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In this study, a precise, rapid, and accurate ultra-performance liquid chromatography–tandem mass spectrometer (UPLC–MS/MS) method for the quantitation of O-demethyl nuciferine in mouse blood was d...

Nuciferine (NF), one of the main and effective components in Nelumbo nucifera Gaertn. leaf extracts, is a promising drug candidate for the treatment of obesity-related diseases, while metformin is a first line therapeutic drug for type 2 diabetes mellitus. Since nuciferine and metformin are likely to be co-administered, the aim of the present study was to evaluate whether co-administration of nuciferine would influence the liver (target tissue) distribution and the anti-diabetic effect of metformin by inhibiting hepatic organic cation transporter 1 (OCT1) and multidrug and toxin extrusion 1 (MATE1). The data demonstrated that nuciferine significantly reduced metformin accumulation in MDCK cells stably expressing human OCT1 (MDCK-hOCT1) or hMATE1 (MDCK-hMATE1), and primary cultured mouse hepatocytes. Furthermore, the presence of nuciferine in the basal compartment caused a concentration-dependent reduction of intracellular metformin accumulation in MDCK-hOCT1/hMATE1 cell monolayers. Compared with the metformin treatment-alone group, co-administration of nuciferine (40 mg/kg) markedly reduced the metformin concentration in mouse livers at 30 and 60 min after a single oral dose of metformin (200 mg/kg), and subsequently impaired the glucose-lowering effect of metformin (200 mg/kg), but the glucose-lowering effect became no different at 90 and 120 min. Therefore, nuciferine influenced the liver concentration and glucose-lowering effect of metformin only for a period of time after dose, administration of nuciferine and metformin with an interval might prevent the drug-drug interaction mediated by OCT1 and MATE1.

Background and purpose Nuciferine, an alkaloid found in Nelumbo nucifera leaves, alleviates dyslipidemia in vivo. However, whether it improves liver injury in diabetic conditions and the underlying mechanism is unclear. The present study aimed to investigate the effects of nuciferine on lipid and glucose metabolism in a murine model of Type 2 diabetes mellitus (T2DM) and to determine the underlying mechanisms of these effects. Experimental approach A murine model of T2DM was induced by high-fat diet (HFD) feeding combined with streptozocin (STZ) injections, and the diabetic mice were treated with nuciferine in their food. The underlying mechanism of the anti-steatotic effect of nuciferine was further explored in HepG2 hepatocytes cultured with palmitic acid. Major signalling profiles involved in fatty acid oxidation were then evaluated, using Western blot, RT-qPCR and si-RNA techniques, along with immunohistochemistry. Key results Nuciferine restored impaired glucose tolerance and insulin resistance in diabetic mice. Hepatic levels of total cholesterol, triglycerides and LDL were decreased, as were the number of lipid droplets, by nuciferine treatment. Furthermore, nuciferine up-regulated β-oxidation related genes in livers of diabetic mice. Luciferase reporter cell assay showed that nuciferine directly reversed palmitic acid-induced inhibition of PPARα transcriptional activity. Silencing PPARγ coactivator-1α (PGC1α) expression in HepG2 cells abolished the effects of nuciferine in accelerating β-oxidation. Conclusions and implications Nuciferine improved lipid profile and attenuated hepatic steatosis in HFD/STZ-induced diabetic mice by activating the PPARα/PGC1α pathway. Nuciferine may be a potentially important candidate in improving hepatic steatosis and the management of T2DM.

