Poster Abstract 1

Design and development of a reporter plasmid for high-throughput screening of botanical extracts and small molecule libraries to discover prenylated flavonoid ligands of FXR

Isabelle E. Logan, Department of Biochemistry and Biophysics, Linus Pauling Institute, Oregon State University
Mary L. Fantacone, Linus Pauling Institute, Oregon State University
Dylan Nelson, College of Pharmacy, Oregon State University
Claudia S. Maier, Department of Chemistry, Linus Pauling Institute, Oregon State University
Jan F. Stevens, College of Pharmacy, Linus Pauling Institute, Oregon State University
Adrian F. Gombart, Department of Biochemistry and Biophysics, Linus Pauling Institute, Oregon State University

The farnesoid X receptor (FXR) is a bile acid nuclear receptor and biological sensor for the regulation of bile acid biosynthesis. When a bile acid ligand binds FXR, it forms a heterodimer with the retinoid X receptor (RXR) and enters the cell’s nucleus. The FXR/RXR heterodimer then binds to FXR response elements (FXREs) present in the promoters of its target genes. FXR activation leads to transcriptional repression of genes involved in de novo lipogenesis and gluconeogenesis and plays an important role in obesity, and metabolic syndrome. Interestingly, mice fed a high-fat diet supplemented with  xanthohumol (XN), a prenylated flavonoid found in hops (Humulus lupulus), showed a dose-dependent decrease in body weight gain and glucose, leptin, and cholesterol levels. Using hydrogen deuterium exchange mass spectrometry (HDX-MS) combined with computational and fluorescence titration studies we indicated that XN binds within the canonical FXR ligand-binding pocket to modulate FXR target gene expression. We hypothesize that other prenylated flavonoids mediate their beneficial health effects by acting as ligands for FXR. To identify additional FXR ligands, we inserted three FXREs and a minimal thymidine kinase promoter into a two-step transcriptional activator (TSTA) reporter plasmid to drive expression of a Gal4 DNA binding domain-VP16 fusion protein. The fusion protein binds, in turn, to a GAL4 promoter in the TSTA reporter plasmid driving expression of β-Lactamase. This enables monitoring of promoter activity using a Förster resonance energy transfer (FRET)-based reporter system. We have shown that a natural ligand, chenodeoxycholic acid, and a synthetic ligand, GW4064, of FXR increase reporter gene expression. This plasmid will enable a robust, sensitive, and inexpensive high-throughput screening assay of small molecule libraries and botanical extracts for potential prenylated flavonoid ligands of FXR.


Poster Abstract 2

Effects of nitrate and nitrite exposure on zebrafish behavior and brain metabolic phenotyping

Manuel, Garcia-Jaramillo, Linus Pauling Institute, Oregon State University
Elizabeth R. Axton, College of Pharmacy, Department of Environmental and Molecular Toxicology, Linus Pauling Institute, Oregon State University
Laura M. Beaver, Linus Pauling Institute, Biological and Population Health Sciences, Oregon State University
Rosa M. Keller, Oregon State University
Norman Hord, Biological and Population Health Sciences, Oregon State University
Jan F. Stevens, College of Pharmacy, Linus Pauling Institute, Oregon State University

The effects of nitrate and nitrite, present in the environment and our diet, on animal metabolism are controversial. We do not currently know the extent to which nitrate and nitrite exposure affects cognitive behavior and changes the abundance of downstream target molecules in the brain. In order to determine the physiological and cognitive effects derived from the exposure to different doses of nitrite and nitrate, we carried out a study with the aquatic model organism Danio Rerio (Zebrafish). Animals were exposed to sodium nitrite (9.75 and 19.5 mg/L), sodium nitrate (60.7, 303.5 and 606.9 mg/L), or control water. For the last three days of the experiment, two sets of fish were treated with 15N-nitrate or 15N-nitrite, in order to study the incorporation of these molecules into the brain metabolism. After four weeks, animals were euthanized and brains extracted. Samples were analyzed for untargeted metabolomics with a 5600 ABSciex TripleTOF in positive and negative mode. Pathway analysis showed significant differences in more than fifty identified compounds when comparing the metabolites of control and treated fish. The strongest variations were observed in the metabolism of aspartate, glutamate, leucine and taurine. Low incorporation of labeled nitrate and nitrite into the brain leads to the conclusion that the behavioral patterns observed in nitrate- and nitrite-treated fish, including anxiety and learning deficits, are due to indirect effects of those molecules on brain metabolism. Overall, we found a very significant depletion in many metabolites involved in the regulation of neuronal activity. For instance, the lower concentration of leucine in the brain of fish exposed to nitrite and nitrate may explain the reduction in glutamate metabolism and the observed behavioral patterns. Our results suggest that nitrate and nitrite treatment may be related to metabolic changes in zebrafish brain that result in behavioral changes associated with anxiety.

