Synthetic cajaninstilbene acid derivatives eradicate methicillin-resistant Staphylococcus aureus persisters and biofilms.

The Staphylococcus aureus can switch to a transient genotype-invariant dormancy, known as a persister, to survive treatment with high doses of antibiotics. This transient persister is an important reason underlying its resistance. There is an urgent need to find new antibacterial agents capable of eradicating methicillin-resistant S. aureus (MRSA) persisters. In this study, 37 new derivatives of cajaninstilbene acid (CSA) were designed and synthesized, and their biological activity against MRSA persisters was evaluated. Most of the newly synthesized derivatives exhibit more potent antimicrobial properties against S. aureus and MRSA than CSA itself, and 23 of the 37 derivatives show a tendency to eradicate MRSA persisters. A representative compound (A6) was demonstrated to target bacterial cell membranes. It eradicated the adherent biofilm of MRSA in a concentration dependent manner, and showed a synergistic antibacterial effect with piperacilin. In a model mouse abscess caused by MRSA persisters, A6 effectively reduced the bacterial load in vivo. These results indicate that A6 is a potential candidate for treatment of MRSA persister infections.

Cytoprotective effects of paeoniflorin are associated with translocator protein 18 kDa.

Paeoniflorin (PF) is one of the important active components in peony that are known to produce the neuroprotective effects. However, the involved cytoprotective factors on brain astrocytes are remain unclear. Translocator protein 18 kDa (TSPO) and its downstream neurosteroids biosynthesis play a significant role in cytoprotection. Based on these, the role of TSPO and neurosteroids biosynthesis in the cytoprotective effects of PF is evaluated. The astrocyte cells were cultured and AC-5216 (TSPO ligand) was selected as the positive control drug. The cytoprotective effects of PF and the levels of neurosteroids were quantified by water-soluble tetrazolium assay and enzyme linked immunosorbent assay, respectively. The cytoprotective activities of PF were relevant to neurosteroids (e.g. progsterone and allopregnanolone) biosynthesis, while these effects were totally blocked by PK11195, trilostane and finasteride, respectively. In summary, the cytoprotective effects of PF maybe mediated by TSPO and neurosteroids biosynthesis. The findings may provide the new insights into the cytoprotective effects of PF.

Anti-Inflammatory Effects of Cajaninstilbene Acid and Its Derivatives.

Cajaninstilbene acid (CSA) is one of the active components isolated from pigeon pea leaves. In this study, anti-inflammatory effects of CSA and its synthesized derivatives were fully valued with regard to their activities on the production of nitric oxide (NO) and pro-inflammatory cytokines tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) in vitro cell model, as well as their impacts on the migration of neutrophils and macrophages in fluorescent protein labeled zebrafish larvae model by live image analysis. Furthermore, the anti-inflammatory mechanism of this type of compounds was clarified by western-blot and reverse transcription-polymerase chain reaction (RT-PCR). The results showed that CSA, as well as its synthesized derivatives 5c, 5e and 5h, exhibited strong inhibition activity on the release of NO and inflammatory factor TNF-α and IL-6 in lipopolysaccharides (LPS)-stimulated murine macrophages. CSA and 5c greatly inhibited the migration of neutrophils and macrophages in injury zebrafish larvae. CSA and 5c treatment greatly inhibited the phosphorylation of proteins involved in nuclear factor kappa B (NF-κB) and mitogen-activated protein kinase (MAPK) pathways. Moreover, we found that peroxisome proliferator-activated receptor gamma (PPARγ) inhibitor GW9662 could reverse partly the roles of CSA and 5c, and CSA and 5c treatment greatly resist the decrease of PPARγ mRNA and protein induced by LPS stimulation. Our results identified the promising anti-inflammatory effects of CSA and its derivatives, which may serve as valuable anti-inflammatory lead compound. Additionally, the mechanism studies demonstrated that the anti-inflammatory activity of CSA and its derivative is associated with the inhibition of NF-κB and MAPK pathways, relying partly on resisting the LPS-induced decrease of PPARγ through improving its expression.

Novel cajaninstilbene acid derivatives as antibacterial agents.

