**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

**POSTER SESSIONS 3 AND 4 – Wednesday and Thursday –** all even number posters will be on display.

*AWARD WINNERS

BIOMEDICAL

P-149

Viral and bacterial infection elicit distinct changes in plasma lipids in febrile children

PRESENTING AUTHOR: Xinzhu Wang, Imperial College London, United Kingdom

CO-AUTHORS: Ruud Nijman, Stephane Camuzeaux, Caroline Sands, Marieke Emonts, Jethro Herberg, Ian Maconochie, Matthew Lewis, Michael Levin, Myra McClure

Fever is the most common reason that children present to Emergency Departments (EDs) in the UK. Clinical signs and symptoms suggestive of bacterial infection are often non-specific, and no test exists for the accurate diagnosis of infection. As a result, many children with viral infection are prescribed antibiotics unnecessarily, while others with serious bacterial infections are not treated in a timely manner and progress to sepsis. In recent years, the 'omics' approaches to identifying biomarkers from the host-response to bacterial infection are proving promising. In this study, lipidomic analysis was carried out with plasma samples obtained from febrile children with confirmed bacterial infection (n=20) and confirmed viral infection (n=20). We show for the first time that bacterial and viral infection elicit distinct changes in the host lipidome. Glycerophosphoinositol, sphingomyelin, lysophosphotidylcholine and cholesterol sulate were increased in the confirmed virus infected group, while fatty acids, glycerophosphocholine, glycerophosphoserine, lactosylceramide and bilirubin were increased in cases of confirmed bacterial infection. A combination of 20 metabolites increased diagnostic performance and achieved the AUC value of 0.853 (95% CI, 0.672 - 0.995). This pilot study demonstrates the potential of metabolic biomarkers to distinguish bacterial from viral infection in febrile children, to facilitate effective clinical management and to limit inappropriate use of antibiotics.

P-150

The circulating metabolome in the progression to islet autoimmunity and type 1 diabetes

PRESENTING AUTHOR: Santosh Lamichhane, Turku Centre for Biotechnology, University of Turku, Finland

CO-AUTHORS: Santosh Lamichhane, Linda Ahonen, Thomas Sparholt Dyrlund, Esko Kemppainen, Heli Siljander, Heikki Hyöty, Jorma Ilonen, Jorma Toppari, Riitta Veijola, Tuulia Hyötyläinen, Mikael Knip, Matej Oresic

Type 1 diabetes (T1D) is a chronic autoimmune disease caused by specific destruction of the insulin-producing beta cell. Clinically pre-diabetic period in T1D is characterized by presence of beta cell specific autoantibodies. Intriguingly, earlier metabolomics study suggests specific metabolic disturbances before individuals precede to auto-immunity. Here we combine lipidomics and metabolomics approach to analyze circulating metabolites in a prospective series of plasma samples from 40 children who progressed to T1D (PT1D), 40 children who developed at least a single islet autoantibody but did not progress to T1D during the follow-up (P1Ab) and 40 matched controls (CTR). We found sphingomyelins and methionine to be persistently dysregulated in PT1D when compared to the P1Ab and CTR groups. Additionally, phosphatidylcholines and triacylglycerols were downregulated in PT1D as compared to P1Ab, while amino acids including glutamine, aspartic acid were downregulated in PT1D as compared to CTR at the age of 3 months. Furthermore, we found hydroxyphenyllactic acid, indole acetic acid, and 11-eicosenoic acid, metabolites of potential microbial origin, to be significantly downregulated at early age (3 and 6 months) preceding clinical T1D. Our study support findings from earlier studies and suggests novel metabolic signatures that specifically characterize children who progressed to islet autoimmunity or overt T1D, respectively, which may be helpful in the identification of at-risk children before the initiation of autoimmunity.

P-151

Baseline metabolic profiles of early rheumatoid arthritis patients achieving sustained drug-free remission after initiating treat-to-target TCZ, MTX or the combination: insights from systems biology

PRESENTING AUTHOR: Wei Yang, Leiden University, Netherlands

CO-AUTHORS: Xavier Teitsma, Amy C Harms, Alireza Mashaghi, Thomas Hankemeier

For patients with newly diagnosed rheumatoid arthritis (RA), treatment aim is early, rapid, and sustained remission. However, still a significant number of patients respond insufficiently to first-line drug methotrexate (MTX) or new biological drug tocilizumab (TCZ). Metabolic analysis prior to therapy could be potential indicators of disease activity and predictors of treatment response. The aim of this study was to identify relevant metabolites and biological pathways associated with achieving sustained drug-free remission (sDFR) after a treat-to-target TCZ- or MTX-based strategy. Baseline serum samples were analyzed from sDFR and non-sDFR patients of early RA. Metabolomic measurements were performed on oxidative stress, amine and oxylipin platforms followed by pathway analyses in each arm. In a result, baseline metabolic profiles of early RA were found to be different between sDFR and non-sDFR and associated pathways can indicate treatment-specific connections with drug mechanisms of MTX or TCZ. Integrated with our previous observations analyzing relevant transcripts and proteins within the same patients, network analysis showed good correlation across "omics" data. Therefore, signature metabolite biomarkers identified in early RA could potentially act as key prognostic factors for applying personalized care in the future.
**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

**POSTER SESSIONS 3 AND 4 – Wednesday and Thursday –** all even number posters will be on display.

*AWARD WINNERS

BIOMEDICAL

P-152

Analysis of Smoker's and Non-Smoker's Urine using enhanced two-dimensional chromatographic resolution and high performance TOFMS

PRESENTING AUTHOR: Nick Jones, LECO Europe BV, Netherlands

Urine is a favored biological fluid for medical testing, since it is easy to obtain in large quantities and provides a window into an individual's exposure, diet, and general health. In this study, a novel analytical approach based on comprehensive two-dimensional gas chromatography-high performance time-of-flight mass spectrometry was utilized for robust identification of compounds in two urine standard reference materials (smoker's and non-smoker's urine). After sample preparation and derivatization, the urine samples were analyzed using both GC-TOFMS and GCxGC-TOFMS. An increase of confidently identified compounds in urine standards was the direct result of transitioning from GC-TOFMS to high performance GCxGC-TOFMS, which yields cleaner spectra and thus improved spectral similarity scores. The Pegasus BT 4D facilitated fast and confident compound identification through enhanced two-dimensional chromatographic resolution and high performance TOFMS. GCxGC-TOFMS contour plots were highly structured, showing clustered classes of compounds and provided high quality spectral data, that were searched against large, well-established databases. Library hits were filtered using retention index software tools and findings further supported by calculating mass delta values for molecular and fragment ions. Comprehensive data was processed via non-targeted and targeted methods. Comparison of smoker's and non-smoker's results demonstrated increased quantities of tobacco related compounds, such as cotinine and trans-3'-hydroxycotinine, but also phenols, and additional nitrogen-containing compounds.

P-153

Enhanced Quantification of LPA 18:1 in Plasma with SelexION™ Ion Mobility Technology

PRESENTING AUTHOR: Cyrus Papan, Sciex Germany GmbH, Germany

Lysophosphatidic acid (LPA) is a signaling molecule in the class of lipid mediators, functioning by signaling through G-Protein coupled receptors or nuclear receptor proteins. LPA has been associated with a wide range of biological processes. The major bottleneck for accurate quantitation of LPA in plasma is the chemical interferences, using current analytical methods. The most abundant LPA fragments, the best fragments for quantification has strong matrix interferences, while the specific qualifier fragment, the fatty acid, shows very low signal intensity. Here we utilize SelexION Technology for the quantification of LPA 18:1 in plasma which removes the unspecific matrix interferences in plasma sample, allowing to use the abundant MRM transitions for quantitation of LPA with much greater confidence.

P-154

Metabolomics of congenital growth hormone deficiency (GHD) patients treated with recombinant human growth hormone (rhGH)

PRESENTING AUTHOR: Nilson Assunção, Unifesp, Brazil

CO-AUTHORS: Luciani Renata Silveira Carvalho, Emanuel Carrilho, Vinicius Guimarães Ferreria, Paulo Cesar Gonçalves Pereira, Breno San Martin

Diagnosis of GHD is commonly associated with the increase on the occurrence of cardiovascular disease in the patients, that frequently suffers from fatigue, reduced aerobic exercise capacity, abdominal obesity and reduced lean body mass. A variety of these features could be reversed by the use of recombinant human growth hormone (rhGH) replacement, however, the use of rhGH for long periods was previously reported to increase the risks of the patients to suffer cardiac problems. This work aims to better understand the effects of rhGH replacement treatment in the metabolism of the patients, in order to brighten the use of such treatment. Therein, we performed untargeted metabolomics on 46 GHD patients and 12 health controls serum samples. The metabolites were extracted from the serum samples through a protein precipitation protocol, and followed to direct infusion analysis on a high-resolution mass spectrometer (LTQ-Velos Orbitrap, Thermo-scientific). The resulting spectra were first aligned, using the m/z's, by an in-house build software, due the lack of commercial softwares able to handle with direct infusion data. The aligned spectra were then submitted to PCA and PLS-DA analysis using the MetaboAnalyst 4.0 tool. The multivariate analysis indicated a list of higher expressed metabolites in the GDH group, such as glycerol, trimethylamine, citrate, pyruvate, carnitine, betaine, glucose, glycine, tyrosine, leucine, choline. The accuracy of ROC Curve was satisfactory (AUC=0.974). Our preliminary result showed that the most expressive metabolites are related to the process of Lipolysis and the use of lipid reserves in bioenergetic processes.
<figure><img src="image_55.png" /></figure>