Poster Abstract 3
Metabolism of xanthohumol and 8-prenylnaringenin by intestinal microbiota

Ines L. Paraiso, College of Pharmacy, Linus Pauling Institute, Oregon State University
Ryszard Zielke, College of Pharmacy, Oregon State University
Aleksandra E. Sikora, College of Pharmacy, Oregon State University
Jan F. Stevens, College of Pharmacy, Linus Pauling Institute, Oregon State University


Xanthohumol (XN), a flavonoid found in hops (Humulus lupulus), exerts mitigating effects on the metabolic syndrome. XN has several pharmacological targets in vitro but appears to be active at comparatively low concentrations in vivo, suggesting the involvement of bioactive metabolites. To fully understand XN’s mechanism of action in vivo, it is necessary to gain more information on its fate in the body following ingestion, as a certain proportion of ingested secondary plant constituents undergo colonic microbial transformation. To study the metabolism of XN and related prenylated flavonoids, XN and 8-prenylnaringenin (8PN) were incubated with E. ramulus, a strictly anaerobic bacterium detectable in the gastrointestinal tract of most individuals. Evidence from our study shows that both XN and 8PN are extensively transformed by this gut microbe, producing metabolites with similar pharmacological properties. Moreover, co-cultures with E. limosum, another intestinal bacterium previously reported to be involved in XN gut metabolism prove that E. ramulus can utilize metabolites made readily available by another bacterium. Degradation pathways of XN are proposed based on the intermediates detected by high performance liquid chromatography (HPLC) coupled with tandem mass spectrometry (MS/MS).

Poster Abstract 4
Characterization and quantification of phytochemical constituents in the medicinal herb Centella asiatica by high-resolution mass spectrometry

Armando Alcazar Magana, Department of Chemistry, Oregon State University
Maya Caruso, Department of Neurology, Oregon Health & Science University
Kirsten Wright, Department of Neurology, Oregon Health & Science University
Amala Soumyanath, Department of Neurology, Oregon Health & Science University
Joseph Quinn, Department of Neurology, Oregon Health & Science University
Jan F. Stevens, College of Pharmacy, Linus Pauling Institute, Oregon State University
Claudia S. Maier, Department of Chemistry, Linus Pauling Institute, Oregon State University

In Ayurvedic medicine, Centella asiatica (CA) has been used to prevent cognitive decline and for brain function improvement. The results of several studies in humans and rodent models have caused excitement in using CA preparations as a potential complementary medicine to improve memory in aging-related cognitive decline and maybe even Alzheimer’s disease. CA has been reported to have also other biological activities beneficial for human health such as gastric ulcers, anti-inflammatory, wound healing, and immunostimulant properties. Geographical, genetic and post-harvest processing all affect the secondary metabolite composition of CA products, potentially influencing their biological effects and study reproducibility. In our laboratory, we favor the use of ultra-performance liquid chromatography (UPLC) in conjunction with accurate mass high-resolution tandem mass spectrometry for structural analysis and quantification, allowing detailed targeted and untargeted characterization of plant extracts. Aqueous and ethanolic CA extracts were characterized using an ultra-performance liquid chromatograph (UPLC) connected to a Synapt G2 HDMS high-resolution accurate mass spectrometry system. For quantification of phytochemicals known to occur in CA, an AB Sciex Triple TOF 5600 mass spectrometer using a parallel reaction monitoring. The disclosed method allows quantification of a) seven flavonoids, b) three structural isomers of caffeoylquinic acids, c) five di-caffeoylquinic acids, d) six caffeic acids derivatives and e) the major saponins and sapogenins. Recovery experiments were carried out for CA extracts. CA samples were spiked with 24 available standards at two different concentration levels (0.25 ng and 5 ng on column for each compound). Recoveries of individual compounds were in range from 91 to 132 %. Three standard mixtures of known concentrations (low, medium and high) were analyzed. The analytical accuracy was in the range of 87-125 %, confirming the feasibility of the proposed procedure for quantitative analysis of CA samples. Differences were observed in the chemical profiles of the CA extracts, demonstrating that standardization and detailed characterization of CA extracts are pre-requisites to reliably and reproducibly study the biological activity of CA preparations.
Acknowledgement: This project is supported by National Institutes of Health Grant # R01AT008099.