Discovery of novel antibacterial agents with new structural scaffolds that combat drug-resistant pathogens is an urgent task. Cajaninstilbene acid, which is isolated from pigeonpea leaves, has shown antibacterial activity. In this study, a series of cajaninstilbene acid derivatives were designed and synthesized. The antibacterial activities of these compounds against gram-negative and gram-positive bacteria, as well as nine strains of methicillin-resistant staphylococcus aureus (MRSA) bacteria are evaluated，and the related structure-activity relationships are discussed. Assays suggest that some of the synthetic cajaninstilbene acid derivatives exhibit potent antibacterial activity against gram-positive bacterial strains and MRSA. Among these compounds, 5b, 5c, 5j and 5k show better antibacterial activity than the positive control compounds. The results of MTT assays illustrate the low cytotoxicity of the active compounds.

5-Benzylidene-3,4-dihalo-furan-2-one derivatives inhibit human leukemia cancer cells through suppression of NF-κB and GSK-3ß.

It has been demonstrated that PPARγ agonists effectively inhibit proliferation, metastasis as well as induce apoptosis in human cancer cell lines. In this study, twenty-two rosiglitazone analogues, 5-benzylidene-3,4- dihalo-furan-2-one derivatives, which have been identified as PPARγ agonists in our previous work, were evaluated for their antitumor effects. Among these compounds, (Z)-3,4-dibromo-5-(3-methoxy-4-((3,5,6-trimethylpyrazin-2- yl)methoxy)benzylidene)furan-2(5H)-one (6w) shows the best antitumor activity, especially against the leukemia cell line U937, resulting in significant cytotoxicity, increased apoptosis and changes in mitochondrial membrane potential. Up-regulation of pro-apoptosis-associated proteins (Bax, caspase-3 and caspase-9) and cleaved PARP as well as down-regulation of anti-apoptosis protein Bcl-2 are observed in 6w-treated U937 cells. It was shown that the antitumor effect of 6w stems from its ability to inhibit the PPARγ-dependent expression of NF-κB and GSK-3ß.

Design, synthesis and anti-inflammatory evaluation of novel 5-benzylidene-3,4-dihalo-furan-2-one derivatives.

Rosiglitazone has shown promising anti-inflammation effect. To develop preferable anti-inflammatory agents, twenty-two rosiglitazone analogs were synthesized and their anti-inflammatory activity was evaluated. Among these compounds, 6i and 6k displayed excellent inhibitory activities on the production of inflammatory mediators, including nitric oxide (NO), tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6). Furthermore, 6i and 6k showed suppression effects on the nuclear factor-kappa B (NF-κB) and mitogen-activated protein kinase (MAPK) pathways, and this suppression effects could be partially reversed by GW9662, which is a peroxisome proliferator-activated receptor γ (PPARγ) antagonist. Additionally, our docking results exhibited the well combination of 6i and 6k to PPARγ. So the anti-inflammation activity of 6i and 6k was due at least in part, to their interaction with PPARγ.

[Metabolomic study of the action mechanism of nourishing blood effect of fo-shou-san on blood deficiency mice].

The metabolic effect of Fo-Shou-San on blood deficiency mice was studied by using metabolomic method. UPLC-QTOF/MS was used to analyze the plasma metabolome in blood deficiency mice. MS data were processed by MarkerLynx software. With multivariate statistical analysis of plasma metabolite profiles, a clear separation among control, blood deficiency model, and Fo-Shou-San groups was achieved. Potential biomarkers were selected according to the parameters of variable importance in the projection (VIP) and identified according to MS information and database retrieval. The metabolic network of blood deficiency was predicted via MetPA database. Twenty-two potential biomarkers were identified and used to explain the thiamine metabolism, arachidonic acid metabolism, sphingolipid metabolism, glyoxylate and dicarboxylate metabolism, histidine metabolism, nicotinate and nicotinamide metabolism, cysteine and methionine metabolism, tryptophan metabolism, starch and sucrose metabolism, tyrosine metabolism and citrate cycle (TCA cycle). Those metabolic pathways were disturbed in blood deficiency mice, but which could be regulated nearly to normal state after Fo-Shou-San administration. In this study, the metabolomics of blood deficiency mice and the action mechanism of nourishing blood effect of Fo-Shou-San were evaluated. The physiological and metabolic state of the organism could be represented comprehensively by using metabolomics. And metabolomics can be used to evaluate the pharmacodynamics and related mechanisms of Chinese medicine and formulae.

