Enhanced Production of Nitrogenated Metabolites with Anticancer Potential in Aristolochia manshuriensis Hairy Root Cultures.

Aristolochia manshuriensis is a relic liana, which is widely used in traditional Chinese herbal medicine and is endemic to the Manchurian floristic region. Since this plant is rare and slow-growing, alternative sources of its valuable compounds could be explored. Herein, we established hairy root cultures of A. manshuriensis transformed with Agrobacterium rhizogenes root oncogenic loci (rol)B and rolC genes. The accumulation of nitrogenous secondary metabolites significantly improved in transgenic cell cultures. Specifically, the production of magnoflorine reached up to 5.72 mg/g of dry weight, which is 5.8 times higher than the control calli and 1.7 times higher than in wild-growing liana. Simultaneously, the amounts of aristolochic acids I and II, responsible for the toxicity of Aristolochia species, decreased by more than 10 fold. Consequently, the hairy root extracts demonstrated pronounced cytotoxicity against human glioblastoma cells (U-87 MG), cervical cancer cells (HeLa CCL-2), and colon carcinoma (RKO) cells. However, they did not exhibit significant activity against triple-negative breast cancer cells (MDA-MB-231). Our findings suggest that hairy root cultures of A. manshuriensis could be considered for the rational production of valuable A. manshuriensis compounds by the modification of secondary metabolism.

Magnoflorine improves cognitive deficits and pathology of Alzheimer's disease via inhibiting of JNK signaling pathway.

BACKGROUND: Cognitive deficit is the main clinical feature of Alzheimer's disease (AD), and the massive death of neuronal cells is the leading cause of cognitive deficits. So, there is an urgent clinical need to discover effective drugs to protect brain neurons from damage in order to treat AD. Naturally-derived compounds have always been an important source of new drug discovery because of their diverse pharmacological activities, reliable efficacy and low toxicity. Magnoflorine is a quaternary aporphine alkaloid, which naturally exist in some commonly used herbal medicines, and has good anti-inflammatory and antioxidant effects. However, magnoflorine has not been reported in AD. HYPOTHESIS/PURPOSE: To investigate the therapeutic effect and mechanism of magnoflorine on AD. METHODS: Neuronal damage was detected by flow cytometry, immunofluorescence and western blotting. Oxidative stress was measured by detection of SOD and MDA, as well as JC-1 and reactive oxygen species (ROS) staining. The APP/PS1 mice were given drugs by intraperitoneal injection (I.P.) every day for one month, and then the new object recognition and Morris water maze were used to detect the cognitive ability of the mice. RESULTS: We demonstrated that magnoflorine reduced Aß-induced PC12 cell apoptosis and intracellular ROS generation. Further studies found that magnoflorine significantly improved cognitive deficits and AD-type pathology. Most interestingly, the efficacy of magnoflorine was better than that of the clinical control drug donepezil. Mechanistically, based on RNA-sequencing analysis, we found that magnoflorine significantly inhibited phosphorylated c-Jun N-terminal kinase (JNK) in AD models. This result was further validated using a JNK inhibitor. CONCLUSION: Our results indicate that magnoflorine improves cognitive deficits and pathology of AD through inhibiting of JNK signaling pathway. Thus, magnoflorine may be a potential therapeutic candidate for AD.

A Systematic Method for the Identification of Aporphine Alkaloid Constituents in Sabia schumanniana Diels Using UHPLC-Q-Exactive Orbitrap/Mass Spectrometry