**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

**POSTER SESSIONS 3 AND 4 – Wednesday and Thursday –** all even number posters will be on display.

*AWARD WINNERS

BIOMEDICAL

P-155

From Molecular Profiling to Precision Medicine in Metabolic Syndrome

**PRESENTING AUTHOR:** Estelle Pujos-Guillot, Clermont Auvergne University, INRA, UNH, Plateforme d'Exploration du Métabolisme, MetaboHUB Clermont, France

**CO-AUTHORS:** Blandine Comte, Stéphanie Monnerie, Marion Brandolini, Cécile Canlet, Florence Castelli, Benoit Colsch, François Fenaille, Charlotte Joly, Natacha Lenuzza, Bernard Lyan, Jean-François Martin, Carole Migné, José A. Morais, Mélanie Pétéra, Etienne Thévenot, Christophe Junot, Pierrette Gaudreau, Estelle Pujos-Guillot

Metabolic syndrome (MetS), defined as a cluster of cardio-metabolic factors including obesity, hypertension, dysglycemia, and dyslipidemia, and mostly affecting older adults, is now a public health challenge because of its growing prevalence. In the context of personalized medicine/ nutrition, new tools are necessary to bring additional knowledge about MetS etiology, better stratify populations and customise strategies for prevention. The objective of this study was to investigate the integration of data from complementary untargeted metabolomics platforms (HRMS, RMN) and technologies to characterize the MetS phenotypic spectrum. A case-control study was designed within the Quebec NuAge cohort1. Six complementary untargeted metabolomic/lipidomic approaches, available within the MetaboHUB infrastructure2, were performed on serum samples collected at recruitment and 3 years later. Standard operating procedures were designed to guaranty the inter-laboratory standardisation from sample preparation to data processing. Data analyses were performed using reproducible online Galaxy workflows3. A full feature selection strategy was developed to build a comprehensive molecular MetS signature, stable over time. Consistent cross-sectional and longitudinal data were observed with a wide range of metabolites (lipids, carbohydrates, amino-acids, peptides...) reflecting subject stability regarding MetS, and providing new insights about underlying mechanisms. Correlation network analysis contributed to explore the links between the molecular signature and clinical parameters. Additionally, an optimized reduced signature was proposed, allowing good prediction performances (12% misclassification, AUC=0.96, CI:[0.94-0.98]), for future clinical application. These results demonstrated the interest of a multidimensional molecular phenotyping as part of the next generation of medicine tools in the frame of non-communicable diseases.

P-156

Faecal metabolomics by conventional UHPLC-HRMS as well as novel LA-REIMS reveals relevant metabolic perturbations in type 2 diabetes

**PRESENTING AUTHOR:** Lieven Van Meulebroek, Ghent University, Belgium

**CO-AUTHORS:** Cameron Simon, Lapauw Bruno, Takats Zoltan, Vanhaecke Lynn

Currently, treatments of type 2 diabetes hold some essential shortcomings, which concurs with the fact that this disease is still not curable nor a steady-state can be achieved. In addition, diagnostic tests have limited sensitivity and specificity, and are not ideal for large-scale screening. A better understanding of the underlying mechanisms of type 2 diabetes is therefore desired. For this purpose, our study implemented faecal metabolomics whereby faecal samples were collected from individuals with normal glycaemia (n=24) and type 2 diabetes (n=24) (UZ Ghent EC/2016/0673, HbA1c-based classification). Analytical methodologies employed conventional UHPLC-Q-ExactiveTM-MS as well as innovative Q-ToF-based LA-REIMS, with the latter having potential for point-of-care disease management and large-scale diagnostic screening. Based on the faecal UHPLC-HRMS fingerprints (6620 polar and 19831 lipophilic components), PCA-X score plots revealed clustering according to health state, which was confirmed by OPLS-DA modelling (p-values ≤ 1.86e-9 and Q2Y ≥ 0.654). Tentative identification of the discriminating metabolites revealed some interesting chemical classes, for which a role in type 2 diabetes was substantiated by literature. LA-REIMS was successfully developed and applied for faecal analysis, thereby using a FELS-25A IntelliguideTM CO2 laser. A major asset of the LA-REIMS approach concerned the short total acquisition time per sample, being < 0.5 min. Using this high-throughput approach, discrimination according to health status was possible, as indicated by the valid OPLS-DA model (p-value of 6.65e-17 and Q2Y of 0.767). In conclusion, for the first time, faecal metabolomics was successfully applied to characterize type 2 diabetes, using conventional UHPLC-HRMS and novel LA-REIMS.

P-157

Lipidomic signatures of non-alcoholic fatty liver disease

**PRESENTING AUTHOR:** Aidan McGlinchey, Örebro University, Sweden

**CO-AUTHORS:** Cecilia Carlsson, Quentin Anstee, Tuulia Hyötyläinen, Matej Orešič

Nonalcoholic fatty liver disease (NAFLD), a major risk factor for chronic liver disease and type 2 diabetes, has no non-invasive diagnostic techniques currently available for stages such as steatosis, nonalcoholic steatohepatitis (NASH) and fibrosis. We previously identified specific serum lipid signatures associating (1) with total liver fat1 and (2) NASH2. Here, we investigated lipidomic profiles of the European project Elucidating Pathways of Steatohepatitis (EPoS) cohort (n=688), comprising various stages of NAFLD (n=666), NASH (n=661) and fibrosis (n=511). In line with previous studies1, we show that steatosis grade strongly associates with (1) certain low-carbon-number, low-double-bond-count triglycerides increasing and (2) specific phospholipids decreasing. As NAFLD progresses from an earlier steatosis state to a later, severe fibrotic stage, fibrosis grades are used as clinical markers of progression to and severity of NASH. Preliminary analysis of 511 subjects with grades of fibrosis versus those without, revealed distinct relationships existing between circulating lipids and fibrosis stage, the profile changing appreciably between steatosis and fibrosis. We therefore suggest dysregulation of lipid metabolism across stages of NAFLD are reflected in circulation and may hold diagnostic value and insight into NAFLD pathogenesis. Further analysis of these markers is warranted and currently being undertaken taking these preliminary findings further toward diagnostic utility.
<figure><img src="image_56.png" /></figure>

**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

**POSTER SESSIONS 3 AND 4 – Wednesday and Thursday –** all even number posters will be on display.

*AWARD WINNERS

BIOMEDICAL

P-158

Identification of metabolite biomarkers of incident chronic kidney disease in Pre- and Type 2 Diabetes patients

PRESENTING AUTHOR: Jialing Huang, Helmholtz Zentrum München, Deutsches Forschungszentrum für Gesundheit und Umwelt (GmbH), Germany

CO-AUTHORS: Cornelia Huth, Martina Troll, Barbara Thorand, Christian Gieger, Jerzy Adamski, Annette Peters, Rui Wang-Sattler

Background and objective. Hyperglycemia exerts detrimental effects on the kidney and is one of the leading causes of chronic kidney disease (CKD). Identification of candidate metabolite biomarkers of CKD in hyperglycemic patients and evaluated their involvement in the pathogenesis of CKD are the aims of this study. Methods and Results. We used targeted metabolomics approach and examined longitudinal association of baseline concentrations of 125 metabolites with CKD incidence over 7 years in the KORA (Cooperation Health Research in the Region of Augsburg) cohort. We found that 19 metabolites nominally associated (P < 0.05) with incident CKD in 397 hyperglycemic participants in two multivariate logistic regression models by adjusting the effect of known CKD risk factors (e.g. age, systolic blood pressure, fasting glucose, serum triglycerides, baseline estimated glomerular filtration rate and baseline albumin-to-creatinine ratio). Furthermore, three metabolites were selected as candidate biomarkers via Priority-Lasso and a backward stepwise selection, and they were also FDR significant among 125 metabolites in both models. Higher levels of these three metabolites were found to increase the risk of incident CKD in hyperglycemia. Moreover, these three metabolites significantly improved the predictive value of the clinical predictors as the AUC (area under the receiver operating characteristic curve) value increased from 0.854 to 0.887(P = 0.037). Conclusion. We identified three novel metabolites to be predictors for CKD incidence in hyperglycemia, which may help develop new strategies to prevent CKD in hyperglycemic patients. Our results need to be replicated in an independent study.

P-159

Plasma phospholipid profile may reflect beneficial effect of physical activity on psychological stress

PRESENTING AUTHOR: Stefania Noerman, University of Eastern Finland, Finland

CO-AUTHORS: Elina Järvelä-Reijonen, Anton Mattsson, Urho Kujala, Sampsa Puttonen, Riitta Korpela, Marjukka Kolehmainen, Kati Hanhineva

Psychological stress seems to increase risk factors of metabolic diseases, but the underlying mechanisms have not been completely understood. Here we present the application of non-targeted metabolite profiling to reveal a panel of metabolites that may potentially intermediates of cross-talk between psychological and physiological health. Subjects prone to metabolic syndrome and psychological distress were recruited from three different cities in Finland and randomised to different 8-week lifestyle interventions. Metabolic profiling was performed on plasma samples of 64 controls and 60 subjects from face-to-face intervention group who showed the largest improvement of stress level between baseline (week 0) and follow-up (week 36). Non-targeted metabolite profiling was performed with high-performance liquid chromatography coupled with tandem mass spectrometry analysis. Physical activity seems to be associated with recovery and negatively associated with stress and adiposity. Several phospholipids that were elevated in the intervention group, including phosphatidylcholine (PC) (18:1/22:6), and two unknown PC (817.596@12.58 and 839.603@12.72) were associated with recovery and peak oxygen consumption (VO2max) and negatively associated with adiposity, stress index, and inflammation marker interleukin-1-receptor antagonist. Conversely, some other PC that were suppressed in intervention group including PC (16:0/16:1), (16:1/18:2), and (18:0/20:3), correlated positively with adiposity and negatively with sleep, recovery, and VO2max. The beneficial effect of physical activity on improvement of stress and body composition hence may be related to changes in the plasma profile of phospholipid, especially ones from PC class. Trial registration: The study was registered at ClinicalTrials.gov on August 17, 2012 with identifier NCT01738256.