Poster Abstract 5
Profiling of specific metabolic signatures in deuterium-labeled and unlabeled broccoli sprouts with X13CMS

Jaewoo Choi, Linus Pauling Institute, Oregon State University
Robert W. Durst, Linus Pauling Institute, Oregon State University
Laura M. Beaver, Biological and Population Health Sciences, Linus Pauling Institute, Oregon State University
Emily Ho, Biological and Population Health Sciences, Moore Family Center for Whole Grain Foods, Linus Pauling Institute, Oregon State University
Jan F. Stevens, College of Pharmacy, Linus Pauling Institute, Oregon State University


Cruciferous vegetables (crucifers) and their bioactive components in food, including indoles and isothiocyanates (ITCs) such as sulforaphane (SFN), appear to modulate cancer risk but observational data to date in humans are inconsistent. Such discrepancies arise in part owing to methodological limitations of accurately assessing dietary exposures on breast and prostate cancer risk. Despite widespread use, classical dietary-intake instruments including food frequency questionnaires and other dietary recall methods are subject to well-known limitations. To circumvent and address the unmet current need for reproducible measures of dietary intake and metabolic impacts of crucifers, we used deuterium-labeled broccoli sprouts in combination with mass spectrometry based metabolomics approaches to quantify and differentiate between broccoli-specific metabolites and their interactions with their molecular targets of action. In methods, broccoli seeds were germinated for 5 days on H2O or 25% deuterium (D2O), refreshed twice per day, and the harvested sprouts were homogenized in methanol (500 μL) using a bullet blender with zirconium oxides beads. The extracts were centrifuged and the supernatants analyzed by UPLC-QToF mass spectrometry on a Sciex 5600 TripleToF instrument. The raw ToF-MS data were converted to mzXML format using MSConvert software. The converted mzXML data were processed with XCMS software for peak detection and retention-time alignment. The XCMS output was subsequently processed with X13CMS software to identify deuterium labeled compounds. XCMS and X13CMS were operated in R software. We processed the raw LC-QToF MS data by XCMS yielding 1428 spectral features, which were re-analyzed using X13CMS software. This step yielded 152 isotopologue groups representing 152 deuterium-enriched metabolites covering glucosinolates derived from methionine (glucoraphanin) and tryptophan (glucobrassicin, neoglucobrassicin) as well as amino acids. The success of this approach does not depend on the level of deuterium incorporation as long

Poster Abstract 6
Diet-gut microbiota interactions mediated by hop-derived xanthohumols in a mouse obesity model

Johana Revel, Department of Chemistry, Linus Pauling Institute, Oregon State University
Liping Yang, Department of Chemistry, Oregon State University
Cristobal Miranda, College of Pharmacy, Linus Pauling Institute, Oregon State University
Joshua Hay, Oregon State University
Adrian Gombart, Department of Biochemistry and Biophysics, Linus Pauling Institute, Oregon State University
Jan F. Stevens, College of Pharmacy, Linus Pauling Institute, Oregon State University
Claudia Maier, College of Chemistry, Linus Pauling Institute, Oregon State University


Xanthohumol (XN) is the principal prenylated flavonoid produced by hops (Humulus lupulus L.). Recent studies conducted by our group with rodents indicate that XN supplementation in food could treat or mitigate metabolic syndrome and obesity. The low bioavailability of XN and flavonoids in general suggests that at least part of the observed health benefits could come from XN biotransformed metabolites generated by the gut microbiome and the liver. We hypothesize that diet supplementation with XN mitigates gut dysbiosis and that the biotransformation products of XN are partly responsible for the observed health benefits. To test this hypothesis, we developed mass-spectrometry-driven workflows for characterizing XN biotransformation products along with metabolites of interest in biofluids. We collected feces from high-fat diet fed C57BL/6J male mice supplemented or not with 30 mg/kg of XN or the XN derivatives, dihydroxanthohumol (DXN) or tetrahydroxantohumol (TXN). We analyzed fecal extracts using a Synapt G2 HDMS mass spectrometry platform coupled to a Waters Acquity I- class UPLC system. We developed a method for the identification and quantification of XN, DXN and TXN along with their metabolites as well as endogenous metabolites, such as bile acids (BAs). We focused primarily on BAs because they are important cholesterol-derived metabolites involved in multiple physiological processes, including fat absorption. DXN and TXN supplemented mice excreted lower amounts of the parent flavonoid and their metabolites compared to XN, which is in agreement with the higher bioavailability of DXN and TXN. Moreover, we observed an overall increase in conjugated bile salts in feces of mice supplemented with DXN and TXN, but not in feces from XN-treated mice, compared to vehicle-treated animals. These results indicate that DXN and TXN treatment enhances BA excretion at the expense of hepatic cholesterol.