Sabia schumanniana Diels (SSD) is a plant whose stems are used in traditional folk medicine for the treatment of lumbago and arthralgia. Previous studies have revealed chemical constituents of SSD, including triterpenoids and aporphine alkaloids. Aporphine alkaloids contain a variety of active components, which might facilitate the effective treatment of lumbago and arthralgia. However, only 5-oxoaporphine (fuseine) has been discovered in SSD to date. In this study, we sought to systematically identify the aporphine alkaloids in SSD. We established a fast and reliable method for the detection and identification of these aporphine alkaloids based on ultra-high-performance liquid chromatography (UHPLC)-Q-Exactive-Orbitrap/mass spectrometry combined with parallel reaction monitoring (PRM). We separated all of the analyzed samples using a Thermo Scientific Hypersil GOLD™ aQ C18 column (100 mm × 2.1 mm, 1.9 µm). Finally, we identified a total of 70 compounds by using data such as retention times and diagnostic ions. No fewer than 69 of these SSD aporphine alkaloids have been reported here for the first time. These findings may assist in future studies concerning this plant and will ultimately contribute to the research and development of new drugs.

[Influence of coexisting components in Coptidis Rhizoma on excretion of magnoflorine in rat bile, urine, and feces].

Magnoflorine is an important aporphine alkaloid in Coptidis Rhizoma. As reported previously, coexisting components in Coptidis Rhizoma can change the pharmacokinetic characteristics of magnoflorine. To illustrate the interactional links of magnoflorine with its coexisting components in Coptidis Rhizoma, the present study investigated the influence of coexisting components in Coptidis Rhizoma on the excretion of magnoflorine in rat bile, urine, and feces. The rats were dosed with magnoflorine(30 mg·kg~(-1)) and water decoction of Coptidis Rhizoma(equivalent to 30 mg·kg~(-1) magnoflorine) via intragastric administration, and magnoflorine(10 mg·kg~(-1)) by intravenous administration, respectively, and the excretion of magnoflorine in rat bile, urine, and feces in 24 h was observed. The excretion rates of magnoflorine in bile and urine in 24 h were 0.90% and 37.11% respectively after intravenous administration of magnoflorine, which suggested that urination was the main excretive way of magnoflorine. The excretion rates of magnoflorine in feces were 8.77% and 6.18% respectively after intragastric administration of magnoflorine and water decoction of Coptidis Rhizoma, which indicated that defecation was the main excretion route of magnoflorine. The cumulative excretion rates of magnoflorine in the bile, urine, and feces in the Coptidis Rhizoma water decoction group were 77.78%, 79.44%, and 70.47% of those in the magnoflorine group. The results showed that the cumulative excretion rates of magnoflorine in rat bile, urine, and feces were not high, suggesting that magnoflorine was metabolized significantly in rats. The coexisting components of Coptidis Rhizoma could inhibit the excretion of magnoflorine in rat bile, urine, and feces, which was consistent with the decrease in the elimination rate of magnoflorine in the pharmacokinetics of Coptidis Rhizoma water decoction. It indicated interactions between drugs. This study is expected to provide references for the development of magnoflorine-containing new drugs and rational clinical medication of Coptidis Rhizoma.

Magnoflorine attenuates inflammatory responses in RA by regulating the PI3K/Akt/NF-κB and Keap1-Nrf2/HO-1 signalling pathways in vivo and in vitro.

BACKGROUND: As a prolonged autoimmune disorder, rheumatoid arthritis (RA) is characterised by synovial hyperplasia and the erosion of bone and cartilage. Magnoflorine (MAG) is the main component purified from Clematis manshurica Rupr. Recent studies have shown that MAG has anti-inflammatory, antioxidant, and immunosuppressive effects, which are relevant to anti-RA activities. OBJECTIVE: The current investigation was conducted to explore the anti-RA effects of MAG and to discover the possible molecular mechanisms. METHODS: In vitro experiments, CCK-8, wound healing, and transwell assays were utilized to evaluate the anti-proliferative, anti-migratory, and anti-invasive activities of MAG, respectively. The rate of cell distribution and cell apoptosis were evaluated by flow cytometry. ROS generation was detected by DCFH-DA staining. Western blotting, quantitative real-time polymerase chain reaction assay, and immunofluorescent staining were employed to test the anti-RA effect of MAG as well as to explore the potential mechanisms by evaluating related gene and protein expression. For in vivo experiments, an adjuvant-induced arthritis (AIA) rat model was established. The related parameters were measured in rats. Then, rats were sacrificed, and ankle joints were collected for histopathological analysis and observation. RESULTS: MAG significantly decreased the proliferation, migration, invasion, and reactive oxygen species levels in IL-1ß-treated MH7A cells. Furthermore, MAG promoted cell apoptosis by increasing Bax levels and decreasing Bcl-2 levels. MAG also induced cell cycle arrest. Inflammatory cytokines (iNOS, COX-2, IL-6, and IL-8) and MMPs (MMP-1, 2, 3, 9, and 13) were reduced by MAG treatment. Molecular analysis revealed that MAG exerted anti-RA effects by partly inhibiting the PI3K/Akt/NF-κB signalling axis and activating the Keap1-Nrf2/HO-1 signalling pathway. In vivo studies have revealed that MAG treatment substantially improved severe symptoms in AIA rats, and these curative effects were linked to the attenuation of inflammatory responses. CONCLUSION: These results first suggested that MAG exhibits anti-arthritic effects in IL-1ß-treated MH7A cells and AIA rat models. Thus, MAG may be used as a new drug to treat RA clinically.