P-160

The PXR-gut microbiota cross talk controls the host hepatic lipid metabolism in a sexually dimorphic way

PRESENTING AUTHOR: Sharon Barretto, INRA-Toxalim, France

CO-AUTHORS: Frederic Lasserre, Lorraine Smith, Celine Lucowikz, Arnaud Polizzi, Colette Bétoulières, Laurence Guzylack, Sandrine Menard, Elodie Person, Sandrine Bruel, Daniel Zalko, Cecile Canlet, Lindsay Peyriga, Edern Cahoreau, Laila Mselli-Lakhal, Nicolas Loiseau, Laurence Gamet-Payrastre, Hervé Guillou, Sandrine Ellero-Simatos

The pregnane X receptor (PXR) is a xenobiotic nuclear receptor, mainly expressed in liver and intestine. In germ-free mice, hepatic PXR activity is significantly reduced, suggesting PXR activation by gut microbiota-derived metabolites. However, its interaction with the gut microbiota remains unclear. We investigated this bi-directional interaction by looking into the effects of gut microbiota suppression by antibiotics integrating transcriptomics (liver, ileum), 1H-NMR based metabolomics (liver, caecal content), lipidomics (liver) and 16S sequencing (caecal content) in Pxr+/+ and Pxr-/- male and female C57BI6/J littermates. Microbiota suppression significantly decreased Pxr activity in the liver and ileum of males while only in the liver of females. Antibiotics significantly affected the hepatic 1H-NMR aqueous metabolic profiles of both Pxr+/+ and Pxr-/- mice. Antibiotics also decreased hepatic triglycerides, relative abundance of hepatic C16 and C16:1n-7 fatty acids and increased C20:1n-9 fatty acid profile of Pxr+/+ males compared to its control group. Similarly, mRNA levels of elongases (Elov1, 2, 3 and 5) were decreased in antibiotic-treated Pxr+/+ males only. Reciprocally, PXR deletion significantly affected gut microbiota composition, with increased proportion of Tenericutes in caecal content of Pxr-/- compared to Pxr+/+ males. Metabolic profiles of caecal content were also significantly different, with a tyrosine-derived gut microbiota metabolite significantly higher in Pxr-/- compared to Pxr+/+ males. No gut microbiota difference was observed in females. In conclusion, gut microbiota activates Pxr both in liver and in ileum of male mice. This crosstalk induces distinct changes in hepatic lipid composition influencing hepatic metabolism and gut microbiota, specific to males only.
**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

**POSTER SESSIONS 3 AND 4 – Wednesday and Thursday –** all even number posters will be on display.

*AWARD WINNERS

BIOMEDICAL

P-161

Intergrated analysis of bile acid metabolome and micriobiome for High-fat-diet effects

PRESENTING AUTHOR: Huiru Tang, School of Life Sciences, Human Phenome Institute, Fudan University, China

CO-AUTHORS: Hong Lin, Yanpeng An, Yulan Wang

Obesity has become a worldwide health issue though mechanistic aspects of obesity development remain to be fully understood. Here, we aim to elucidate the roles of bile acids and their associations with gut microbiota during obesity development with a high fat diet (HFD)-induced obesity rat model. We developed an UHPLC-MS method for bile acids quantification and analysed bile acids fluxes in multiple biological matrixes including feces, plasma, liver tissue and various segments of intestinal tissues. We found that, irrespective of dietary regimes, taurine-conjugated bile acids were the dominant species in the liver whereas unconjugated bile acids were in plasma. However, HFD caused slight increases in the total bile acids pool and particularly the increases in the levels of deoxycholic acid (DCA) (138.67 ± 37.225 nmol/L in controls, 242.61 ± 43.16 nmol/L in HFD group, p = 0.014) and taurodeoxycholic acid (TDCA) (2.8 ± 0.247 nmol/g in controls, 4.5 ± 0.386 nmol/g in HFD group, p =0.0018) in plasma and liver tissues, respectively, which were consistent with the increased levels of DCA in intestinal tissues and feces. These changes are correlated to an increase in abundance of genera Blautia, Coprococcus, Intestinimonas, Lactococcus, Roseburia, and Ruminococcus. Our investigation revealed the fluxes of bile acids and their association with gut microbiota during obesity development and explicatned unfavorable impact of HFD on health.

P-162

Using multivariate and multi-omics approaches to investigate autism spectrum disorder, asthma and Alzheimer's disease

PRESENTING AUTHOR: Beatriz Galindo-Prieto, Inst. for Computational Biomedicine, EIPM, Dept.of Physiology and Biophysics, Weill Cornell Medicine, United States

CO-AUTHORS: Jan Krumsiek

Asthma, autism spectrum disorder (ASD) and Alzheimer's disease (AD) affect millions of people worldwide. Multivariate and multiblock statistical models are widely used for finding relationships and patterns that can help to better understand the underlying biology, and thus move forward to try to reach a cure. Multi-omics data integration entails analyzing multiple -omics data matrices simultaneously by using multiblock approaches that find correlations among the blocks (i.e., data matrices) and relationships between descriptors (e.g., metabolites) and responses (e.g., clinical outcomes). Three examples are provided here: (i) A study carried out at NTNU (Norway) where is shown how multivariate methods such as principal component regression of eye and hand motion tracking data can help to better understand language processing in children with ASD (Vulchanova et al., manuscript in revision, 2019), (ii) a study from UMU and Karolinska Institute (Sweden) of the biological interactions in asthma using the MB- VIOP variable selection method (Galindo-Prieto, 2017) after modeling a 6-block multi-omics dataset (Reinke et al., 2018), and (iii) a currently ongoing study (Arnold et al., bioRxiv, 2019) from Weill Cornell Medicine (USA) with the goal to find correlations between blood and brain in a 4-block multi- omics Alzheimer's dataset. Acknowledgements: We acknowledge past financial support from IDS (Umeå University) (BGP), MKS Instruments (BGP), and the ERCIM 'Alain Bensoussan' Fellowship Programme (BGP), and current financial support from the National Institute on Aging and Weill Cornell Medicine (JK, BGP). We also want to thank Prof. R. Kaddurah-Daouk and the Alzheimer Disease Metabolomics Consortium for their support.

P-163

New biomarkers of 3-hydroxy-3-methylglutaryl CoA lyase deficiency

PRESENTING AUTHOR: Štěpán Kouřil, University Hospital Olomouc, Czech Republic

CO-AUTHORS: Jan Václavík, Lucie Mádrová, Radana Karlíková, David Friedecký, Štěpán Kouřil, Leo AJ Kluijtmans, Ron A Wevers, Tomáš Adam

3-OH-3-Me-glutaryl-coenzyme A lyase deficiency (HMGCLD) is a rare inherited metabolic disorder caused by mutations in HMGCL gene. The mitochondrial enzyme is responsible for catalysing the cleavage of HMG-CoA to acetyl-CoA and acetoacetic acid. Diagnosis is established by tandem MS based newborn selective metabolic screening that contains SRM transition for C5-OH carnitine (262 → 85) which represents four different compounds (Me-malonylcarnitine, 3-OH-isovalerylcarnitine, succinylcarnitine and 2-Me-3-OH-butyrylcarnitine) pointing to different inborn errors of metabolism (IEM). In order to distinguish between these IEM organic acid profiles of urine samples must be acquired. We run untargeted metabolomic analysis of five plasma samples from HMGCLD patients together with a group of controls with the aim to find new diagnostic biomarkers that would be detectable even via newborn screening. Apart from known biomarkers such as 3-OH-isovalerylcarnitine and 3-Me- glutarylcarnitine, other acylcarnitine (AC) species derived from intermediates in the leucine degradation pathway were observed. Furthermore, organic acid counterparts of these AC were also significantly increased compared to controls. Newly found elevated AC could hypothetically be also expected in one other condition, 3-Me-glutaconyl-CoA hydratase deficiency (MGCA). HMGCLD could be distinguished from MGCA by measuring of 3-OH-3-Me-glutarylcarnitine or 3-OH-3-Me-glutarate. The result suggests that 3-OH-3-Me-glutarylcarnitine and 3-OH-3-Me-glutarate could be specific biomarkers of HMGCLD in dried blood spots by FIA-MS screening and thus speed up differential diagnoses among all the diseases screened by C5-OH transition. That would allow for earlier introduction of dietary intervention directly from first screening sample. The work was supported by the Czech Science Foundation Grant (18-12204S) and NPUI (LO1304).
**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

**POSTER SESSIONS 3 AND 4 – Wednesday and Thursday –** all even number posters will be on display.

*AWARD WINNERS

BIOMEDICAL

P-164

Detail LC-MS/MS analysis of organic acids, acylglycines and acylcarnitines for targeted metabolomic profiling

PRESENTING AUTHOR: David Friedecký, University Hospital and Palacky University Olomouc, Czech Republic

CO-AUTHORS: Jaroslava Jáčová, Kateřina Mičová, Tomáš Adam

Analysis of organic acids offers an important insight into central metabolism. It has traditionally been performed using routine GC-MS platform that requires lengthy sample preparation and allows analysis of just a small batches of samples. The GC-MS under common conditions is not capable to analyze acylcarnitines (together with organic acids) and acylglycines as polar compounds suffer from poor detection. The LC-MS/MS multiple reaction monitoring (MRM) method was developed on the UltiMate (Dionex) LC triple quadrupole 6500 (Sciex) MS for the analysis of 124 organic acids, acylcarnitines and acylglycines in biological matrices. Under acidic conditions of formic acid on non-polar C18 column, good separation of isomers was obtained. Through MRM transitions, detection and quantification of compounds present in urine, serum and intracellular content were enabled. The run time takes 26 min. Analytes show good linearity (r2 > 0.99) and reproducibility (CV < 10%) of results. The method was tested on metabolomic experiment on plasma samples of glutaric aciduria I and maple syrup urine disease, where good segregation of groups was obtained, and the diagnostic markers of the disorders were found as the most discriminating compounds. The method is also usable for diagnostic purpose to find inborn errors of metabolism, where selected diseases were tested (SCADD, methylmalonic, glutaric and isovaleric aciduria). The method that minimizes GC-MS drawbacks can be applied as a sensitive and simple approach for targeted clinical metabolomics. It was approved using samples of patients with inherited metabolic disorders. Moreover, it enables accurate and early diagnosing more than 60 disorders.

