by the rHV3 drops treatment. Thus, our data suggest that rHV3 drops are pharmacologically effective for the protection against STZ-induced diabetic cataracts in rats.

[Study on quality standard of Psammosilene tunicoides based on absorbed active components in rat plasma].

A method for determintion of allantion in Psammosilene tunicoides was established by HPLC. Using alcohol as the extraction solvent, the subsequent filtrate of P. tunicoides was analysed by HPLC. Allantoin was successfully detected and separated by ZORBAX NH2 column (4.6 mm x 150 mm,5 microm) at wavelength of 220 nm and column temperature of 40 degrees C, with acetonitrile-water (93: 7) as mobile phase at a flow rate of 1.0 mL x min(-1). The results showed that it had a good linear relationship between the concent ration of allantion and chromatographic peak area. The linear correlation coefficient of allantion was 0.999 5 in 0.010 4-0.166 g x L(-1). The relative standard deviation of six parallel injections was less than 2.1%. The average recoveries were ranged from 95.47% to 100.9%. This method was sensitive and accurate for the determination of allantion in P. tunicoides.

[Antitussive constituents of Disporum cantoniense].

The antitussive activity assay for the root extraction of Disporum cantoniense was carried out with coughing mice induced by ammonia liquor. The results showed that the ethanol and water extractions of D. cantoniense possess strong antitussive activity, and the high dose of the former was better than positive control, and then the constituents of the ethanol extraction were separated and purified by various modern chromatographic techniques. Their structures were identified by physico-chemical properties and spectroscopic data. As a result, eight compounds were isolated and identified as stigmast-4-en-3-one(1), (22E, 24R)-ergosta-5, 7, 22-trien-3beta-ol(2), obtucarbamate A(3), obtucarbamate B(4), neotigogenin(5), azo-2, 2'-bis[Z-(2,3-dihydroxy-4-methyl-5-methoxy) phenyl ethylene] (6),dimethyl {[carbonylbis (azanediyl)] bis( 2-methyl-5, 1-phenylene) j dicarbamate (7) , and quercetin-3-O-pB-D-glucopyranoside(8). All compounds were isolated from this plant for the first time, and the result of bioactivity-directed isolation showed that compounds 3, 4, and 6 had obvious effect on antitussive activity, and compound 6 had the same level as positive control.

[Chemical constituents from roots of Psammosilene tunicoides].

Three compounds, including a new one (1), were isolated from the roots of Psammosilene tunicoides. Based on analysis of NMR data as well as their physical and chemical properties, their structures were elucidated as 3,3'-(4,5-dimethoxynaphthalene-2,7-diyl) bis (1-nitropropan-1-one) (1), alpha-spinasterol-3-O-beta-D-glucopyranoside (2) and alpha-spinasteryl-3-O-beta-D-glucoside-6'-O-palmitate(3). Compounds 2 and 3 were isolated from this genus for the first time.