Metabolic characteristics and pharmacokinetic differences of Qiwei Tongbi oral liquid in rheumatoid arthritis rats.

Qiwei Tongbi oral liquid (QWTB), a classical traditional Chinese medicine formula, is widely used to treat arthritis-related diseases in clinical practice. Currently, in vivo metabolic characteristics and pharmacokinetic studies are lacking. This study analyzed the prototype components of QWTB absorbed in the blood and their metabolic transformation process after intragastric administration and compared the differences in pharmacokinetic properties between healthy and rheumatoid arthritis model rats. In sum, 17 prototype components and 21 related metabolites were identified in the plasma and urine of the treated rats. Metabolites were derived from sinomenine and magnoflorine. Through systematic methodology verification, an accurate and stable detection method for sinomenine and magnoflorine in plasma samples was established and applied to pharmacokinetic research of QWTB. At the three dose levels, the AUC0-∞ (area under the curve) of the two components showed a good positive correlation with the dose (R2 > 0.9). Compared with healthy rats, the Tmax , t1/2z , and AUC of sinomenine were markedly increased, and Cmax was decreased in rheumatoid arthritis model rats, indicating that the rate of absorption and elimination rate decreased, but the body exposure increased. However, there were no significant differences in the pharmacokinetic parameters of magnoflorine under healthy and pathological conditions. In summary, the main active ingredients of QWTB are sinomenine and magnoflorine, which exhibit linear kinetic characteristics within a set dose range, and the rheumatoid arthritis pathological state is more conducive to the absorption and efficacy of sinomenine. The results of this study demonstrate the rationality of the clinical application of the QWTB.

Natural isoquinoline alkaloids: Pharmacological features and multi-target potential for complex diseases.

Complex diseases such as neurodegenerative disorders and cancer constitute a growing public health problem due to the rising incidence and lack in effective therapies. Since pharmacotherapy based on a single target has been insufficient for drug development in complex diseases, the emerging multi-target approach is a promising strategy for the search of new drug candidates. Plant-derived isoquinoline alkaloids comprise a vast source of multimodal agents with unique structural diversity, and variated range of pharmacological activities. This review offers an exhaustive compilation of the pharmacological relevance and multi-target potential of natural isoquinolines, emphasizing their features and promising activity in complex diseases such as Alzheimer, Parkinson, and Cancer. Selected examples were discussed in depth to illustrate the most relevant structural motifs and their possible relationship with the multimodal activity offering a comprehensive baseline in the search and optimization of isoquinoline scaffolds with polypharmacological potential for complex diseases.

Amelioration of diabetic retinopathy in db/db mice by treatment with different proportional three active ingredients from Tibetan medicine Berberis dictyophylla F.