P-165

Correlation between the metabolome and mtDNA heteroplasmy in mitochondrial disease

PRESENTING AUTHOR: Marianne Venter, North-West University, South Africa

CO-AUTHORS: Jeremie Zander Lindeque, Kazuto Nakada, Roan Louw

Primary mitochondrial disease caused by variation in mitochondrial DNA (mtDNA) pose difficulties in diagnostics and treatment, due to extensive heterogeneity in clinical presentation of patients: different mtDNA mutations can lead to similar clinical presentations, while the same mtDNA mutation can present with distinct and unrelated clinical phenotypes. Factors involved in this clinical heterogeneity may include heteroplasmy levels (where two or more species of mtDNA can co-exist in a single mitochondrion/cell/tissue), haplogroup context, or other adaptive mechanisms, but remain largely unclear. Only one mouse model has successfully been engineered to harbour an mtDNA deletion, known as ΔMitomice. This mouse model has a large mtDNA deletion and presents with a Leigh Syndrome-like phenotype at high heteroplasmy levels, making it useful for in vivo investigations of mitochondrial disease. The aim of this study was to give new insight into the role of heteroplasmy levels of defective mtDNA in disease progression and severity, using the ΔMitomice disease model. We employed an untargeted metabolomics approach to investigate the changes in metabolite levels along an mtDNA heteroplasmy gradient (0-80% heteroplasmy in tail snips). While some metabolites correlated in a linear fashion with heteroplasmy levels, other metabolites indicated a sudden increase at 60% heteroplasmy, indicating the influence of a threshold effect on these compounds. To our knowledge, this is the first study to utilise linear statistics to investigate the relationship between mtDNA heteroplasmy levels and the metabolome. The findings identified specific metabolites that can be utilised to monitor disease progression and treatment efficacy in mitochondrial disease patients.

P-166

Workflow and normalization strategies employed on CHRIS epidemiological study for targeted metabolomics

PRESENTING AUTHOR: Vinicius Veri Hernandes, Researcher, Italy

CO-AUTHORS: Johannes Rainer, Andrew Hicks, Peter Pramstaller, Sigurdur Smarason

The Cooperative Health Research in South Tyrol (CHRIS) is a population-based study which aims to investigate mainly neurological and cardiovascular conditions, as well as the process of ageing. More than 13,000 volunteers have been part of the program, which includes clinical tests (blood glucose, antinuclear antibodies, iron metabolism markers and liver function markers, among others), psychological and life style surveys and blood sampling for genotyping and metabolomics study. Using both targeted and untargeted approaches, the objective is to enhance metabolite coverage, maximizing the information obtained for this cohort. Targeted method is based on the Absolute IDQ p180 Kit from Biocrates, which provides a reproducible workflow for sample preparation and the quantification of around 180 metabolites through a liquid chromatography - mass spectrometry (LC-MS) system. The workflow is optimized according to the LC-MS equipment employed at the analysis step (in this case, a QTRAP 6500 from AB Sciex) and each sample set is prepared in the way that 2 derivative subsets are acquired. First subset is used for reversed phase analysis (covering amino acids and biogenic amines) and the second is submitted to a flow injection analysis (FIA) for accessing hexose, acyl carnitines and glycero and sphingolipids. Here we present the overall workflow and methodology for the targeted metabolomics approach of the CHRIS study and results from the employed normalization strategy and quality assessments.
*AWARD WINNERS

BIOMEDICAL

P-167

Mutations in PCYT2 disrupt etherlipid biosynthesis and cause a complex hereditary spastic paraplegia

PRESENTING AUTHOR: Fred Vaz, Amsterdam UMC, Netherlands

CO-AUTHORS: John H. McDermott, Mariëlle Alders, Saskia B. Wortmann, Stefan Kölker, Mia L. Pras-Raves, Martin A. T. Vervaart, Henk van Lenthe, Angela C. M. Luyf, Hyung L. Elfrink, Kay Metcalfe, Sara Cuvertino, Peter E. Clayton, Rebecca Yarwood, Martin P. Lowe, Simon Lovell, Richard C. Rogers, Antoine H. C. van Kampen, Jos P.N. Ruiter, Ronald J. A. Wanders, Sacha Ferdinandusse, Michel van Weeghel, Marc Engelen, Siddharth Banka

CTP:phosphoethanolamine cytidylyltransferase (ET), encoded by PCYT2, is the rate-limiting enzyme for phosphatidylethanolamine synthesis via the CDP-ethanolamine pathway. Phosphatidylethanolamine is one of the most abundant membrane lipids and is particularly enriched in the brain. We identified five individuals with biallelic PCYT2 variants clinically characterised by global developmental delay with regression, spastic para- or tetraparesis, epilepsy and progressive cerebral and cerebellar atrophy. Using patient fibroblasts we demonstrated that these variants are hypomorphic, result in significantly decreased ET protein levels and enzyme activity without affecting mRNA levels. The significantly better survival of hypomorphic CRISPR-Cas9 generated pcyt2 zebrafish knockout compared to a complete knockout, in conjunction with previously described data on the pcyt2 mouse model, indicates that complete loss of ET function may be incompatible with life in vertebrates. Lipidomic analysis revealed profound lipidomic abnormalities in patient fibroblasts impacting both neutral etherlipid and etherphospholipid metabolism. Plasma lipidomics studies also identified changes in etherlipids that have the potential to be used as biomarkers for ET deficiency. In conclusion, our data establish PCYT2 as a disease gene for a new complex hereditary spastic paraplegia and confirm that etherlipid homeostasis is important for the development and function of the brain.

P-168

HPLC-MS/MS acylcarnitine profiling gives insight into mitochondrial and peroxisomal processes of amino acid and fatty acid metabolism

PRESENTING AUTHOR: Pieter Giesbertz, Department of Nutritional Physiology, Technische Universität München, Germany

CO-AUTHORS: Josef Ecker, Alexander Haag, Jarlei Fiamoncini, Hannelore Daniel

Acylcarnitines are intermediates of fatty acid and amino acid metabolism derived from the conversion of Acyl-CoA species by the enzymatic action of carnitine acyltransferases. In contrast to CoA species, acylcarnitines are present in plasma and urine. In these body fluids, they are important diagnostic markers for inherited diseases of peroxisomal and mitochondrial oxidation. Although a direct injection tandem mass spectrometry might be suitable as a high-throughput strategy, for instance in neonatal screening, the quantification of isomeric and low-concentrated compounds can become challenging. Here, we describe an HPLC-supported MS/MS method for the comprehensive quantification of 85 acylcarnitine species. The method includes most amino acid-derived compounds, as well as odd-numbered carbon species, like pentadecanoyl- (C15) and heptadecanoylcarnitine (C17). Within the context of the metabolic syndrome, the method was applied to human plasma and murine plasma and tissue samples of obesity, insulin resistance and type 2 diabetes. In samples from both species, significant changes were found for acylcarnitine species derived from branched-chain amino acid metabolism. In addition, significant changes in acylcarnitine species derived from peroxisomal alpha and omega oxidation pathways were observed. In a second step, the rather unexplored pathways of peroxisomal fatty acid metabolism were studied in more detail in murine models of peroxisomal stimulation. In conclusion, the method described here extends the current spectrum of acylcarnitine species as markers of mitochondrial and peroxisomal metabolism. The application to human and murine samples gives novel insight in the changes of mitochondrial and peroxisomal metabolism in conditions of aberrant health.

P-169

Ultra high performance tandem mass spectrometric determination of steroid hormones in adipose tissues of two species

PRESENTING AUTHOR: Jutta Lintelmann, Helmholtz Zentrum München, Research Unit Molecular Endocrinology and Metabolism, Germany

CO-AUTHORS: Jutta Lintelmann, Katharina Schuh, Helga Sauerwein, André Tchernof, Janina Tokarz, Cornelia Prehn, Jerzy Adamski

The reliable quantification of steroid hormones in various tissues is an indispensable tool for the investigation and elucidation of metabolic processes involved in course and development of many diseases. We developed and applied a UHPLC-MS/MS method for the determination of 17 steroid hormones in bovine and human adipose tissue. The method includes homogenization and methanolic extraction of the steroids followed by robotized solid phase extraction on well plates of the AbsoluteIDQ® Stero 17 Kit from Biocrates (Biocrates Life Sciences AG). Resulting extracts are separated on a core-shell column (Kinetex XB-C18, 1.7 µm, 150 x 2.1 mm I.D., Phenomenex) using an acetonitrile-methanol gradient and 0.1 % formic acid at 0.3 mL/min and 45 °C (1290 Infinity II, Agilent). Analytes are detected by using a triple quadrupole mass spectrometer (QTRAP 5500, Sciex) with electrospray ionization in positive and negative MRM mode. Method validation resulted in recoveries between 20 % and 63 % making the use of individual internal standards constraining. Intra- and interday imprecisions were acceptable with coefficients of variation from 3 % to 18 %; and detection limits were between 0.01 ng/g and 0.5 ng/g adipose tissue. To demonstrate the applicability of the method, human and bovine subcutaneous adipose tissues were analyzed. First results showed concentrations for the steroid hormones detected lying between 0.07 ng/g (11-deoxycorticosterone) and 15 ng/g (dehydroepiandrosterone) in human adipose tissue and between 0.04 ng/g (estradiol) and 46 ng/g (progesterone) for bovine adipose tissue. The respective patterns and concentrations were highly different between the two species compared.
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P-170