Purposes: Nuciferine, a main aporphine alkaloid component found in lotus leaf (Nelumbo nucifera), has been demonstrated to possess the property of reducing fat mass and alleviating dyslipidemia in vivo. The purpose of this study is to explore the effects of nuciferine on the proliferation and differentiation of 3T3-L1 cells and further investigate the possible underlying molecular mechanisms.Methods: 3T3-L1 preadipocytes were treated with 0∼20 μM nuciferine for 24∼120 h, the cell viability was assessed using CCK8. 3T3-L1 preadipocytes and human primary preadipocytes were then induced differentiation and the effects of nuciferine on the lipid metabolism in differentiating and fully differentiated adipocytes were observed by the methods of intracellular triglyceride (TG) assay, Oil Red O staining, RT-qPCR and western blot. Transient transfection and dual luciferase reporter gene methods were used to assess the effects of nuciferine on FAS promoter activities.Results: Nuciferine inhibited the proliferation of 3T3-L1 preadipocytes in a dose- and time-dependent manner. 20 μM nuciferine significantly attenuated lipid accumulation and reduced intracellular TG contents by 47.2, 59.9 and 55.4% on the third, sixth and ninth day of preadipocytes differentiation, respectively (all p < 0.05). Moreover, the mRNA levels of PPARγ, C/EBPα, C/EBPβ, FAS, ACC, HSL and ATGL were notably decreased by 39.2∼92.5% in differentiating preadipocytes when treated with 5∼20 μM nuciferine (all p < 0.05). In fully differentiated adipocytes treated with 20 μM nuciferine for 48 h, the mRNA levels of FAS, ACC and SREBP1 were remarkably downregulated by 22.6∼45.2% compared with the controls (0 μM) (all p < 0.05), whereas the expression of adipokines FGF21 and ZAG were notably promoted by nuciferine. Similarly, in fully differentiated human primary adipocytes, the mRNA levels of FAS, ACC, SREBP1 were decreased and the expression of FGF21 and ZAG were elevated after treated with nuciferine (all p < 0.05). Further mechanism studies showed that 2.5∼20 μM nuciferine significantly decreased FAS promoter activities in 3T3-L1 preadipocytes.Conclusion: Nuciferine inhibited the proliferation and differentiation of 3T3-L1 preadipocytes. The inhibitory effects of nuciferine on adipogenesis might be due to the downregulation of PPARγ, C/EBPα and C/EBPβ, which led to the reduction of intracellular lipid accumulation in 3T3-L1 cells and by downregulating the expression of critical lipogenic enzymes, especially of FAS, which was achieved by inhibiting the FAS promoter activities. Besides, nuciferine promoted the expression of adipokines in fully differentiated adipocytes.

Folium nelumbinis was used as vegetable, functional food and herbal medicine in Asia. In this study, p-sulfonatocalix[6]arenes (SC6A) assisted extraction of nuciferine from the Folium nelumbinis has been carried out to develop an eco-friendly extraction process with high extraction efficacy and facilitated operation. A single factor test was adopted to obtain the optimal extraction conditions, involved the use of 0.4:1:20 (g/g/mL) SC6A / Folium nelumbinis / solution ratio, below 180 μm granulometric fraction and 1 h extraction time with soaking method. Nuciferine and SC6A were separated easily by alkalization, the purity and recovery of nuciferine extracted with SC6A were increased 29.24 and 35.73 times compared with aqueous solution. Moreover, a good reusability of SC6A in the extraction of nuciferine was demonstrated. Competitive fluorescence titration, DSC and FT-IR characterization indicated that SC6A could form host-guest complexes with nuciferine at 1:1 ratio with a binding constant value of 5.55 × 105 M-1. The study provided an eco-friendly, safe and effective method for further development of nutrition supplements contains nuciferine

Objectives: The aim of this study was to analyse the role of two alkaloid, Protopine and Nuciferine, in the prevention and the treatment of the low and mild grade adverse events related to the use of HIVEC® (Hyperthermic IntraVEsical Chemotherapy) instillations. Materials and methods: From September 2017 to September 2019, 100 patients were prospectively randomized into two groups: Group A = Protopine and Nuciferine syrup, 10 ml, once a day, for 8 weeks; Group B = placebo (flavoured coloured water), 10 ml, once a day, for 8 weeks. The primary endpoint was the evaluation of the efficacy of the therapy with Protopine and Nuciferine in controlling of the irritative symptoms. The secondary endpoint was the evaluation of the influences of the treatment on the uroflowmetric parameters. Results: The patients of Group A showed a better International Prostatic Symptoms Score (IPSS) score, a better control of urgency symptoms (PPIUS) and tolerate well the pain (VAS score). The treatment doesn’t modify Uroflow-Qmax and seems to improve the Uroflow-Voided Volume (ml) without influencing the Uroflow-Post Void Residual volume (PVR). Moreover, the treatment with Protopine and Nuciferine has been proven to be effective in the treatment of overactive bladder (OAB) symptoms. Patients’ evaluation of the two different treatments assessed with Patient Global Impression of Improvement questionnaire (PGI-I), demonstrated improvements in the Group A, while the Group B showed a lower satisfaction.Conclusions: Protopine and Nuciferine can be interesting nutraceutical compounds useful to control irritative and pain related symptoms of intravesical chemo/immunotherapy.