ETHNOPHARMACOLOGICAL RELEVANCE: Berberis dictyophylla F., a famous Tibetan medicine, has been used to prevent and treat diabetic retinopathy (DR) for thousands of years in clinic. However, its underlying mechanisms remain unclear. AIM OF THE STUDY: The present study was designed to probe the synergistic protection and involved mechanisms of berberine, magnoflorine and berbamine from Berberis dictyophylla F. on the spontaneous retinal damage of db/db mice. MATERIALS AND METHODS: The 14-week spontaneous model of DR in db/db mice were randomly divided into eight groups: model group, calcium dobesilate (CaDob, 0.23 g/kg) group and groups 1-6 (different proportional three active ingredients from Berberis dictyophylla F.). All mice were intragastrically administrated for a continuous 12 weeks. Body weight and fasting blood glucose (FBG) were recorded and measured. Hematoxylin-eosin and periodic acid-Schiff (PAS) stainings were employed to evaluate the pathological changes and abnormal angiogenesis of the retina. ELISA was performed to assess the levels of IL-6, HIF-1α and VEGF in the serum. Immunofluorescent staining was applied to detect the protein levels of CD31, VEGF, p-p38, p-JNK, p-ERK and NF-κB in retina. In addition, mRNA expression levels of VEGF, Bax and Bcl-2 in the retina were monitored by qRT-PCR analysis. RESULTS: Treatment with different proportional three active ingredients exerted no significant effect on the weight, but decreased the FBG, increased the number of retinal ganglionic cells and restored internal limiting membrane. The results of PAS staining demonstrated that the drug treatment decreased the ratio of endothelial cells to pericytes while thinned the basal membrane of retinal vessels. Moreover, these different proportional active ingredients can markedly downregulate the protein levels of retinal CD31 and VEGF, and serum HIF-1α and VEGF. The gene expression of retinal VEGF was also suppressed. The levels of retinal p-p38, p-JNK and p-ERK proteins were decreased by drug treatment. Finally, drug treatment reversed the proinflammatory factors of retinal NF-κB and serum IL-6, and proapoptotic Bax gene expression, while increased antiapoptotic Bcl-2 gene expression. CONCLUSIONS: These results indicated that DR in db/db mice can be ameliorated by treatment with different proportional three active ingredients from Berberis dictyophylla F. The potential vascular protection mechanisms may be involved in inhibiting the phosphorylation of the MAPK signaling pathway, thus decreasing inflammatory and apoptotic events.

Magnoflorine prevent the skeletal muscle atrophy via Akt/mTOR/FoxO signal pathway and increase slow-MyHC production in streptozotocin-induced diabetic rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Tinospora cordifolia (TC) is being used as a blood purifier in Ayurveda since ancient time. It is a very popular immunomodulator and holds anti-inflammatory and anti-oxidative potential, hence anti-aging properties. Therefore, it is also known as 'Amrita' in Ayurveda and is widely used to treat diabetes mellitus type II (T2DM) and its secondary complications; however, its underlying mechanism was not expedited to date. AIM-: To explore the in vivo therapeutic efficiency and mechanism of action of TC and its secondary constitute magnoflorine on the skeletal muscle atrophy in the rat model of T2DM. METHOD: Animal model of T2DM was developed using streptozotocin (STZ) injection followed by intervention with TC, metformin, and magnoflorine for three weeks. Confirmation of T2DM and abrogation of atrophic markers and possible mechanisms on supplementation of TC and magnoflorine were explored using histology, bio-assays, Western blotting, and q-PCR. RESULT: TC and Magnoflorine supplementations significantly (p ≤ 0.05) decreased the fasting blood glucose (FBG) levels in T2DM rats. Both treatments prevented the lean body, individual skeletal muscle mass, and myotubes diameter loss (p ≤ 0.05). Magnoflorine significantly reduced the degradation of the protein indicated by biochemical markers of atrophy i.e. decreased serum creatine kinase (CK) levels and increased myosin heavy chain-ß (MyHC-ß) levels in muscles. Q-PCR and western blotting supported the findings that magnoflorine significantly increased the mRNA and protein abundances (~3 fold) of MyHC-ß.TC and magnoflorine efficiently decreased the expression of ubiquitin-proteasomal E3-ligases (Fn-14/TWEAK, MuRF1, and Atrogin 1), autophagy (Bcl-2/LC3B), and caspase related genes along with calpains activities in T2DM rats. Both TC and magnoflorine also increased the activity of superoxide dismutase, GSH-Px, decreased the activities of ß-glucuronidase, LPO, and prevented any alteration in the catalase activity. In contrast, magnoflorine increased expression of TNF-α and IL-6 whereas TC and metformin efficiently decreased the levels of these pro-inflammatory cytokines (p ≤ 0.05). However, magnoflorine was found to increase phosphorylation of Akt more efficiently than TC and metformin. CONCLUSION: TC, and magnoflorine are found to be effective to control fasting blood glucose levels significantly in T2DM rats. It also promoted the Akt phosphorylation, suppressed autophagy and proteolysis that might be related to blood glucose-lowering efficacy of magnoflorine and TC. However, increased muscle weight, specifically of the soleus muscle, expression of IL-6, and slow MyHC indicated the increased myogenesis in response to magnoflorine and independent from its hypoglycemic activity.