Salivary metabolomics as a new tool for unravelling obesity pathways in adolescents

PRESENTING AUTHOR: Kathleen Wijnant, University of Ghent, Belgium

CO-AUTHORS: Kathleen Wijnant, Lieven Van Meulebroek, Beata Pomain, Stefaan De Henauw, Nathalie Michels, Lynn Vanhaecke

High prevalence and treatment resistance of obesity urges further exploration in early diagnosis and prevention. This is especially important at young age, when metabolic and psychological development are ongoing. Indeed, genetic susceptibility, bacterial symbiosis, but also environmental factors favor obesity. Researchers struggle with the complex processes towards disease susceptibility. Therefore, this project aims to elucidate pathways involved in obesity and obesity-related comorbidities by using salivary metabolomics, both targeted and untargeted. As saliva can be non-invasively collected while its metabolic composition parallels that of blood, it is a very interesting bio-fluid for both research, diagnosis and prognosis. Following the development of a salivary UHPLC-Q-HRMS profiling and fingerprinting method, its 'fit-for-purposeness' was proven through comprehensive validation (targeted 33 endogenous components, untargeted 8564 components) in which excellent precision (CV <15% or 30% respectively), linearity (R2 >0.99 or 0.90 respectively) and recovery were observed. The potential of this method was demonstrated in a first setup, where 13 obese were compared to 13 healthy children (10-16y). The constructed OPLS-DA model enabled us to discriminate adolescents (obese versus healthy weight), reflected by a promising Q2 of 0.669 and R2 of 0.541, good permutation testing and CV-ANOVA p <0.001. Subsequently, 6 metabolites with differentiating potential were retained with satisfying VIP-score (≥1), jack-knifed confidence interval and S-plot descriptors. Integrative metabolomics data analysis strategies will be applied for the identification of differentiating metabolites, pathway analysis and network mapping. This first model proves the potential of salivary metabolomics as a tool for unravelling mechanistic information and biomarker discovery in obesity.

P-171

Lipid remodelling of adipocyte cell membranes is associated with transcriptional and epigenetic changes during genetic and diet-induced obesity in mice

PRESENTING AUTHOR: Julian Griffin, Imperial College London, United Kingdom

CO-AUTHORS: Ke-di Liu, Animesh Acharjee, Lee D. Roberts, Melanie K Gulston, Antonio Murgia, Xinzhu Wang, Yajing Chu, James A. West, Robert C Glen, Andrew J. Murray

Obesity is a complex disorder where the genome interacts with diet and environmental factors to influence ultimate body mass, composition and shape. Numerous studies have investigated how bulk lipid metabolism of adipose tissue changes with obesity, and in particular how the composition of triglycerides (TGs) change with increased adipocyte expansion. However, reflecting the analytical challenge posed by examining non-TG lipids in extracts dominated by TGs, the glycerophospholipid (PLs) composition of cell membranes has been seldom investigated. We conducted a comprehensive lipidomic study of white adipose tissue in mice who become obese either through genetic modification (ob/ob), diet (high fat diet) or a combination of the two using solid phase extraction and LC-MS/MS to study the lipidome and targeted methods to study metabolites involved in one-carbon metabolism and the methylation of cytosine. Pre-fractionation of lipids using SPE allowed the characterisation of a diverse range of PLs which would normally be a minor component of a total lipid extract from adipose tissue. We demonstrate that the changes in TGs that dominate the overall lipid composition of white adipose tissue are distinct from the compositional changes of PLs, the predominant components of the cell membranes. PLs correlate better with transcriptional and one-carbon metabolism within the cell. To investigate epigenetic changes of the genome we developed a targeted analysis of methyl-cytosine and cytosine from DNA. This demonstrates that both genotype and diet alter global methylation of DNA, suggesting the compositional changes that occur in cell membranes have diverse functional consequences during adipocyte expansion.

P-172

Atlas of Circadian Metabolism Reveals System-wide Coordination and Communication between Clocks

PRESENTING AUTHOR: Anna Artati, Helmholtz Zentrum Muenchen, Germany

CO-AUTHORS: Kenneth A. Dyar, Dominik Lutter, Nicholas J. Ceglia, Yu Liu, Danny Armenta, N. Henriette Uhlenhaut, Pierre Baldi, Jerzy Adamski, Matthias H. Tschöp, Kristin Eckel-Mahan, Paolo Sassone-Corsi

Previous studies revealed that circadian clocks are integral part of the regulation of biological processes in all tissues of organisms. Advanced knowledge of the molecular and cellular underpinnings of circadian biology indicated that circadian disruption can play a role in a wide range of pathologies. Metabolic diseases are often characterized by circadian misalignment in different tissues in organisms. However, how altered coordination and communication among tissue clocks relate to specific pathogenic mechanisms remains largely unknown. In this study we present a temporal and spatial atlas of circadian metabolism in the context of systemic energy balance and under chronic nutrient stress (high-fat diet [HFD]) by performing 24-hours metabolomics profiling of eight mouse tissues simultaneously. Suprachiasmatic nucleus, medial prefrontal cortex, gastrocnemius skeletal muscle, interscapular brown adipose tissue, epididymal white adipose tissue, liver, serum, and cauda epididymal sperm were collected every 4 hours across the light/ dark cycle from a single cohort of C57BL/6J mice after 10 weeks of ad-libitum access to standard chow or HFD. The metabolites in each sample were monitored and relatively quantified with UHPLC-MS/MS and GC-MS/MS. Comparative analysis reveals how the tissue metabolic pathways are linked and gated to specific temporal and spatial windows and how tissue-specific and inter-organ dynamic clocks emerge by nutrient challenge. Overall, we show in this study how dynamic metabolic relationships among tissues can be reconstructed across time and space and how integration of circadian metabolomics data from multiple tissues can improve our understanding of health and disease.
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P-174 Analytical pipeline to determine the healthy fecal metabolome in cryogenically collected samples

PRESENTING AUTHOR: Kajetan Trost, Steno Diabetes Center Copenhagen, Denmark

CO-AUTHORS: Linda Ahonen, Tommi Suvitaival, Nina Christiansen, Trine Nielsen, Maja Thiele, Suganya Jacobsen, Aleksander Krag, Peter Rossing, Torben Hansen, Lars Ove Dragsted, Cristina Legido-Quigley

Introduction: Fecal metabolomics is a valuable tool to discover links between fecal microbiota and disease and it is especially pertinent for research in gastrointestinal disorders, clinical nutrition and metabolic syndromes. Large interindividual differences exist in the fecal composition, however little is known about the actual within-sample heterogeneity in healthy sample donors. Methods: We collected and immediately froze one stool sample from 10 healthy volunteers and cryogenically drilled every sample in four areas along the specimen. We detected relatively polar metabolites by derivatization followed by two-dimensional gas chromatography and time of flight mass spectrometry, while we analyzed lipids using ultra high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry. Metabolite features with analytical variation above 30% were removed, the ratio of between subjects to within subject variation was used to assess sample homogeneity. Results: After exclusion of 2326 features that could not be annotated, we detected a total of 298 molecular features, of which 182 showed analytical variation x< 30%. Metabolites included amino acids, fatty acid derivatives, carboxylic acids and phenolic compounds. Lipids predominantly belonged to the groups of diacylglycerols, triacylglycerols and ceramides. As many as 79% (144 of 182) molecules showed less variability within the same individual than the variability observed between subjects. Conclusions: Feces show extensive biological variation, both within and between healthy individuals. Initial results show that molecules in abundance are those synthesized by the microbiome, ceramides, triacylglycerols and diacylglycerols.

P-175 Bariatric surgery impacts metabolism of branched chain amino acids and branched chain fatty acids

PRESENTING AUTHOR: Alicja Pakiet, University of Gdansk, Poland

CO-AUTHORS: Maciej Wilczyński, Olga Rostowska, Justyna Korczyńska, Patrycja Jabłońska, Łukasz Kaska, Monika Proczko-Stepaniak, Piotr Stepnowski, Tomasz Śledziński, Adriana Mika

Bariatric surgery is the most effective form of treatment of morbid obesity leading to substantial weight loss and remission of type 2 diabetes. In the targeted metabolomic analysis of serum amino acids by liquid chromatography–mass spectrometry (LC-MS) we investigated the effect of omega-loop gastric bypass (OLGB) in obese patients (n=50) at the time of the surgery and at 6-9 months after the surgery, and in lean controls (n=30). Obese subjects were characterized by elevated levels of branched chain amino acids (BCAA) – leucine and isoleucine, while post-OLGB patients exhibited reduced valine, leucine and isoleucine content. BCAA concentrations were positively correlated with marker HOMA-IR (R=0.3, p=0.007), proving the impact of BCAA levels on insulin resistance. Furthermore, we used gas chromatography–mass spectrometry (GC-MS) lipidomic approach to profile fatty acids in serum of study subjects. Statistical analysis showed significant increase in levels of several iso- and anteiso- (BCFA) in post-OLGB patients, which was negatively correlated with HOMA-IR (R=0.28, p=0.012). BCFA are beneficial to health due to their anticancer, immunosuppressive and antibacterial properties. Since BCAA can act as precursors for synthesis of BCFA, levels of mRNA of enzymes associated with BCAA catabolism were investigated. We found that visceral adipose tissue expression of branched-chain keto acid dehydrogenase E1 subunit beta (BCKDHB) and branched chain amino acid transaminase (BCAT) 1 and 2 was severely reduced in obese individuals. It is therefore possible that the improvement in levels of beneficial BCFA following bariatric surgery is a result of normalization of BCAA metabolism in adipose tissue.
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BIOMEDICAL