Gut microbiota dysbiosis has a significant role in the pathogenesis of metabolic diseases, including obesity. Nuciferine (NUC) is a main bioactive component in the lotus leaf that has been used as food in China since ancient times. Here, we examined whether the anti-obesity effects of NUC are related to modulations in the gut microbiota. Using an obese rat model fed a HFD for 8 weeks, we show that NUC supplementation of HFD rats prevents weight gain, reduces fat accumulation, and ameliorates lipid metabolic disorders. Furthermore, 16S rRNA gene sequencing of the fecal microbiota suggested that NUC changed the diversity and composition of the gut microbiota in HFD-fed rats. In particular, NUC decreased the ratio of the phyla Firmicutes/Bacteroidetes, the relative abundance of the LPS-producing genus Desulfovibrio and bacteria involved in lipid metabolism, whereas it increased the relative abundance of SCFA-producing bacteria in HFD-fed rats. Predicted functional analysis of microbial communities showed that NUC modified genes involved in LPS biosynthesis and lipid metabolism. In addition, serum metabolomics analysis revealed that NUC effectively improved HFD-induced disorders of endogenous metabolism, especially lipid metabolism. Notably, NUC promoted SCFA production and enhanced intestinal integrity, leading to lower blood endotoxemia to reduce inflammation in HFD-fed rats. Together, the anti-obesity effects of NUC may be related to modulations in the composition and potential function of gut microbiota, improvement in intestinal barrier integrity and prevention of chronic low-grade inflammation. This research may provide support for the application of NUC in the prevention and treatment of obesity., Obesity: Getting leaner on lotus leaves A natural compound found in lotus leaf could treat and prevent obesity by stabilizing disrupted gut microbiota and reducing the associated chronic. Obesity is a considerable health burden worldwide, yet treatment options are limited. The composition of an individual’s gut microbiota influences the development of obesity; an imbalance in the ratio of two bacterial species in particular can accelerate the disease. Mingyan Liu and Minjie Wei at the China Medical University, Shenyang, and co-workers demonstrated that nuciferine, a bioactive component of lotus leaf, reduced weight gain and fat accumulation in rats fed a high-fat diet. Nuciferine changed the diversity and composition of the rats’ gut microbiota, and modified the expression of genes involved in processes such as lipid metabolism. Nuciferine also enhanced intestinal integrity, and reduced chronic low-level inflammation.

Multidrug resistance (MDR) is the major barrier to the successful treatment of chemotherapy. Compounds from nature products working as MDR sensitizers provided new treatment strategies for chemo-resistant cancers patients.We investigated the reversal effects of nuciferine (NF), an alkaloid from Nelumbo nucifera and Nymphaea caerulea, on the paclitaxel (PTX) resistance ABCB1-overexpressing cancer in vitro and in vivo, and explored the underlying mechanism by evaluating drug sensitivity, cell cycle perturbations, intracellular accumulation, function and protein expression of efflux transporters as well as molecular signaling involved in governing transporters expression and development of MDR in cancer.NF overcomes the resistance of chemotherapeutic agents included PTX, doxorubicin (DOX), docetaxel, and daunorubicin to HCT-8/T and A549/T cancer cells. Notably, NF suppressed the colony formation of MDR cells in vitro and the tumor growth in A549/T xenograft mice in vivo, which demonstrated a very strong synergetic cytotoxic effect between NF and PTX as combination index (CI) (CI0.1) indicated. Furthermore, NF increased the intracellular accumulation of P-gp substrates included DOX and Rho123 in the MDR cells and inhibited verapamil-stimulated ATPase activity. Mechanistically, inhibition of PI3K/AKT/ERK pathways by NF suppressed the activation of Nrf2 and HIF-1α, and further reduced the expression of P-gp and BCRP, contributing to the sensitizing effects of NF against MDR in cancer.This novel finding provides a promising treatment strategy for overcoming MDR and improving the efficiency of chemotherapy by using a multiple-targets MDR sensitizer NF.

Ethnopharmacological relevance San-Ye-Tang-Zhi-Qing formula (SYTZQ) is an effective prescription for the treatment of pre-diabetes disorders of glycolipid metabolism in type 2 diabetes mellitus (T2DM). It consists of five Chinese herbs including Mori Folium, Nelumbinis Folium, Crataegi Folium, Salviae Miltiorrhizae Radix et Rhizoma and Paeoniae Radix Rubra. Aim of the study This study was aimed to reveal the pharmacological mechanism of pharmacokinetic target components of SYTZQ for the treatment of T2DM. Materials and methods A rapid, precise and sensitive ultra-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method was developed to quantify simultaneously nuciferin, vitexin-4″-O-glucoside, vitexin-2″-O-rhamnoside, paeoniflorin and rosmarinic acid in rat plasma after oral administration of SYTZQ. The network pharmacology was used to analyze the effect of the compounds absorbed into the blood of SYTZQ on T2DM. The effects of paeoniflorin, nuciferine and rosmarinic acid on adipogenic differentiation were validated in vitro experiments. Results The separation was performed on an ACQUITY UHPLC HSS T3 column (2.1 mm × 100 mm, 1.7 μm) using acetonitrile and 0.1% (v/v) formic acid in water as the mobile phase in gradient elution. The calibration curves of five analytes showed good linearity (r ≥ 0.9991) with the lower limits of quantification (LLOQ) between 0.3 and 5.0 ng/mL. The recoveries and matrix effects of five analytes ranged from 81.1% to 113%. The RSDs of inter-day and intra-day precision were all within 13.7%. The validated method was successfully applied to the pharmacokinetic study of five ingredients after oral administration of SYTZQ to rat. 39 major targets and 22 candidate pathways of five compounds absorbed into the blood of rats after administration of SYTZQ were identified and successfully constructed a compound-target-disease-pathway network. It was confirmed that paeniforin, nuciferine and rosmarinic acid could decrease the adipogenicity differentiation in vitro experiments. Conclusions The pharmacokinetic parameters indicated that the five components (nuciferin, vitexin-4″-O-glucoside, vitexin-2″-O-rhamnoside, paeoniflorin and rosmarinic acid) were absorbed and eliminated quickly in vivo. These five absorbed components were associated with 22 pathways, including insulin resistance, regulation of lipolysis in adipocytes, PI3k/AKT-, TNF-, cAMP- and cGMP-PKG-signaling pathway. Paeoniflorin, nuciferine and rosmarinic acid have the effect of inhibiting adipocyte differentiation. This study could provide more reference for quality control, and provide a firm basis for evaluating the clinical efficiency of SYTZQ.