Magnoflorine-Isolation and the Anticancer Potential against NCI-H1299 Lung, MDA-MB-468 Breast, T98G Glioma, and TE671 Rhabdomyosarcoma Cancer Cells.

Magnoflorine (MGN) is a quaternary aporphine alkaloid that exhibits numerous therapeutic properties, including neuropsychopharmacological, anti-anxiety, immunomodulatory, anti-inflammatory, antioxidant, or antifungal activities. The aim of the present study was an investigation of the influence of MGN on viability, proliferation, induction of apoptosis, and cell cycle arrest in NCI-H1299 lung, MDA-MB-468 breast, T98G glioma, and TE671 rhabdomyosarcoma cancer cells. MGN was isolated from the roots of Berberis cretica L. by counter-current partition chromatography (CPC). Cell viability and proliferation assessments were performed by means of MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and 5-bromo-2'-deoxyuridine (BrDU) assays, respectively. The induction of apoptosis and cell cycle progression was measured using fluorescence-activated cell sorting analysis. MGN in high doses inhibits proliferation, induces apoptosis, and inhibits cell cycle in S/G2 phases in a dose-dependent manner. MGN seems to be a promising anti-cancer compound in therapy of some types of lung, breast, glioma, and rhabdomyosarcoma cancers, for which current standard therapies are limited or have severe strong side effects.

Magnoflorine inhibits human gastric cancer progression by inducing autophagy, apoptosis and cell cycle arrest by JNK activation regulated by ROS.

The antitumor effect of magnoflorine (Mag), an alkaloid isolated from Coptidis Rhizoma, in gastric cancer (GC) cells has not been reported. In the study, Mag suppressed the proliferation of GC cells, but showed no influence on normal gastric cells. Mechanistically, Mag induced autophagy in GC cells, as evidenced by the up-regulated expression of LC3B-II and increased autophagosome formation. Furthermore, we found that Mag-triggered autophagic cell death was regulated by reactive oxygen species (ROS)-induced suppression of serine/threonine-protein kinases (AKT) signaling. What's more, Mag treatment led to apoptosis in GC cells through enhancing cleaved Caspase-3 and PARP expressions. In addition, up-regulated expression of p27 and p21, as well as down-regulated expression of Cyclin-A and Cyclin-B1 was detected in Mag-treated GC cells, contributing to the S/G2 cell cycle arrest. Importantly, Mag incubation resulted in a significant increase in jun N-terminal kinase (JNK) phosphorylation but not p38 and ERK1/2, which was involved in the modulation of apoptosis and S/G2 phase arrest. Moreover, ROS production was highly induced by Mag treatment, and Mag-exhibited these functions was largely dependent on the generation of ROS in GC cells. Consistently, the GC cell xenograft mouse model confirmed the anti-tumor role of Mag in vivo. Collectively, these results indicated that Mag showed anti-GC effects, which could be a potential therapeutic target for GC treatment.