Poster Abstract 7
A novel nutritional formulation containing the prebiotic human milk oligosaccharide 2’- fucosyllactose reduces gastrointestinal symptoms and beneficially alters the gut microbiome in adults with gastrointestinal dysfunction

Jennifer J. Ryan, National University of Natural Medicine
Kirti Salunkhe, The Functional Medicine Research Center, Metagenics, Inc.
Nikhat Contractor, The Functional Medicine Research Center, Metagenics, Inc.


Background: Gut dysbiosis, disruption in the homeostasis of the intestinal microbiota, contributes to the pathogenesis of many gastrointestinal disorders. Human milk oligosaccharides (HMOs), which are naturally occurring in human breast milk, are considered “bifidogenic” and “butyrogenic”. In breast-fed infants, they serve as primary substrates for select Bifidobacterium spp. and are metabolized into butyrate by butyrate-producing gut microbiota. UGIR is a formulation that provides nutritional support for adults with gastrointestinal dysfunction; it contains a combination of essential macro and micronutrients and prebiotics, including 2’-fucosyllactose (2’FL), the most abundantly produced HMO. This study reports novel data on the effect of 2’FL, in the context of a comprehensive nutritional formulation, in adults with gastrointestinal dysfunction. Methods: Adults with IBS, ulcerative colitis, Crohn’s disease or celiac disease were recruited from four U.S. medical practices. Participants received one serving of UGIR twice daily for six weeks. Outcome measures included the Gastrointestinal Quality of Life Index (GIQLI) and a stool analysis panel. Results: Twelve participants completed the study. GIQLI total score, gastrointestinal symptoms domain, and social function domain scores improved (P<0.05). Butyrate, acetate, and total SCFAs increased (P<0.05). Several commensal bacteria increased including Bifidobacterium spp., Bifidobacterium longum, Faecalibacterium prausnitzii, Aneurotruncus colihominis, and Pseudoflavonifractor spp. (P<0.05). Conclusions: UGIR consumption was associated with reduced gastrointestinal symptoms, increased fecal SCFAs, increases in several beneficial gut microbes (including species that have been shown to consume 2’FL as a substrate in vitro), increases in butyrate-producing species, and increases in species that have been previously shown to be low in patients with IBS and IBD. It is plausible that the improvements in butyrate levels and commensal gut microbiota contributed to the clinical benefits demonstrated on the questionnaire. These results suggest that UGIR is a promising novel nutritional formulation that could be used in the management of gastrointestinal dysfunction associated with gut dysbiosis.

Poster Abstract 8
Supplementation of mice fed a high fat diet with xanthohumol and its derivatives changes the composition of the fecal microbiota and is associated with observed metabolic improvements

Yang Zhang, School of Public Health and Human Sciences, Linus Pauling Institute, Oregon State University
Cristobal Miranda, College of Pharmacy, Linus Pauling Institute, Oregon State University
Thomas J. Sharpton, Department of Microbiology, Department of Statistics, Oregon State University
Gerd Bobe, Department of Animal Sciences, Linus Pauling Institute, Oregon State University
Johana Revel, Department of Chemistry, Linus Pauling Institute, Oregon State University
Paul Blakemore, Department of Chemistry, Oregon State University
Andriy Morgun, College of Pharmacy, Oregon State University
Natalia Shulzhenko, College of Veterinary Medicine, Oregon State University
Claudia S. Maier, Department of Chemistry, Linus Pauling Institute, Oregon State University
Jan F. Stevens, College of Pharmacy, Linus Pauling Institute, Oregon State University
Adrian F. Gombart, Department of Biochemistry and Biophysics, Linus Pauling Institute, Oregon State University