Onychopetalum is an Amazonian botanical genus close-related with Bocageopsis and Unonopsis, whose species are used in traditional medicine. Although these genera are widely investigated from the phytochemical viewpoint, the knowledge about Onychopetalum remains almost unexplored. In order to perform a comprehensive characterization of isoquinoline-derived alkaloids from Onychopetalum amazonicum, an integrative approach based on the alkaloid profile analysis by leaf spray mass spectrometry (LS-MS), followed by a robust dereplication through high-performance liquid chromatography coupled with diode array detector and atmospheric pressure chemical ionization tandem mass spectrometry, (HPLC-DAD-APCI-MS/MS) and nuclear magnetic resonance (NMR) analysis was adopted. LS-MS allowed the tentative identification of reticuline, norjuziphine, nornuciferine, O-methylisopiline, nuciferine, N-methyl-O-methylisopiline, anonaine, and N-methylanonaine. HPLC-DAD-MS/MS analysis supported the tentative identification of N-methylasimilobine, asimilobine, norushinsunine, N-methylisopiline, isopiline, liriodenine, and lysicamine. Moreover, the integration of MS and NMR data allowed the additional identification of stepholidine and isocorypalmine. The applied integrative approach proposed herein enabled a comprehensive analysis of the alkaloid content from the O. amazonicum, showing to be a useful strategy for the dereplication of isoquinoline-derived alkaloids.

Label-free cell phenotypic assays were performed to establish a β2-adrenoceptor (β2-AR) target model in A431 cells and a β1-AR target model in transfected HEK293-β1 cells, using known β2-AR and β1-AR agonists and antagonists. A list of natural compounds was screened on the target models, among which seven new compounds were found to be antagonistically active against β2-AR. After receptor specificity evaluations on hydroxyl carboxylic acid receptor-2 (ΗΧΑ-2), histamine receptor (H1R), and β1-adrenoceptor (β1-AR), six out of the seven compounds, including nuciferine, epiberberine, harmaline, harmine, palmatine, and columbamine, exhibited specific antagonistic activity against β2-AR. Epiberberine and palmatine showed the strongest antagonistic activities against β2-AR with IC50 values of 2.3 ± 0.2 μM and 2.6 ± 0.3 μM, respectively. Docking palmatine to the crystal structure of human β2-AR (PDB 5X7D) suggested that the ligand forms a hydrogen bond with N312 and hydrophobic interaction with several amino acid residues in the binding pocket, such as D113 and V114. The kinetic binding profile of palmatine was further investigated using co-stimulation assays. Results suggested that palmatine was a competitive antagonist for β2-AR. The six novel β2-AR antagonists provide a promising chemical starting point for identification and optimization of drugs used for treating hypertension, glaucoma, and infantile hemangiomas. This study also lays the foundation for the in-depth investigation of biochemical mechanisms and pharmacological properties of natural compounds.

Lotus (Nelumbo nucifera Gaertn.) contains various bioactive compounds, with benzylisoquinoline alkaloids (BIAs) as one of the major groups. The biosynthetic pathways of two major bioactive BIAs in this plant, nuciferine and N-nornuciferine, are still not clear. Therefore, several genes related to BIA biosynthesis were searched from the lotus database to study the role of key genes in regulating these pathways. In this study, the expression profiles of NCS, CNMT, 6OMT, CYP80G2, and WRKY TFs were investigated in mechanically wounded lotus leaves. It was found that the accumulation of nuciferine and N-nornuciferine significantly increased in the mechanically wounded lotus leaves in accordance with the relative expression of putative CYP80G2 and one WRKY transcription factor (NNU_24385), with the coregulation of CNMT. Furthermore, the role of methyltransferase-related genes in this study suggested that methylation of the isoquinoline nucleus to yield a methylated-BIA structure may occur at the N position before the O position. Altogether, this study provides improved understanding of the genes regulating BIA biosynthesis under stressed conditions, which could lead to improvements in BIA production from the commercial lotus.