Advances in Chemistry and Bioactivity of Magnoflorine and Magnoflorine-Containing Extracts.

The review collects together some recent information on the identity and pharmacological properties of magnoflorine, a quaternary aporphine alkaloid, that is widely distributed within the representatives of several botanical families like Berberidaceae, Magnoliaceae, Papaveraceae, or Menispermaceae. Several findings published in the scientific publications mention its application in the treatment of a wide spectrum of diseases including inflammatory ones, allergies, hypertension, osteoporosis, bacterial, viral and fungal infections, and some civilization diseases like cancer, obesity, diabetes, dementia, or depression. The pharmacokinetics and perspectives on its introduction to therapeutic strategies will also be discussed.

Development of a chemical fingerprint as a tool to distinguish closely related Tinospora species and quantitation of marker compounds.

Tinospora species are morphologically similar. Several cases of human toxicity have been reported in association with T. crispa. A chemical fingerprint was developed to differentiate T. crispa from its closely related species and to quantitate its major furanoditerpenes namely as borapetosides B, C and F. The rapid, sensitive and repeatable method was established using ultra-high performance liquid chromatography coupled with photodiode array and single quadrupole electrospray mass spectrometry detectors using a flavonoid, two alkaloids, an amide and six diterpenoids. Qualitative and quantitative determination was performed by UHPLC-UV and confirmed by MS. The intra-day RSD for replicates was between 0.9 and 6.8% and inter-day RSD was between 1.2 and 9.1%. Recovery was 97-103 %. The method is useful to achieve decisiveness in not only identifying but also differentiating T. crispa from T. sinensis and other closely related Tinospora species. Seventeen Tinospora plant samples and seventeen dietary supplements claiming T. crispa, T. sinensis and T. cordifolia were analyzed. The newly developed and validated method successfully resulted in the conclusive identification of two dietary supplements to be mislabeled.

Profiling and Pharmacokinetic Studies of Alkaloids in Rats After Oral Administration of Zanthoxylum nitidum Decoction by UPLC-Q-TOF-MS/MS and HPLC-MS/MS.

Zanthoxylum nitidum (Roxb.) DC (Rutaceae), called as "liangmianzhen" in China, is well known for its anti-inflammation and analgesic effect. Alkaloids are its main active constituents. However, little has been known about the absorption of main alkaloids in vivo. In this study, an ultra-performance liquid chromatography coupled with quadrupole-time-of-flight mass spectrometry was employed for identification of absorbed alkaloids in rats after oral administration of Z. nitidum decoction. By analyzing the fragmentation patterns, a total of nineteen alkaloids were exactly or tentatively identified in rat plasma after treatment, of which magnoflorine, α-allocryptopine, and skimmianine are dominant. Moreover, a high performance liquid chromatography coupled mass spectrometry method was developed for simultaneous quantification of magnoflorine, α-allocryptopine, and skimmianine, and successfully applied to pharmacokinetic study in rats after oral administration of Z. nitidum decoction. The research would contribute to comprehensive understanding of the material basis and function mechanism of Z. nitidum decoction.

Antifungal activity of magnoflorine against Candida strains.

Candida albicans is a major invasive pathogen, and the development of strains resistant to conventional antifungal agents has been reported in recent years. We evaluated the antifungal activity of 44 compounds against Candida strains. Magnoflorine showed the highest growth inhibitory activity of the tested Candida strains, with a minimum inhibitory concentration (MIC) of 50 µg/mL based on microdilution antifungal susceptibility testing. Disk diffusion assay confirmed the antifungal activity of magnoflorine and revealed that this activity was stable over 3 days compared to those of berberine and cinnamaldehyde. Cytotoxicity testing showed that magnoflorine could potentially be used in a clinical setting because it didn't have any toxicity to HaCaT cells even in 200 µg/mL of treatment. Magnoflorine at 50 µg/mL inhibited 55.91 ± 7.17% of alpha-glucosidase activity which is required for normal cell wall composition and virulence of Candida albicans. Magnoflorine also reduced the formation of C. albicans' biofilm. Combined treatment with magnoflorine and miconazole decreased the amount of miconazole required to kill various Candida albicans. Therefore, magnoflorine is a good candidate lead compound for novel antifungal agents.