[Research on nourishing and tonifying blood effects of the herb pair consisting of Angelica sinensis and Ligusticum chuanxiong on the basis of drug interaction].

OBJECTIVE: To study the pharmacodynamic interaction of nourishing and tonifying blood effects of the herb pair consisting of Angelica sinensis and Ligusticum chuanxiong by response surface method. METHODS: The blood deficiency rat model was induced by injecting N-acetylphenylhydrazine and cyclophosphamide. The effects of Angelica sinensis and Ligusticum chuanxiong in different proportions (0:1, 1:5, 2:5, 2:3, 1:1, 3:2, 5:2, 5:1, and 1:0) and at different concentrations on the peripheral blood index and the organ indices were observed. Then all indices were integrated to the total nourishing effect value by comprehensive index method. The interaction was analyzed by response surface method. The model parameters were estimated with nonlinear regression. The three-dimensional response surfaces were constructed with Matlab Software. RESULTS: In the response surface, most compatibility of Angelica sinensis and Ligusticum chuanxiong showed synergistic action, some showed addition action, and few of them showed obvious antagonist action. The proportion of Angelica sinensis and Ligusticum chuanxiong from 4:1 to 2:1 and the dose of Angelica sinensis and Ligusticum chuanxiong from low to high showed addition action, while the other proportions showed obvious addition action at low dose and synergistic action at high dose. CONCLUSIONS: The research results could provide scientific evidence for reasonable application of Angelica sinensis and Ligusticum chuanxiong in clinics of Chinese medicine. The quantitative analysis on drug interactions of herbal compatibility by response surface method could provide reference for relative studies.

[Study on antioxidant interaction of different preparations and proportions of Danggui-Chuanxiong drug pair].

OBJECTIVE: To observe the in vitro antioxidant interaction of different preparations and proportions of Danggui-Chuanxiong drug pair in the DPPH free radical scavenging rate with the response surface methodology. METHOD: The 2,2-diphenyl-1-picryl-hydrazyl (DPPH) free radical scavenging rate method was adopted for determining the antioxidant activity of extracts from Danggui-Chuanxiong with 10 proportions and three extraction processes. The response surface methodology was used to determine the parameters of the dose-effect curve and establish a three-dimensional response surface model. The three-dimensional response surface graph was constructed with Matlab software. RESULT: All of the 30 samples with different proportions and preparations had antioxidant effect in scavenging free radicals and a remarkable dose-effect relationship. Their water extracts had a narrow synergistic range, with only spot distribution. Their antagonist ranges were districted in six bands of various widths. The synergistic ranges of ethanol extracts were districted in small bands, with the antagonist ranges scattered in points. The synergistic ranges of their water-alcohol extracts were distributed in three bands, with their antagonist ranges scattered in points. In short, the water-alcohol extracts showed a wider synergistic range than ethanol extracts, followed by water-extracts. All of the three extraction processes showed no obvious synergistic and antagonist effects. CONCLUSION: The quantitative study on the interaction of traditional Chinese medicines with different compatibilities with the response surface methodology provides reference of thoughts and methods for relevant studies.

[Evaluate activating blood circulation and dissipating blood stasis effect mechanism of foshousan on acute blood stasis rats on basis of metabolomic approach].

Ice water bath and subcutaneous injection of adrenaline were used to establish the acute blood stasis model of rats. Ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS) was used to study the urine metabolic changes of acute blood stasis rats. Potential biomarkers were selected by variable importance projection, and identified on basis of MS information and databases. The metabolic pathways were predicted via MetPA database. To study the effect of Foshousan on endogenous metabolites of acute blood stasis model rats, find potential biomarkers, and explore the effect mechanism of Foshousan on activating blood circulation and dissipating blood stasis. Eleven potential biomarkers were identified with multivariate statistical analysis of urine metabolite profiles, and which also were used to explain the phenylalanine metabolism, tryptophan metabolism and sphingolipid metabolism. Those disturbed metabolic pathways in acute blood stasis rats could be regulated closely to normal state after Foshousan administration. Metabolomics has a bright prospect in the efficacy evaluation and effect mechanism elucidation of the traditional Chinese medicines.