In this study, inhibitory kinetics of Nuciferine and Methyl Ganoderate extrated from Lotus Leaves and Ganoderma lucidum on Mucor miehei Lipase were studied first. The molecular structure of Nuciferine and Methyl Ganoderate were determined. The inhibitory effects of two extracts on lipase were reversible, with the IC50 values of 0.194 and 0.332 mg/mL, respectively. The inhibition kinetic analysis by Lineweaver-Burk plots showed that they were a mixed-type inhibitor of lipase, with inhibition constants KI of 0.16 and 0.29 mg/mL, and KIS of 0.36 and 0.49 mg/mL, respectively. Results of spectral analysis showed that the UV absorption and the molecule fluorescence spectrum of the lipase hydrolyzate were significantly decreased after the inhibitor was added. The molecular docking further suggested that the interaction site between the active substance and inhibitor was located in an α-helix and a β-sheet of the lipase, and the lipase active site was interfered by the inhibitor near the cap structure. In addition, the proliferation and differentiation of 3 T3-L1 preadipocytes were inhibited by two extracts. Total triglycerides and cholesterol were significantly reduced in the cells. The results confirmed that Nuciferine and Methyl Ganoderate can be used as potential obesity treatment drugs.

Recently, type H vessels were reported to couple angiogenesis and osteogenesis during osteoclastogenesis, and tartrate-resistant acid phosphatase (Trap)+ preosteoclasts were found to secrete increased PDGF-BB to promote type H vessel formation. Therefore, utilization of type H vessels may be a strategy to treat diseases involving bone loss. In the present study, we found that nuciferine, a natural bioactive compound, has various effects, including inhibiting osteoclastogenesis and promoting type H vessel formation. Nuciferine inhibited osteoclastogenesis and bone resorption but increased the relative number of Trap+ preosteoclasts. Nuciferine restrained the expression of osteoclast-specific genes and proteins, promoted PDGF-BB production and potentiated related angiogenic activities by inhibiting the MAPK and NF-κB signaling pathways in vitro. We confirmed the bone-protective effects of nuciferine in ovariectomized mice and found that nuciferine treatment increased the PDGF-BB concentration and the number of type H vessels in the femur. In conclusion, our results demonstrated that nuciferine can decrease multinucleated osteoclast formation and promote type H vessel formation through preservation of Trap+ preosteoclasts via inhibition of the MAPK and NF-κB signaling pathways and may be an excellent agent for the treatment of diseases involving bone loss.

Background A quality marker (Q-marker) is defined as an inherent chemical compound that is used for the quality control of a drug. Its biological activities are closely related to safety and therapeutic effects. Generally, a multiple-component herbal medicine may have many Q-markers. We therefore proposed a concept of “super Q-marker” satisfying both the criterion of Q-markers and PK-markers to be used in more effective quality control of herbal medicine. Purpose The first aim was to find suitable prototype-based PK-markers from Tangzhiqing tablets (TZQ), a Chinese patent medicine. Then super Q-markers were expected to be identified from the prototype-based PK-markers based on an in vitro-in vivo correlation study. Methods Potentially eligible prototype-based PK-markers were identified in a single- and multiple-dose pharmacokinetic study on TZQ in 30 healthy volunteers. The in vitro dissolution and permeation profiles of the prototype-based PK-markers of TZQ were evaluated by the physiologically-based drug dissolution/absorption simulating system (DDASS). An in vitro-in vivo correlation analysis was conducted between the dissolution/permeation behaviors in DDASS and the actual absorption profiles in human to test the transferability and traceability of the promising super Q-markers for TZQ. Results In human, plasma paeoniflorin and nuciferine as prototype-based PK-markers exhibited the appropriate pharmacokinetic properties, including dose-dependent systemic exposure (AUC, Cmax) and a proper elimination half-life (1∼3h). In DDASS, it was predicted that paeoniflorin and nuciferine are highly permeable but the absorption rates are primarily limited by the dissolution rates. Moreover, the established in vitro-in vivo correlations of paeoniflorin and nuciferine were in support of the super Q-markers features. Conclusion Paeoniflorin and nuciferine are identified as the super Q-markers from the prototype-based PK-markers of TZQ based on findings from a combination of in vitro, in vivo, and in vitro-in vivo correlation studies. This method is practical for optimal identification of qualified Q-markers, thus helping improve the quality control of herbal medicines.

Correction for ‘Synthesis and evaluation of nuciferine and roemerine enantiomers as 5-HT2 and α1 receptor antagonists’ by Hui Li Heng et al., Med. Chem. Commun., 2018, DOI: 10.1039/c7md00629b.