Immunomodulatory effects of Tinospora crispa extract and its major compounds on the immune functions of RAW 264.7 macrophages.

The in vivo immunomodulatory activities of Tinospora crispa have been reported but its molecular mechanisms underlying its immunomodulatory properties remains obscure and the active constituents contributing to the activities have not been identified. The present study was aimed to investigate the immunomodulatory effects of T. crispa extract (TCE) and its chemical constituents on RAW 264.7 macrophages. Six known compounds including magnoflorine and syringin were isolated by various chromatographic techniques from TCE and their structures were determined spectroscopically. A validated HPLC method was used to quantify magnoflorine and syringin in the extract. The immunomodulatory effects of TCE and its isolated compounds on chemotaxis, phagocytosis, production of inflammatory mediators including reactive oxygen species (ROS), nitric oxide (NO), prostaglandin E2 (PGE2) and pro-inflammatory cytokines which include tumor necrosis factor-α (TNF-α), interleukin (IL)-1ß, IL-6 and monocyte chemoattractant protein-1 (MCP-1) on macrophages were assessed. TCE increased the chemotaxis and phagocytic activity of macrophages and significantly enhanced the production of ROS, NO and pro-inflammatory cytokines. All alkaloids isolated, specifically magnoflorine showed remarkable inducing effects on the chemotaxis, phagocytic activity, ROS and NO productions and the secretions of IL-1ß, TNF-α, IL6, PGE2 and MCP-1. In contrast, syringin potently reduced the chemotaxis, phagocytic activity, ROS and NO productions and secretions of IL-1ß, TNF-α, IL6, PGE2 and MCP-1. TCE showed strong immunostimulant effects on various components of the immune system and these activities were possibly contributed mainly by the alkaloids specifically magnoflorine. TCE has potential to be developed as an effective natural immunostimulant for improvement of immune-related disorders.

Qualitative and quantitative analysis of chemical constituents of Ptychopetalum olacoides Benth.

Ptychopetalum olacoides is a folk medicinal plant for health care in market, especially in Brazil. Fourteen known compounds were isolated from P. olacoides and their chemical structures were elucidated by extensive spectroscopic data, including 1D NMR, 2D NMR, UV, IR and HR-ESI-MS. The 14 known compounds were identified as N-trans-feruloyl-3,5-dihydroxyindolin-2-one (1), magnoflorine (2), menisperine (3), 4-coumaroylserotonin (4), moschamine (5), luteolin (6), 4'-methoxyluteolin (7), 3-methoxyluteolin (8), 3, 7-dimethoxyluteolin (9), caffeic acid (10), ferulic acid (11), vanillic acid (12), syringic acid (13) and ginsenoside Re (14). To our knowledge, compounds (1-6, 13-14) were isolated from the plant for the first time. Additionally, quantitative analysis results indicated that calibration equations of compounds (1-3, 6, 9, 11-13) exhibited good linear regressions within the test ranges (R2 ≥ 0.9990) and magnoflorine and menisperine were the major constituents in the barks of P. olacoides. The contents of magnoflorine and menisperine accounted for 75.96% of all analytes. However, the content of phenolic components was smaller and the highest content was no more than 1.04 mg/g. Collectively, these results suggested that alkaloids are the dominant substances in P. olacoides, which can make a difference for the quality control and further use of P. olacoides.