P-176

Diet or surgery - A multi-platform metabolomics approach aiming to unravel dysregulated metabolic pathways in type 2 diabetes

PRESENTING AUTHOR: Katharina Herzog, Lund University, Sweden

CO-AUTHORS: Mahmoud Al-Majdoub, Johan Berggren, Andreas Lindqvist, Jan Hedenbro, Leif Groop, Nils Wierup, Peter Spégel

Type 2 diabetes (T2D) is a devastating, chronic metabolic disease that presently affects 8.5% of the world's population, resulting in an unmet need for improved treatments. Roux-and-Y gastric bypass surgery (RYGB) is an effective approach to induce weight loss and T2D remission in obese individuals. The mechanisms driving T2D remission are, however, not fully understood. In addition, it is debated whether a low-calorie diet (LCD) may induce the same effects observed after RYGB. In this study, we used blood plasma from morbidly obese individuals with (n=10) or without T2D (n=9) to investigate the alterations in metabolite levels in response to LCD and subsequent RYGB. We applied three complementary metabolomics platforms based on ultrahigh-performance liquid chromatography (UHPLC) coupled to quadrupole time-of-flight mass spectrometry (QTOF-MS). Together, these platforms cover >300 identified metabolites, including a variety of lipids, low molecular weight metabolites, i.e. amino acids, and metabolites with intermediate polarity, i.e. acylcarnitines. After data processing using commercial and open-source tools and in-house libraries, we applied multivariate data analyses techniques such as linear mixed-effect regression models and orthogonal projections to latent structures effects projection (OPLS-EP), an approach that allows multivariate modeling of dependent samples. Our data suggest that the effects of LCD, and the combination of LCD and RYGB, elicit different changes in the fasting metabolic state. These effects were driven by changes in acylcarnitines and various lipids. Furthermore, analyzing metabolic profiles in response to mixed-meal tests identified differences between individuals with and without T2D, which may aid unraveling the mechanisms driving T2D remission.

Functional Microbiomics

Nutrition – Microbiome – Host Interaction in Health and Disease

BIOCRATES

LIFE SCIENCES

THE DEEP PHENOTYPING COMPANY

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• Neurology

• Oncology

• Gastroenterology

• Cardiology

NEW
MxP® Quant 500 Kit

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Choline Metabolism

• Choline, TMAO

• Betaine, homocysteine

Tryptophan Metabolism

• Tryptophan, kynurenine

• Serotonin, indoles

Cholesterol Metabolism

• Secondary bile acids

• Cholesteryl esters

Fatty Acid Metabolism

• Eicosapentaenoic acid (ω-3)

• Arachidonic acid (ω-6)
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P-177*

Profiling the effect of Adult Onset Hypothyroidism (AOH) on the mouse heart in both sexes integrating untargeted GC- and LC-MS metabolomics

**PRESENTING AUTHOR:** Maria-Konstantina Ioannidi, Metabolic Engineering & Systems Biology Lab, Institute of Chemical Engineering Sciences, FORTH, Greece

**CO-AUTHORS:** Marigoula Margarity, Maria I. Klapa

**Background:** The present study is part of a multi-tissue investigation (brain, heart, liver) of the effect of Adult-Onset-Hypothyroidism (AOH) on the metabolic physiology of a chemical induction mouse model, intending to comparatively reconstruct the metabolic fingerprint of this pathophysiology at the organism level in both sexes. Here, we present the heart metabolic profiling, integrating gas and liquid chromatography-mass spectrometry (GC-/LC-MS)-based metabolomics with systems and network biology approaches. **Methods:** Tissue collection of 24 mice (male/female in sets of six euthyroid/hypothyroid) was performed on the 124th postnatal day, after applying an optimized tissue perfusion protocol with saline solution. An adapted to mouse heart metabolite extraction protocol was performed. Standardized data deposition and analysis was conducted using our software suite, M-IOLITE, and multivariate statistics tools. The differential metabolites were visualized within the mouse heart primary metabolism network that we have reconstructed from literature and our data. **Results and Discussion:** Indeed, the integration of GC-MS with LC-MS metabolomic analyses provided a broader overview of the mouse heart central carbon metabolism under AOH. Our results validated a lipid-centric metabolism for the heart, which becomes de-regulated under AOH characterized by decreased energy metabolism and TCA cycle activity. No extended inter-sex differences were observed, with the female exhibiting greater metabolic AOH insulation through an increased lipid oxidation activity. These results will be presented in comparison with the already acquired brain region metabolic profiles of the same animals towards a multi-tissue AOH metabolic physiology model. **Acknowledgments:** The work was supported by BITAD_MIS_5002469, ELIXIR-GR_MIS 5002780, EATRIS-GR_MIS 5028091, INSPIRED_5002550.

P-178

High-performance metabolomics diagnostic model for Non-alcoholic fatty liver disease (NAFLD)

**PRESENTING AUTHOR:** Takeshi Kimura, St. Luke's International University, Japan

**CO-AUTHORS:** Masahiro Nojima, Yutaka Aoki, Kuniyoshi Hayashi, Hirotaka Fujimoto, Yuji Heike, Mariko Asami, Kazuhiko Suzuki, Kevin Urayama, Masaaki Matsuura, Takaaki Sato, Katsunori Masuda

Non-alcoholic fatty liver disease (NAFLD) is an important disease due to its association with a broad range of disorders. We aimed to identify metabolome biomarkers for NAFLD using gas chromatography/mass spectrometry (GC/MS) and develop a diagnostic model for disease prediction. **Methods:** We enrolled 3,733 visitors of St. Luke International Hospital, Center for Preventive Medicine (Tokyo, Japan) between October 2015 and September 2016 who applied exclusion criteria for NAFLD diagnosis, e.g. daily alcohol intake (Men≥30g, Women≥20g), concurrent liver disease or malignancy, medication for metabolic diseases. Fatty liver was diagnosed by ultrasonography. Metabolome analysis was performed on blood serum samples using the GCMS-TQ8040 (Shimadzu, Japan). **Results:** The mean age of the participants was 51.8 years, and 826 (22.1%) were diagnosed with NAFLD. Compared to the non-NAFLD group, 57.0% of the metabolites (65/114 investigated metabolites) were significantly increased in the NAFLD population. The metabolite that demonstrated the strongest association was glutamic acid (P = 6.4 x 10-89 and AUC = 0.759), followed by 2-oxoglutaric acid and valine. These metabolites were significantly associated with NAFLD even after adjustment for sex, age, and BMI using logistic regression. Finally, using LASSO (least absolute shrinkage and selection operator), we constructed a diagnostic model including multiple metabolites (n=70), which achieved an AUC of 0.870. **Discussion and Conclusions:** We identified metabolites associated with NAFLD which were observed to be independent of obesity. A high-performance diagnostic model for NAFLD through integration of metabolomics analysis and machine learning was developed which may have potential for clinical application in the future.
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BIOMEDICAL

P-179

Curcumin alleviates palmitate-induced inflammation and glycerolipid accumulation in C2C12 skeletal muscle cells

PRESENTING AUTHOR: Dean Ashley, University of Cambridge, United Kingdom

CO-AUTHORS: McNally BD, Murfitt S, Middleton A, Sassano G, Russell P, Aleksic M, Griffin J

Increased plasma concentrations of palmitate in obese individuals is suggested to enhance inflammation, lipotoxicity and insulin resistance in peripheral tissues, including skeletal muscle. Upon treatment with palmitate expression of the inflammatory cytokines IL6 and Tnfa were increased as measured by rtPCR, while co-treatment with curcumin ameliorated these increases. Published studies have shown that curcumin can increase AMPK activity in skeletal muscle of mice fed a high-fat diet, ameliorating inflammation, mitochondrial dysfunction and insulin resistance. However, the mechanism underpinning these changes has not been fully elucidated. As AMPK is an important regulator of lipid metabolism, we applied mass spectrometry to assess curcumin-induced alterations in the lipid profile of C2C12 myotubes exposed to 750 µM palmitate. Lipids were extracted using a modified Bligh and Dyer method, and lipid changes were assessed using open profiling C18-liquid chromatography-mass spectrometry. Treatment with palmitate raised concentrations of triacylglycerides (TAGs), diacylglycerides (DAGs), ceramides and phosphatidylcholine species. The elevated levels of TAGs and DAGs were reduced with curcumin co-treatment, while concentrations of other lipid species were unaffected. These results suggest the reduced inflammation and insulin resistance exerted by curcumin is driven by increased catabolism of DAGs and TAGs by alleviating the palmitate induced DAG activation of protein kinase C, JNK and IKK pathways.