In the present study, various lotus leaf extracts were prepared by multiple methods, characterized by 12 alkaloids, 14 flavonoids, and superoxide anion radical scavenging (OS) and xanthine oxidase inhibitory (XOI) activities, and divided into 3/4 groups with different characteristics. Both alkaloids and flavonoids contributed to XOI activity of representative lotus leaf extract. Norcoclaurine, O-nornuciferine, N-nornuciferine, nuciferine, luteolin, diosmetin, quercetin, and quercetin-3-O-glucuronide were defined as quality control standards for lotus leaf extracts with dual OS and XOI activities. Enzymatic hydrolysis coupled with ultrasound-assisted extraction method using 50% (v/v) ethanol was the most favorable approach for preparing food-grade lotus leaf extract with dual functions. Network pharmacology revealed that this extract having abundant alkaloids and flavonoids (especially flavonoid aglycones) could improve hyperuricemia by multiple actions on uric acid synthesis and transport, inflammation response, oxidative stress and lipid metabolism. Our study provided crucial data about clear quality control standards for plant-based functional food with potential hypouricemic effect.

Osteoarthritis (OA), a chronic disease featured by severe pain and persistent inflammation, significantly affects human health and lowers the quality of life. Nuciferine (Nuf) is major alkaloids abundant in lotus leaf and has various biological activities. In current study, we aimed to explored the protective effect of Nuf against OA and its underlying mechanism in vitro and in vivo. It was found that Nuf alleviated the degradation of extracellular matrix (ECM) under the stimulation of IL-1β and inhibited the production of inflammation cytokines such as tumor necrosis factor (TNF)-α, interleukin (IL)-6, inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in vitro. Meanwhile, it was demonstrated that Nuf suppressed the activation of PI3K/Akt/NF-κB signaling pathway induced by IL-1β. Besides, there was favorable binding ability between Nuf and PI3K. Nuf significantly alleviated the development of OA in vivo in the rat OA model established by anterior cruciate ligament transaction (ACLT). Together, our results indicated that Nuf attenuates the progression of OA by targeting PI3K/Akt/NF-κB signaling pathway, which makes Nuf as a promising candidate for the treatment of OA.

The incidence and prevalence of metabolic syndrome has steadily increased worldwide. As a major risk factor for various diseases, metabolic syndrome has come into focus in recent years. Some natural aporphine alkaloids are very promising agents in the prevention and treatment of metabolic syndrome and its components because of their wide variety of biological activities. These natural aporphine alkaloids have protective effects on the different risk factors characterizing metabolic syndrome. In this review, we highlight the activities of bioactive aporphine alkaloids: thaliporphine, boldine, nuciferine, pronuciferine, roemerine, dicentrine, magnoflorine, anonaine, apomorphine, glaucine, predicentrine, isolaureline, xylopine, methylbulbocapnine, and crebanine. We particularly focused on their impact on metabolic syndrome and its components, including insulin resistance and type 2 diabetes mellitus, endothelial dysfunction, hypertension and cardiovascular disease, hyperlipidemia and obesity, non-alcoholic fatty liver disease, hyperuricemia and kidney damage, erectile dysfunction, central nervous system-related disorder, and intestinal microbiota dysbiosis. We also discussed the potential mechanisms of actions by aporphine alkaloids in metabolic syndrome.

Phytochemical investigation of the CHCl3 extract of the seed embryos of Nelumbo nucifera Gaertn resulted in the isolation of a new naturally occurring bisbenzylisoquinoline alkaloid, O-methylneferine (1), together with five known alkaloids, neferine (2), armepavine (3), (–)-(1R)-N-methylcoclaurine (4), nuciferine (5), and pronuciferine (6). The structures of these compounds were characterized by spectroscopic methods and comparison of physical properties with those reported in the literature. Among them, compounds 1 and 2 exhibited significant activity against human colon adenocarcinoma cell line (HT-29), with IC50 values of 0.70 and 1.61 µM, respectively, which were 8- and 3.5-fold higher than that of the reference anticancer drug, doxorubicin (IC50 5.63 µM). Moreover, compounds 1 and 2 displayed less cytotoxic activity against the non-cancerous HEK 239 cells with the IC50 values of 42.48 and 12.19 μM, respectively, whereas the cytotoxicity of doxorubicin against this cell line was 0.22 μM. The very potent cytotoxicity against HT-29 cell line and very high selectivity index (60.6-fold) of the alkaloid 1 is of particular significant; it could be considered as a promising structure lead for anti-colon cancer drug development.