Simple and Rapid Differentiation of Mutong of Aristolochiaceae by Direct Spray Mass Spectrometry / 分析化学

Direct spray mass spectrometry was used to simply and rapidly differentiate Mutong of Aristolochiaceae from other two kinds of Mutong medicinal materials (Lardizabalaceae and Ranunculacea) by analyzing the chemical profile of Mutong of Aristolochiaceae.A novel method for determination of magnoflorine content in Mutong of Aristolochiaceae was established.The results showed that Mutong of Aristolochiaceae could be identified according to the symbolic component, magnoflorine.Under positive ion mode, semi-quantitative result based on the signal strength ratio of magnoflorine and nuciferin was obtained by choosing nuciferin as an internal standard.The method showed good linear coefficient in the concentration range of 0.50-20.00 mg/L of magnoflorine.The limit of detection was 0.1 mg/L.The method was simple and fast, and could be used for direct and rapid in-situ analysis and identification of Mutong of Aristolochiaceae from other closely related Mutong herbal species without sample pre-treatment.The results were important for the quality control of Mutong herbal medicine.

A novel systems pharmacology platform to dissect action mechanisms of traditional Chinese medicines for bovine viral diarrhea disease.

Due to the large direct and indirect productivity losses in the livestock industry caused by bovine viral diarrhea (BVD) and the lack of effective pharmacological therapies, developing an efficient treatment is extremely urgent. Traditional Chinese medicines (TCMs) that simultaneously address multiple targets have been proven to be effective therapies for BVD. However, the potential molecular action mechanisms of TCMs have not yet been systematically explored. In this work, take the example of a herbal remedy Huangqin Zhizi (HQZZ) for BVD treatment in China, a systems pharmacology approach combining with the pharmacokinetics and pharmacodynamics evaluation was developed to screen out the active ingredients, predict the targets and analyze the networks and pathways. Results show that 212 active compounds were identified. Utilizing these lead compounds as probes, we predicted 122 BVD related-targets. And in vitro experiments were conducted to evaluate the reliability of some vital active compounds and targets. Network and pathway analysis displayed that HQZZ was effective in the treatment of BVD by inhibiting inflammation, enhancing immune responses in hosts toward virus infection. In summary, the analysis of the complete profile of the pharmacological activities, as well as the elucidation of targets, networks and pathways can further elucidate the underlying anti-inflammatory, antiviral and immune regulation mechanisms of HQZZ against BVD.

Anti-inflammatory and antinociceptive activities of Croton urucurana Baillon bark.

ETHNOPHARMACOLOGICAL RELEVANCE: Croton urucurana (Euphorbiaceae) is popularly used in Brazil to treat inflammatory processes, pain, and gastric ulcers. AIM OF STUDY: To evaluate the anti-inflammatory and antinociceptive properties of the methanol extract from the bark of C. urucurana (MECu) in mice and identify its chemical constituents. MATERIALS AND METHODS: The extract was characterized by UHPLC-DAD-ESI-Q-TOF-MS/MS. Extract doses of 25, 100, and 400mg/kg were employed in the biological assays. Evaluation of anti-inflammatory activity was based on paw edema and leukocyte recruitment into the peritoneal cavity of mice, both induced by carrageenan. Abdominal writhing caused by acetic acid and duration of formalin-induced paw-licking were the models employed to evaluate antinociceptive activity. RESULTS: Ten compounds were identified in the extract: (+)-gallocatechin (1), procyanidin B3 (2), (+)-catechin (3), (-)-epicatechin (4), tembetarine (5), magnoflorine (6), taspine (7), methyl-3-oxo-12-epi-barbascoate (8), methyl-12-epi-barbascoate (9), and hardwickiic acid (10). This is the first report of compounds 2, 4, 6, 7, and 10 in C. urucurana and compound 5 in the genus Croton. In addition to inhibiting paw edema and leukocyte recruitment (particularly of polymorphonuclear cells) into the peritoneal cavity of mice, MECu reduced the number of abdominal writhings induced by acetic acid and the duration of formalin-induced paw licking. CONCLUSIONS: The methanol extract of C. urucurana bark exhibited anti-inflammatory and antinociceptive properties, corroborating its use in folk medicine. These effects may be related to the presence of diterpenes, alkaloids, and flavonoids.