[Research on Chinese medicine pairs (VII)--Angelicae sinensis radix-chuanxiong rhizoma].

Angelicae Sinensis Radix-Chuanxiong Rhizoma has the effects of nourishing and tonifying blood, activating blood and dissipating blood stasis, regulating menstruation and analgetic, which is commonly used Chinese medicine pair (CMP) in traditional Chinese medicine (TCM) clinic. It might be an independent formula, and is also usually used in many gynecological formulae and modern TCM prescriptions. This paper mainly analyzed and summarized the compatibility theory, bio-active constituents, compatibility effects and action mechanism, and clinical applications of the CMP, which can provide a basis for the depth research and development of the CMP.

[Analysis on correlation between general efficacy and chemical constituents of danggui-chuanxiong herb pair based on artificial neural network].

OBJECTIVE: To evaluate the effect of danggui-chuanxiong herb pair with different prescription proportions and formulas on nourishing and tonifying blood (NTB), activating blood circulation and dissolving blood stasis (ADBS), regulating menstruation and relieving pain (RMRP) in an all-round way, in order to reveal the correlation between the effect and constituents and their degrees. METHOD: First, the indexes of effect were standardized. All effects were integrated by using multi-index aggregative index method. The weight coefficients of each index were worked out by the combining the statistics of testing frequency of each effect index given by the specialist-scored method and in relevant literatures released in the latest 10 years. Then, the total effect values were obtained by multiplying the standardized value of each index with the weight coefficients and then adding their results together. Finally, a fitting analysis was made on the contents of seven major active components in danggui-chuanxiong herb pair, total aromatic acids and total phthalide lactones and their total effect by using the artificial neural network, respectively. RESULT: Water extracts of danggui-chuanxiong (1.5:1) had the best effect on total NTB; alcohol extracts of danggui-chuanxiong herb pair (1:1) had the best effect on total ADBS; alcohol extraction of danggui-chuanxiong herb pair (1.5:1) had the best effect on RMRP. By using the same extraction method, extracts of danggui had better effect on total NTB than Chuanxiong; while extracts of chuanxiong had better effect on total ADBS and RMRP than danggui. With the same prescription proportion, water extracts of danggui-chuanxiong herb pair had the best effect on total NTB; alcohol extracts of danggui-chuanxiong herb pair had the best effect on total ADBS and RMRP. Aromatic acids were the main effective components for NTB, especially chlorogenic acid and caffeic acid. Both aromatic acids and phthalide lactones were active components for ADBS, particular chlorogenic acid and senkyunolides I and H showed greater effect on ADBS. Both aromatic acids and phthalide lactones were also the important active constituents for the effect of RMRP, especially ligustilide, caffeic acid, ferulic acid, and senkyunolide I showed greater effect on RMRP. CONCLUSION: Both aromatic acids and phthalide lactones of danggui-chuanxiong herb pair contributed to NTB, ADBS and RMRP. Aromatic acids laid particular stress on NTB and ADBS, while phthalide lactones laid particular stress on RMRP. In this study, determined effect indexes of the herbal medicines were integrated by using multi-index aggregative index method, and a fitting correlation analysis was made on the component content of the herbal medicines and their integration effect by using the artificial neural network, in order to specify the active constituents of the herbal medicines with different effects and their contributions to the general efficacy and provide new ideas and methods for basic study on complicated active constituents of the herbs medicines.

[Comparison of in vitro anti-oxidative activities among Siwu Decoction Serial Recipes, their composed crude herbs, and main aromatic acids, as well as their dose-effect correlation].