Pharmaco- or psycho-active compounds in traditional medicines or in plants used during neoshamanic rituals can sometimes lead to the (re-) discovery of new drugs for chronic pain, anxiety, depression or schizophrenia. Neoshamanic rituals fulfill the needs for healing and transformation of a number of people, who are partly dissatisfied with the absence of certain healing aspects in western medicine. A recent neoshamanic ritual is based on the administration of a resin from the Blue Nile Flower (Nymphaea caerulea), also referred to as Blue Lotus or Sacred Blue Lily of the Nile). This extract contains a number of alkaloids such as aporphine and nuciferine, although the quality of commercial available extracts is highly variable. We will discuss the biological effects of nuciferine, a compound which may be of value for the treatment of psychiatric disorders, and discuss the use of Blue Nile Flower in rituals from a transpersonal psychological perspective. Nuciferine has an enriched pharmacological profile, with affinities for a number of serotonergic and dopaminergic receptors. Nuciferine and its derivatives might lead to a new family of atypical antipsychotic compounds. Furthermore, a recent identified mechanism of action related to its anti-inflammatory activity, suggest this molecule might also play a role in the treatment of depression and posttraumatic stress disorder.

Ethnopharmacological relevance Jiang-Zhi-Ning (JZN) is a traditional Chinese medicine formula, which has the effect of lowering blood lipid level and softening blood vessels. It is clinically used in the treatment of hyperlipidemia with significant curative effect. Aim of the study This study aims to screen the active components of JZN that are responsible for its blood lipids lowering effect and lay the foundation for elucidating pharmacodynamic material basis of the hypolipidemic effect of the formula. Materials and methods The hyperlipidemia model was used to evaluate the efficacy of the JZN effective extraction with the TC and TG of rat plasma as evaluation index. Then the established ultra-high performance liquid chromatography coupled with electrospray ionization-quadrupole-time of flight-mass spectrometry (UPLC-ESI-Q-TOF-MSn) method was utilized to analyze the components of JZN effective extraction and the absorbed components in rat plasma, the potential active components were screened by using the combined analysis results of in vivo and in vitro component identification. Then an established ultra-high performance liquid chromatography-triple quadrupole mass spectrometry (UPLC-QqQ-MSn) method was used to determine the content of potential active components and its natural ratio in JZN effective extraction, and a potential active components combination (PACC) was formed accordingly. Then a HepG2 cell hyperlipidemia model induced by sodium oleate was used to study the hypolipidemic activity of PACC by detecting the content of TG level in the model. Meanwhile, the real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) was used to conduct preliminary research on its hypolipidemic mechanism. Then combined with the concept of “combination index” in the “median-effect principle”, to calculate the half inhibitory concentration (IC50) values of PACC and each monomer component on inhibiting the TG level in the cell model. Subsequently, the “activity contribution study” was carried out, and the components with the sum of the “activity contribution value” of 85% were finally selected as the hypolipidemic active components of JZN. Results The pharmacodynamics results showed that JZN effective extraction has displayed a good hypolipidemic effect. 45 components were identified in vitro, 108 components were identified from rat plasma, and 17 potential active components were screened out. The content determination result showed that the ratio of each potential active components in PACC as following: cassiaside C: rubrofusarin-6-O-gentiobioside: aurantio-obtusin-6-O-glucoside: hyperoside: isoquercitrin: quercetin-3-O-glucuronide: (E)-2,3,5,4′-tetrahydroxystilbene-2-O-glucoside: rutin: emodin-8-O-glucoside: astragalin: armepavine: N-nornuciferine: coclaurine: O-nornuciferine: nuciferine: N-norarmepavine: higenamine = 3.30: 16.06: 9.15: 23.94: 98.40: 417.45: 189.68: 8.62: 1.28: 5: 3.51: 14.57: 1.06: 1.35: 1: 5.64: 6.06, and the activity study results showed that it has displayed a good hypolipidemic activity. Finally, the hypolipidemic active components screened out by the “activity contribution study” were: quercetin-3-O-glucuronide, (E)-2,3,5,4′-tetrahydroxystilbene-2-O-glucoside, isoquercitrin, O-nornuciferine, hyperoside and rubrofusarin-6-O-gentiobioside. Conclusions A scientific and rational approach of screening the hypolipidemic active ingredients of JZN has been developed in the current study. In addition, the research revealed the blood lipid lowering mechanism of those ingredients, which provide a solid basis for further elucidating the hypolipidemic pharmacodynamic material basis and action mechanism of JZN.