P-180

Metabolomics as a diagnostic tool in inborn errors of metabolism

PRESENTING AUTHOR: Michel van Weeghel, Amsterdam UMC, Netherlands

CO-AUTHORS: Mia L. Pras-Raves, Martin A. T. Vervaart, Angela C. M. Luyf, Lindsey Welling, Mendy Karssies, Sacha Ferdinandusse, Antoine H. C. van Kampen, Riekelt H. Houtkooper, Ronald J. A. Wanders, Annet M. Bosch, Hans R. Waterham, Frédéric M. Vaz

Metabolomics involves the (semi-)quantitative and qualitative measurement of metabolites in biological matrices and is used as an analytical tool for biomarker discovery in research but have not frequently been applied to diagnostics. Inborn errors of metabolism (IEM) are routinely identified by targeted analysis of intermediary metabolites. Recent developments in high-resolution mass spectrometry, tracer-based metabolomics and ion mobility techniques, opens new avenues to perform (untargeted) metabolomics for the identification of new biomarkers in IEM. In our study, we investigated the metabolome profiles from a set of 120 plasma samples from patients with 10 known inborn errors of peroxisomal metabolism. We used reversed-phase, normal-phase, and HILIC chromatography followed by full-scan orbitrap-MS on the Q Exactive plus in positive and negative ionization mode and annotated over 200 polar metabolites and over a 1200 lipid species. We identified specific biomarkers for the different inborn errors of peroxisomal metabolism and discovered new potential biomarkers. Because this is a semi-targeted approach, identities of some of the newly found biomarkers remain to be characterized. Furthermore, we performed stable isotope tracer experiments on three classical and three variant (no illness at diagnosis by newborn screening, erythrocyte galactose-1-phosphate uridyltransferase activity 4-9%) galactosemia patients and were able to distinguish mild from severe patient (the galactosemia index), which ultimately could lead to personalized treatment of these patients. The ultimate goal is to use this type of metabolomics analysis for first line screening of IEMs.

P-181

Metabolic Signature Associated with Insulin Sensitivity Improvement Following a Weight Loss Intervention

PRESENTING AUTHOR: Jarlei Fiamoncini, University of São Paulo - School of Pharmaceutical Sciences, Brazil

CO-AUTHORS: Milena Rundle, E. Louise Thomas, Barbara Gelhaus, Ronny Scheundel, Alexander Haag, Christian Hoffmann, Guus Roeselers, Denise Sonntag, Jimmy D Bell, Gary Frost, Hannelore Daniel

Metagenomics and metabolomics analysis are allowing the exploration of the connection between intestinal microbiota and host metabolism. Within the NutriTech project, 72 healthy subjects (male and female, ± 60 y.o.) were enrolled in a weight loss intervention (20% dietary energy restriction for 12 weeks), losing in average 6.4% of their body weight. In some volunteers, weight loss was associated with an improvement in insulin sensitivity, while in others only minor changes in insulin sensitivity were observed, despite weight loss. Looking for metabolic signatures that could help understand these outcomes, two PLS-DA models were built: one using the results of body composition and metabolite profiling of plasma samples collected during an OGTT and a mixed meal test; the second using data from intestinal microbiota composition. Marker metabolites of both groups were identified: in particular urea, specific bile acids, amino acids and acylcarnitines, as well as several discriminant bacterial species. Individuals without improvement in insulin sensitivity following the weight loss, displayed higher plasma concentrations of tauroursodeoxycholic acid (a taurine-conjugated bile acid), had higher proportions of Bilophila and other urease-positive bacteria within their microbiome, but lower concentrations of urea and deoxycholic acid. The differences in microbiota composition are in line with the differences in plasma urea and bile acids concentrations. This analysis brings new phenotypic information associated with differential weight loss and allows the generation of hypothesis. Further studies are warranted and validation of the findings in larger cohorts is of course necessary.
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P-182

Batch independent screening of Inborn Errors of Metabolism with untargeted metabolomics

PRESENTING AUTHOR: Michiel Bongaerts, Erasmus Medicale Centre, Netherlands

CO-AUTHORS: Ramon Bonte, Serwet Demirdas, Henk Blom, George Ruijter

It has been demonstrated that untargeted metabolomics can be used for the screening of Inborn Errors of Metabolism (IEM). However, most studies have used control samples from the same experimental run for determining Z-scores of metabolites. This limits both the amount and matching (sex, age) of the controls with the patient of interest. We explored the use of controls originating from different experimental runs / batches. In this way a better reference population can be defined for a given patient. 6 batches were independently processed and subsequently merged. In total 195 controls and 33 patients were measured including 20 different IEM's. We explored multiple normalization methods to correct for signal drift and batch effects. Internal standard based normalization methods performed better than statistical based methods. We developed a normalization method, Metchalizer, which uses a mixed effect model and internal standards for normalization. This method showed the best performance on our data compared with other methods. After normalization with Metchalizer 56 out of 69 biomarkers for patients with known IEM's were detected by using a reference population from other batches. This study identified multiple age/ sex related metabolites by using a regression model. The variance in control samples is determined by both biological and technical variance. The biological variance for a given metabolite might strongly depend on age and/or sex and therefore matching patient and reference is important when determining Z-scores. We show that using controls from other batches can solve this issue as long as batch effects can be corrected.

P-183

Metabolomic alterations in hypocholesterolemic drug users

PRESENTING AUTHOR: Minako Matsumoto, Keio University, Japan

CO-AUTHORS: Sei Harada, Miho Iida, Suzuka Kato, Ayako Kurihara, Ayano Takeuchi, Kazuyo Kuwabara, Daisuke Sugiyama, Tomonori Okamura, Toru Takebayashi

BACKGROUND: Although hypocholesterolemic drugs are the first-line therapy for cardiovascular disease prevention, the most highly used stains are known for their various adverse effects such as muscle inflammation, liver damage, and hyperglycemia. This study sought to determine variations of plasma metabolite concentrations related to hypocholesterolemic drugs intake using capillary electrophoresis-mass spectrometry.
METHODS: All data and samples were collected from participants of the Tsuruoka Metabolomics Cohort Study which was conducted in Japan. Information on medical history, lifestyle, and medications was collected through a standardized, self-administered questionnaire. After excluding those with a history of cancer, stroke, and myocardial infarction, we selected participants on hypocholesterolemic drugs alone as drug users (n=385) and those with LDL-cholesterol 140 mg/dl or above but without any drug as controls (n=1143). Associations between the drug intake and plasma metabolite concentrations were examined by linear regression. Then, analysis of co-variance was conducted to examine the association between levels of fasting plasma glucose (FPG) and hypocholesterolemic drug-related metabolites. RESULTS: Out of 115 polar metabolites analyzed, five were identified as hypocholesterolemic drug-related metabolites. These associations were independent of sex, age, systolic blood pressure, FPG, smoking, alcohol intake, physical activity, dietary energy, and liver function. Three out of five metabolites (betaine, threonine, and cystine) were included in the trans-sulfuration pathway. Cystine was also found to be associated with elevated levels of FPG, which may reflect the effect of statin intake on hyperglycemia. CONCLUSION: Alterations of plasma metabolite levels in transsulfuration pathway were observed with hypocholesterolemic drug intake.

P-184

Metabolomics study on the effect of combination treatment between pioglitazone and piceatannol on Non-Alcoholic Fatty Liver Disease in Rats

PRESENTING AUTHOR: Shun Wan Chan, Technological and Higher Education Institute of Hong Kong, Hong Kong

CO-AUTHORS: Shun-Wan Chan, Chi-On Chan, Tung-Ting Sham, Huan Zhang, Yam-Fung Ng, Daniel Kam-Wah Mok

Non-alcoholic fatty liver disease (NAFLD) is one of the most common liver disorders worldwide. NAFLD prevalence increases recently and becomes an important public health concern. Given the importance of insulin resistance in the pathogenesis of NAFLD, insulin sensitizers were broadly investigated in treating NAFLD. However, the long-term usage related side effects of insulin sensitizers have limited its utility. Combination drug therapy utilizes more than one medication but each agent is given at a dose much lower than the normal therapeutic dose (i.e., minimal side effects are anticipated) resulting in synergistic therapeutic outcomes. In this study, we used metabolomics approaches to investigate a novel drug therapy using pioglitazone (a commonly used insulin sensitizer) in combination with piceatannol, a natural analog of resveratrol, to treat NAFLD in rat fed with high fat diet (HFD). Our in vivo study clearly demonstrated that Sprague-Dawley rats fed with HFD for 4 weeks could establish NAFLD. Treatment with pioglitazone in combination with piceatannol would could significantly reduce the adverse effects induced by HFD. It could lower the liver's lipid content and the total cholesterol level in serum. It was found that the metabolomics profile of combination drug therapy group was significantly different from that of the HFD group but it was getting closer to that of the control group. Additionally, more than 20 metabolites were found as potential biomarkers indicating that this combination drug therapy could effectively modulate the lipid metabolism and result in improvements in NAFLD.
**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

**POSTER SESSIONS 3 AND 4 – Wednesday and Thursday –** all even number posters will be on display.

*AWARD WINNERS

BIOMEDICAL

P-185 In vivo metabolic profiling in brown and white adipose tissue using open flow microperfusion (OFM)

PRESENTING AUTHOR: Elmar Zügner, Joanneum Research Forschungsgesellschaft mbH, Austria

CO-AUTHORS: Simon Schwingenschuh, Thomas Altendorfer-Kroath, Sonja Kainz, Yasemin Erdem, Günther Rauter, Petra Kotzbeck, Joanna Hummer, Thomas Birngruber, Christoph Magnes

One of the major challenges for the healthcare system is the increasing prevalence of obesity. Several therapeutic approaches to overcome obesity modify adipose tissue metabolism to induce sustained weight loss. However, how treatment effects are mediated on multiple metabolic pathways remain elusive. We present a novel method to investigate tissue-specific metabolism in brown and white adipose tissue (BAT, WAT) in rats. We performed open flow microperfusion (OFM) allowing collection of interstitial fluid of BAT and WAT, and subsequent metabolic profiling of the collected samples. Metabolic profiling was performed in OFM samples with volumes as small as 15 µl each. Analyzation was done using HILIC-HRMS (Vanquish UHPLC, QExactive mass spectrometer Thermo Fischer Scientific) in positive and negative electro spray ionisation mode with a gradient elution and a run time of 37 minutes. Metabolic profiling identified 141 metabolites including lipids, fatty acids or amino acids that were classified into metabolites usable for multivariate (MVA) or for univariate (UVA) data analysis. 94 MVA metabolites showed a median standard deviation of peak intensity in pooled quality control samples of 7.1%, low ppm and no retention-time deviation, making them usable for principal component analysis (PCA) and analysis of variance (ANOVA). UVA metabolites were only usable for ANOVA. PCA showed a distinct signature for 2 metabolite classes in WAT and BAT samples. MVA and some UVA metabolites from these classes also showed statistically significant differences in the ANOVA. These results serve as a proof-of-concept to directly investigate adipose tissue metabolism in-vivo.