OBJECTIVE: To assess and compare the in vitro anti-oxidative activities among Siwu Decoction Serial Recipes, their composed crude herbs, and main aromatic acids they contained. METHODS: Their anti-oxidative activities (including Siwu Decoction and correlated recipes such as Taohong Siwu Decoction, Xiangfu Siwu Decoction, Shaofu Zhuyu Decoction, and Xiongqiong Decoction, 16 kinds of crude herbs, and main aromatic acids they contained) were determined using 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical clearance method. RESULTS: The selected five decoctions showed obvious activities of scavenging free radicals. Siwu Decoction was better than other decoctions in scavenging free radicals and Xiongqiong Decoction was the least. Among the 16 kinds of crude herbs, red peony root, white peony root, safflower, ligustici chuanxiong, common aucklandia root showed the strongest activities, while peach seed showed the poorest activities. Among aromatic acids, gallic acid, protocatechuic acid, vanillic acid, caffeic acid, chlorogenic acid, p-coumaric acid, and ferulic acid showed obvious anti-oxidative activities in scavenging free radicals, showing obvious dose-effect correlation. p-hydroxybenzoic acid, benzoic acid, and cinnamic acid showed no activities on scavenging free radicals (P > 0.05). CONCLUSION: Siwu Decoction and aromatic acids contained in correlated decoctions played significant roles in anti-oxidative activities.

[Interaction of nourishing and tonifying blood effects of the combination of Angelicae sinensis Radix and Astragali Radix studied by response surface method].

The combination of Angelicae sinensis Radix (Danggui, DG) and Astragali Radix (Huangqi, HQ) is a popular herb pair commonly used in clinic for the treatment of blood deficiency syndrome in China. The aim of this paper is to study the interaction of DG and HQ nourishing and tonifying blood effects by response surface method. The blood deficiency mice were induced by injecting N-acetylphenylhydrazine (sc) and cyclophosphamide (ip). The blood deficiency mice were administrated intragastrically with DG-HQ extracts (0:1, 1: 5, 2:5, 2:3, 1:1, 3:2, 5:2, 5:1, 1:0). The changes of the peripheral blood indexes and organ indexes were observed. The indexes were integrated by comprehensive index method; the interactions of DG and HQ were analyzed by the response surface diagram established with Matlab software. The results showed that DG and HQ at most of their combination ratios had synergic effect. Within the range of 1:5 - 5:1, all of the extracts of DG-HQ showed synergic effect, and among which, high-doses had better effects than low-doses. The highest value (-1) of the synergic effect was showed when DG was 10 - 40 g at the same time of HQ as 90 -180 g, and DG was 50 - 100 g at the same time of HQ as 20 - 100 g. DG-HQ at all combination dosages within Chinese Pharmacopeia (DG: 6 - 12 g, HQ: 9 - 30 g) had certain synergic effect, and Danggui Buxue Decoction (DG: 6 g, HQ: 30 g) also was at this range. The results provided scientific basis to the clinical application of DG and HQ. And the response surface method was firstly applied to quantitatively evaluate the bio-activity change of herb combination, which provided a novel way for modern basic research on the interaction of herbs.

[Detection for alpha-glucosidase-inhibiting polyhydroxyalkaloid ingredients of Commelina communis L. by ESIMS].

For detecting polyhydroxyalkaloids-type alpha-glucosidase-inhibiting ingredients of Commelina communis L grown in China, total alkaloids were obtained from the plant by extraction with water, removal of precipitation after the addition of alcohol, enrichment and purification by ion exchange resin and sephadex LH 20 chromatography. Polyhydroxyalkaloids in the total alkaloids were detected by ion trap electron-spray ionization mass spectra (ESIMS). Several reported and unreported polyhydroxyalkaloids in the plant were detected from the material collected from Jixi county, Anhui province. The crude drug growing in China contains alpha-glucosidase-inhibiting polyhydroxyalkaloids and can be used to therapy in diabetes.

[Alkaloid constituents from silkworm dropping of Bombyx mori].

OBJECTIVE: To extract and isolate chemical constituents from the total alkaloids of silkworm dropping (Can Sha) of Bombyx mori L. METHODS: Chemical constituents were isolated by column chromatography with macroporous adsorbent resin, ion-exchange resin and sephadex LH 20. The structures of the isolated compounds were determined by spectral means. RESULTS: Three compounds were isolated and identified as 1-deoxynojirimycin (1) , fagomine (2) , and 3-epifagomine (3) on the basis of their 1H-NMR, 13C-NMR spectra and ESIMS data. CONCLUSION: Compounds 1-3 are isolated and identified from Can Sha for the first time.