This study aimed to elucidate the mechanisms of nuciferine (a main aporphine alkaloid of lotus leaf extract), which can induce relaxation in contracted tracheal rings. Under Ca2+-free and 2 mM Ca2+ conditions, we found that nuciferine had no effect on the resting muscle tone of tracheal rings. In contrast, nuciferine relaxed high K+-contracted mouse tracheal rings in a dose-dependent manner and inhibited both Ca2+ influx and voltage-dependent L-type Ca2+ channel currents induced by high K+. Similarly, nuciferine also inhibited acetylcholine-induced contractions in mouse tracheal rings in a dose-dependent manner. Meanwhile, both acetylcholine-induced intracellular Ca2+ influx and whole-cell currents of nonselective cation channels were blocked by nuciferine. Together, the results indicate that nuciferine-induced relaxation in tracheal rings mainly occurred due to the inhibition of extracellular Ca2+ influx through the blockade of voltage-dependent L-type Ca2+ channels and/or nonselective cation channels. These results suggest that nuciferine has a therapeutic effect on respiratory diseases associated with the aberrant contraction of airway smooth muscles and/or bronchospasm.

The alkaloids from lotus (Nelumbo nucifera Gaertn) are effective in lowering hyperlipemia and level of cholesterol. However, there is not a general method for their separation. In this work, a general ionic liquid pH-zone-refining countercurrent chromatography method for isolation and purification of six alkaloids from the whole lotus plant was successfully established by using ionic liquids as the modifier of the two-phase solvent system. The conditions of ionic liquid pH-zone-refining countercurrent chromatography, involving solvent systems, concentration of retainer and eluter, types of ionic liquids, the content of ionic liquids as well as ionic liquids posttreatment, were optimized to improve extraction efficiency. Finally, the separation of these six alkaloids was performed with a two-phase solvent system composed of n-hexane-ethyl acetate-methanol-water-[C4mim][PF6] at a volume ratio of 5:2:2:8:0.1, where 10mM TEA was added to the organic stationary phase as a retainer and 3mM HCl was added to the aqueous mobile phase as an eluter. As a result, six alkaloids including N-nornuciferine, liensinine, nuciferine, isoliensinine, roemerine and neferine were successfully separated with the purities of 97.0%, 90.2%, 94.7%, 92.8%, 90.4% and 95.9%, respectively. The established general method has been respectively applied to the crude samples of lotus leaves and lotus plumules. A total of 37.3mg of liensinine, 57.7mg of isoliensinine and 179.9mg of neferine were successfully purified in one run from 1.00g crude extract of lotus plumule with the purities of 93.2%, 96.5% and 98.8%, respectively. Amount of 45.6mg N-nornuciferine, 21.6mg nuciferine and 11.7mg roemerine was obtained in one step separation from 1.05g crude extract of lotus leaves with the purity of 96.9%, 95.6% and 91.33%, respectively.

Nuciferine, a major alkaloid found in Nelumbinis Folium, exhibits a broad spectrum of bioactivities, such as antiobesity, anti-diabetes and anti-inflammatory. However, many research regarding nuciferine focused on the extraction, isolation and biological activity, the metabolism is not comprehensively explained in vivo. Thence, the present of this paper is to establish a simple method for speculating metabolites of nuciferine. A total of 15 metabolites were detected and tentatively identified through ultra high performance liquid chromatography-diode array detection-quadrupole time-of-flight mass spectrometry (UHPLC-DAD-QTOF-MS), including 7 new metabolites. Among them, we also discovered a previously unmentioned metabolically active site at the C1-OCH3 position. These metabolites suggested that demethylation, oxidation, glucuronidation and sulfation were major metabolic pathways. This study provided significant experiment basis for its safety estimate and valuable information about the metabolism of nuciferine, which will be advantageous for new drug development.

Once breast cancer cells grow aggressively and become lodged in the skeleton through migration and invasion, they interact with bone microenvironment and accelerate much more tumor growth and bone destruction. We investigated whether liensinine and nuciferine, major active components in Nelumbo nucifera (lotus), could prevent breast cancer cell-mediated bone destruction. Liensinine and nuciferine inhibited the growth of MDA-MB-231 and MCF-7 human breast cancer cells by inducing apoptosis and inhibiting proliferation via cell cycle arrest. Liensinine treatment led to the increased Bax/Bcl-2 ratio, activation of caspase-3, and subsequent cleavage of PARP. Liensinine also displayed significant inhibition on the migration and invasion of both MDA-MB-231 and MCF-7 human breast cancer cells compared with nuciferine. In addition, liensinine and nuciferine inhibited the receptor activator of nuclear factor kappa-B ligand- (RANKL-) induced osteoclast differentiation in mouse bone marrow macrophage cells and mature osteoclast-mediated bone resorption. Furthermore, oral administration of liensinine reduced the osteolysis in nude mice with intratibial injection of MDA-MB-231 cells. Collectively, liensinine and nuciferine may be promising candidates for preventing and treating breast cancer bone metastasis and the resulting osteolytic bone loss by targeting both cancer cells and osteoclasts. Liensinine has more potent anticancer and antibone resorptive activities than nuciferine.