Despite our understanding of the underlying physiology and current prevention and treatment options, obesity still has reached epidemic proportions globally. The prenylated flavonoid xanthohumol (XN) found in hops and beer improves dysfunctional glucose and lipid metabolism in preclinical animal models of diet-induced obesity (DIO) and metabolic syndrome (MetS). We have shown that XN binds to the farnesoid X receptor (FXR) and regulates host genes involved in metabolism of cholesterol into bile acids and the cathelicidin antimicrobial peptide (CAMP) gene expression. In a 14-week study with C57BL/6J mice fed a high-fat diet containing XN, α,β-dihydro-XN (DXN) or tetrahydro-XN (TXN), the treatments resulted in abrogation of neuro-metabolic impairments. We hypothesize that consumption of XN or its hydrogenated derivatives by mice fed a high-fat diet induces CAMP and shapes the composition of the gut microbiota. Mice administered XN, DXN or TXN showed similar improvements of impaired glucose tolerance compared to the control; however, the derivatives decreased plasma insulin and leptin to a greater extent than XN. We sequenced the 16 rRNA genes of the microbes present in the feces of each animal. We observed statistically significant shifts in the structure and membership of the microbiota. The compounds significantly decreased the percentages of Bacteroidetes and Tenernicutes. In contrast, they significantly increased Firmicutes, Proteobacteria (DXN and TXN), and Verrucomicrobia (DXN and TXN). Specifically, Oscillospira (Firmicutes), an under-studied anaerobic bacterial genus, increased from 7% to 14% abundance after DXN and TXN supplementation. Moreover, we identified statistically significant correlations with weight gain, food intake, food efficiency, fasting glucose and levels of insulin and leptin with Oscillospira. We postulate that Oscillospira, along with other defined and undefined genera, may mediate some of the benefits from consumption of XN and its derivatives.

Poster Abstract 9
Hepatic and gut microbial metabolism of the hop hutraceutical, xanthohumol, in humans

Lea-Sophie Ullrich, College of Pharmacy, Linus Pauling Institute, Oregon State University
Ralph Reed, College of Pharmacy, Linus Pauling Institute, Oregon State University
Ines L. Paraiso, College of Pharmacy, Linus Pauling Institute, Oregon State University
Sandra Uesugi, Linus Pauling Institute, Oregon State University
Jan F. Stevens, College of Pharmacy, Linus Pauling Institute, Oregon State University


Xanthohumol, a prenylated flavonoid, is a yellow substance isolated from hops (Humulus lupulus) and found primarily in beer. Over the past years it has received a lot of attention due to its biological effects. This natural compound efficiently improves dysfunctional glucose and lipid metabolism in animal models of metabolic syndrome and could therefore have health protective actions against type 2 diabetes mellitus and cardiovascular diseases. In order to facilitate clinical studies in the future it is essential to establish a broad view of xanthohumol’s metabolism. To determine xanthohumol and its metabolites simultaneously, our group has developed an extraction method using a paper strip for improved analyte detection and quantitation followed by a sensitive and selective high pressure liquid chromatography tandem mass spectrometry method that separates xanthohumol, isoxanthohumol, α,β- dihydroxanthohumol, 6-prenylnaringenin, and 8-prenylnaringenin. The present study focused on xanthohumol and its metabolites found in human urine in a crossover, double-blinded, placebo- controlled human intervention study at three dose levels of xanthohumol (OSU Institutional Review Board approval #6119). Healthy human subjects consumed a non-alcoholic beverage without and with xanthohumol for three weeks, with a washout period of three weeks. Findings show a spontaneous cyclization of xanthohumol into isoxanthohumol, which can be followed by a hepatic or gut microbial demethylation into 8-prenylnaringenin, an estrogenic natural compound. α,β-dihydroxanthohumol and 6- prenylnaringenin were also detectable in urine from subjects taking xanthohumol. The detection of α,β- dihydroxanthohumol, which lacks estrogenicity, suggests that human gut microbiota have the capability to hydrogenate the α,β-unsaturated bond of xanthohumol.

Acknowledgment: This project was supported by TAXAN Inc., Wiesbaden, Germany.

Poster Abstract 11
Green tea treatment in obese mice with nonalcoholic steatohepatitis lowers hepatic NFκB activation in association with altered relative abundance of phosphatidylcholine and bile acid metabolites

Geoffrey Y. Sasaki, Human Nutrition Program, The Ohio State University
Jinhui Li, Human Nutrition Program, The Ohio State University
Morgan J. Cichon, Foods for Health Discovery Theme, The Ohio State University
Ken M. Riedl, Nutrient & Phytochemical Analytic Shared Resource ,The Ohio State University
Rachel E. Kopec, Human Nutrition Program, The Ohio State University
Richard S. Bruno, Human Nutrition Program, The Ohio State University


Green tea extract (GTE) limits NFκB-mediated inflammation during nonalcoholic steatohepatitis (NASH). Our objective was to identify shifts in the global hepatic metabolome associated with GTE-mediated lowering of NFκB activation during NASH. Male C57BL/6J mice were fed a low-fat (LF) or high-fat (HF) diet for 12 wk to induce NASH. They then continued on these diets supplemented with 0 or 2% GTE (n = 10/ group) for an additional 8 wk prior to assessing metabolomics profiles. GTE attenuated histological evidence of NASH (hepatic steatosis, hepatocellular ballooning), hepatic NFκB activation, and lipid peroxidation that were otherwise increased in HF controls. Principal component analysis indicated that GTE in HF-fed mice restored the hepatic metabolome that was otherwise shifted away from LF-fed mice. Compared with HF controls, 129 metabolites were altered (≥2-fold; P<0.01) in response to GTE. GTE in HF mice decreased (P<0.05) the relative abundance of phosphatidylcholine catabolites (e.g. lysophosphatidylcholine, glycerophosphocholine) that were otherwise increased in HF controls. Compared with HF controls, GTE increased primary and secondary bile acid metabolites (e.g. chenodeoxycholic acid, cholic acid, sulfoglycolithocholate) and decreased hepatic cholesterol to levels not different from LF mice. Phosphorylated p65 and hepatic MDA were correlated with phosphatidylcholine metabolites (P<0.05; r = 0.42-0.63) whereas bile acid metabolites were inversely associated with hepatic cholesterol and phosphorylated p65 (P<0.05; r = -0.39 to -0.65). These findings suggest that metabolic shifts by GTE treatment function to lower NFκB activation in NASH by limiting lysophosphatidylcholine-mediated hepatic injury and increasing bile acid metabolites that promote FXR activation.

Poster Abstract 12
Effects of dietary resveratrol on the fecundity of a novel model, Nothobranchius guentheri

Andrew C. Drake, Department of Biochemistry and Biophysics, Linus Pauling Institute, Oregon State University
Youngjoo Lee, Department of Biochemistry and Biophysics, Linus Pauling Institute, Oregon State University
Nicholas O. Thomas, Department of Biochemistry and Biophysics, Linus Pauling Institute, Oregon State University
Kate Petersen Shay, Linus Pauling Institute, Oregon State University


The loss of female reproductive ability occurs midway through the lifespan in humans and is an early phenotype for human aging. Yet the number of women delaying pregnancy or postponing decisions about reproduction continues to rise in industrialized societies due to personal or socioeconomic circumstances, often resulting in subfertility or difficulty conceiving. There are few defined mechanisms associated with this etiology and equally few effective therapies. We used a novel emerging model, Nothobranchius guentheri, with an age-associated spectrum of changes analogous to that found in human fertility to test a possible solution to this problem. Our hypothesis is that resveratrol (RSV) will activate SirT1, an oxidative stress sensor and longevity assurance enzyme, and thus improve female fecundity in mid-life. RSV, a polyphenol found in grapes and red wine, has been presented both commercially and in studies as an anti-aging dietary supplement due to its ability to prolong both lifespan and health span. SirT1 is an NAD+ dependent histone deacetylase, whose activity is regulated by the nicotinamide to NAD+ salvage pathway, especially by the rate-limiting enzyme NAMPT. We found that female N. guentheri fed 600 μg RSV/g food into mid-life (~20 weeks), beginning at sexual maturity, showed increased fecundity compared to those on Control diet. Furthermore, ovarian NAMPT was found at higher concentrations in the RSV-fed fish which we expect leads to greater availability of NAD+ for sirtuin activity. This suggests that dietary RSV has a positive effect on female fertility and that it may become an effective therapy to regulate sirtuin activity and combat reproductive senescence.

Poster Abstract 13
Antiviral activity of scorpion venom peptides

Huerta-E, V.M., PrevenOx Orthomolecular Biomedical Research
V. Banafsheh, H-Roostaeyan, PrevenOx Orthomolecular Biomedical Research


Arachnid venoms, utilized as a tool for defense and attack, by killing or immobilizing their predators, are composed by rich molecular diversity and complex mixture of proteins and peptides. More than 200 viruses are known to cause human diseases. Considered most common pathologies in humans, cardiovascular and infectious diseases and cancer are among the leading causes of deaths. The very few antiviral drugs commercially available can induce severe and considerable adverse effects, especially to those patients receiving lifelong treatment for diseases such as HIV. Furthermore, viruses possess rapid mutational capacity to trick and infect host cells. All these facts together have propelled the prospection for new antiviral drugs, particularly from natural products, as they constitute more than 25% of the new drug prototypes approved in the last decades. Among sources of natural products, scorpion venoms have revealed a great potential for drug discovery, and despite their harmful action mechanism most of them have components holding potential medicinal properties to cure diseases. It is widely reported that scorpion venoms are rich sources of antimicrobial substances, and contain a vast array of active biological compounds with distinct chemical structures. Scorpion venom is a complex mixture of hundreds of molecules, mostly peptides, which possesses a large array of biological activities betwixt: antiviral activity. In addition in scorpions the biologically active peptides are classified as disulfide-bridged peptides (DBPs) and non-disulfide-bridged peptides (NDBPs), with the former being the main components of scorpion venoms, responsible for the neurotoxic effects. Usually these DBPs target the ion channels of excitable and non-excitable cell membranes. These properties make these molecules interesting prototypes of drugs for the treatment of diverse diseases, particularly those affecting the neural system. In conclusion scorpion peptides exhibit direct virucidal activity. Because of the limited efficiency of commonly used drugs and emerging resistance of viruses, antiviral scorpion peptides may have the potential as putative therapeutic agents with antiviral activities.

Poster Abstract 14
To whom the side-effect of licorice, pseudoaldosteronism, will occur: A case-control study

Tetsuhiro Yoshino, Center for Kampo Medicine, Keio University, Linus Pauling institute, Oregon State University
Akihiro Komatsu, Department of Pharmacy, Keio University
Tomonori Nakamura, Faculty of Pharmacy, Keio University
Kenji Watanabe, Center for Kampo Medicine, Keio University


Background: Pseudoaldosteronism is a well-known side effect of licorice or glycyrrhizin (GL), one of major components in licorice. Pseudoaldosteronism mimics primary aldosteronism which causes hypokalemia, hypertension, and extremities’ edema. GL can be used to treat chronic hepatitis, but also plays important role in pseudoaldosteronism. The importance of multidrug resistance protein 2 (Mrp2) in GL excretion has been reported recently. Dysfunction of Mrp2 causes elevated direct bilirubin (DB), GL, and its metabolites. Hence, elevated DB can be a risk predictor of pseudoaldosteronism. The relationship between pseudoaldosteronism and elevated DB, however, has not been studied as yet. Objectives: To evaluate the relationship between elevated DB and hypokalemia, the most sensitive marker of pseudoaldosteronism.
Methods: This case-control study included patients with chronic hepatitis who visited the division of hepatology at Keio University Hospital between January 2009 and December 2015. Inclusion criterion was availability of laboratory data on DB and change in serum potassium.
Results: Data from 1392 patients (796 men) were used in the analysis, and the most common cause of chronic hepatitis was hepatitis type C virus infection. GL was used for 79 patients (GL+) and wasn’t for 1313 (GL-). Mean age was 60.5 ± 14.2 years for GL+, and 58.3 ± 15.8 years for GL-. Hypokalemia tended to be noted more in GL+ patients (OR 1.69, 95%CI:0.99-2.85, p=0.06). When we divided patients with or without elevated DB, hypokalemia was noted more in GL+ patients with elevated DB (OR 2.89, 95%CI:1.15-8.00, p=0.02), but not in GL+ patients without elevated DB (OR 0.84, 95% CI:0.31-2.20, p=1). From multi-variable logistic regression in GL+ patients, elevated DB and lower serum albumin level were related to hypokalemia, but age, sex, daily dose of GL, and duration of GL intake were not.
Conclusions: Elevated DB might be a predictor of pseudoaldosteronism due to GL.