P-186 Liquid chromatography-mass spectrometry-based approach to evaluate lipidomic changes of plasma in Yijin-Tang-treated obese mice

PRESENTING AUTHOR: Jueun Lee, Korea Basic Science Institute, South Korea

CO-AUTHORS: So Min Lee, Eunjung Kang, Hye-Lin Kim, Jeeyoun Jung, Geum-Sook Hwang

Obesity is one of the most widespread health problems and is characterized by an aberrant lipid metabolism. The herbal remedy Yijin (Erchen)-tang (YJT) is widely used to treat obesity-related disorders. However, the change of lipid metabolism by YJT still remains unknown. In this study, liquid chromatography (LC)-mass spectrometry (MS)-based approach was applied to investigate specific changes in lipid metabolism associated with the therapeutic effects of YJT-treated obese mice. C57BL/6N mice were fed a high-fat and high-cholesterol (HFHC, 40% fat and 1% cholesterol) diet for 8 weeks and treated them with YJT for an additional 6 weeks. We then performed untargeted lipidomic analysis of plasma using UPLC-QTOF MS coupled with multivariate statistical analysis. Partial least squares-discriminant analysis score plots showed that YJT altered the lipid metabolic pattern of HFHC mice. In particular, ceramides and triglycerides with saturated fatty acids and monounsaturated fatty acids were significantly changed by YJT, which were significantly associated with insulin resistance, the AGE-RAGE signaling pathway in diabetic complications and adipocytokine signaling pathway in pathway enrichment analysis. In addition, the changes of total WBC, CRP and blood glucose, and correlation analysis between lipids and those clinical markers showed the effect of YJT treatment on ameliorating inflammation and improving insulin resistance. These data suggest that YJT ameliorates obesity-induced systemic inflammation and insulin resistance by regulating lipid metabolism, and demonstrated that lipidomic profiling is a useful method to investigate the therapeutic effects of herbal decoctions in traditional Korean and Chinese medicine.

P-187 Anabolic androgenic steroids induce weight loss in male Wistar rats in relation to free fatty acids in plasma

PRESENTING AUTHOR: David Balgoma, Uppsala University, Sweden

CO-AUTHORS: Sofia Zelleroth, Alfhild Grönbladh, Mathias Hallberg, Curt Pettersson, Mikael Hedeland

The abuse of anabolic androgenic steroids (AASs) in supratherapeutic doses is a source of public concern. Among other effects, AASs affect lipogenesis and lipolysis in different tissues. Lipidomics has the potential of unraveling the mechanism of the effects of AASs in health and disease. Consequently, we treated Wistar rats every third day with vehicle, nandrolone decanoate, and testosterone undecanoate (supratherapeutic doses). The weight of the animals was measured every third day and, on day 18, the animals were euthanized, blood was collected, and plasma isolated. After extraction, the plasma lipidome was analyzed on an Acquity UPLC-Synapt G2S Q-ToF equipped with a BEH C18 column. Free fatty acids showed a specific regulation in relation to the treatment with AASs. They increased in the treatment with nandrolone decanoate, but they did not change with testosterone undecanoate. Weight decreased with the treatment with nandrolone decanoate but weight did not change with testosterone undecanoate. As free fatty acids in plasma are known to reflect the levels of triacylglyceride lipolysis in the adipose tissue, weight regulation is associated with the levels of free fatty acids in plasma. This effect might play a role in the effect of androgens in health and disease.
**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

**POSTER SESSIONS 3 AND 4 – Wednesday and Thursday –** all even number posters will be on display.

*AWARD WINNERS

BIOMEDICAL

P-188

Development of a New Diagnostic Tool for the Early Detection of Type 2 Diabetes (DeTecT2D)

**PRESENTING AUTHOR:** Rui Wang-Sattler, Helmholtz Zentrum München, German Research Center for Environmental Health, Germany

**CO-AUTHORS:** Stefan Brandmaier, Isobel D. Stewart, Andreas Lechner, Norbert Stefan, John Chambers, Peter Bergsten, Therese Koal, Christian Gieger, Jerzy Adamski, Annette Peters

It is alarming that 1 in 2 people (212 million) with diabetes were undiagnosed according to the International Diabetes Federation. One reason for this problem is that the only sensitive assay for the early detection of Type 2 Diabetes (T2D) is an oral glucose tolerance test (OGTT), which is time consuming and costly. The goal of the H2020 EIT Health funded DeTecT2D consortium therefore is to develop a new metabolomics-based assay for the sensitive detection of pre-diabetes and T2D from a single fasting blood sample. A set of 3 metabolite biomarkers for pre-diabetes and T2D was previously identified by members of the DeTecT2D consortium. Based on clinical studies, the consortium extended this set to 6 metabolites, which were further evaluated in two population-based human cohorts (n=9870). In KORA and Fenland studies, the set of biomarkers showed comparable AUC values (83% and 94%) to detect individuals with pre-diabetes and newly diagnosed T2D, respectively. We further evaluated whether the set of 6 metabolites was capable of replacing the standard OGTT. Based on individuals with normal fasting glucose, we obtained AUC values of 74% and 77% for pre-diabetic and diabetic individuals in the KORA S4 study, respectively. Although our extended set of biomarkers does not outperform the OGTT, it is highly predictive for newly diagnosed T2D. Thus, our set of biomarkers holds great promise for use as a routine screening tool for individuals with pre-diabetes and undiagnosed T2D.

P-189

Mimicking protein restriction by inhibiting SLC6A19 (B0AT1) – a potential target to treat metabolic disorders

**PRESENTING AUTHOR:** Kiran Javed, Australian National University, Australia

**CO-AUTHORS:** Stefan Broer

Recent studies have established that dietary protein restriction has beneficial impacts on the metabolic health and it promotes improved glucose homeostasis by increasing the induction of FGF21 from liver. B0AT1 (SLC6A19) is the major neutral amino acid transporter in the lumen of the intestine that carries out the bulk of amino acid absorption from the diet. It also reabsorbs neutral amino acids in proximal tubule of the kidney. Mice lacking B0AT1 show signs of protein restriction such as elevated levels of FGF21 and reduced mTORC1 activity. Moreover they have improved glucose homeostasis and are protected from diet induced obesity, making it a potential target to treat metabolic diseases such as phenylketonuria and type 2 diabetes. We determined the postprandial levels of amino acids and other metabolites under low, standard and high protein diet of SLC6A19ko, wt and hz mice. All essential amino acids showed a significant positive correlation with the protein content of the diet in WT mice. The neutral AAs that requires SLC6A19 for its absorption were reduced in the blood of SLC6A19ko mice whereas other amino acids showed no significant difference as compared to WT mice. The reduced levels of neutral AA were more prominent under conditions of high protein diet. We also found a few bacterial metabolites of AA fermentation that were correlating with the protein content of the diet in the blood of SLC6A19ko mice. This study highlights the potential of this transporter as a target to induce protein restriction for the treatment of metabolic disorders.

P-190

A Reliable Mass Spectrometry Methodology for Podocyturia Evaluation in Genetic and Non-Genetic Kidney Diseases

**PRESENTING AUTHOR:** Christiane Auray-Blais, Université de Sherbrooke, Canada

**CO-AUTHORS:** Tristan Martineau, Michel Boutin, Anne-Marie-Côté, Bruno Maranda, Daniel Bichet

Increased podocyte levels in urine (podocyturia) is an early sign of kidney abnormalities in patients. Currently, kidney damages are evaluated by using proteinuria measurements and the estimated glomerular filtration rate (eGFR) which, in some cases, might not always be efficient for early detection of patients. Moreover, most analytical techniques for the evaluation of podocyturia are tedious, time-consuming, and may lead to marked results variability. The primary objective of this research project was to develop and validate a multiplex, quantitative ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) approach to evaluate two proteins related to podocyturia, podocalyxin and podocin in urine samples. The secondary aim targeted the analysis of these biomarkers in samples from women with preeclampsia, Fabry disease patients, and related gender-matched controls. One mL of a random urine sample was centrifuged, followed by the addition of cleavable peptide standards to the supernatant. Trypsin digestion was performed (2h at 37oC) and the peptides were purified by solid-phase extraction and evaporated. A 10-min UPLC-MS/MS was developed and validated. Normal values were established. Our results show that women with preeclampsia had abnormal urine levels of both proteins with a higher sensitivity for podocalyxin. Fabry male patients presented a slight elevation of podocin whereas untreated Fabry females had increased concentrations of both podocalyxin and podocin. Correlations were established with these biomarkers and some clinical parameters. This reliable methodology might be an efficient tool for early diagnosis of women with preeclampsia and Fabry disease patients. It is also applicable to other genetic and non-genetic kidney diseases.
**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

**POSTER SESSIONS 3 AND 4 – Wednesday and Thursday –** all even number posters will be on display.

*AWARD WINNERS

BIOMEDICAL

P-191

Fecal metabolome and microbiota composition in obese Göttingen minipigs as influenced by source of carbohydrates

PRESENTING AUTHOR: Mihai Victor Curtasu, Aarhus University, Denmark

CO-AUTHORS: Valeria Tafintseva, Zachary Bendiks, Maria L. Marco, Achim Kohler, Yetong Xu, Helle Nygaard Lærke, Knud Erik Bach Knudsen, Mette Skou Hedemann.

The objective of this study was to explore the effects of ad libitum feeding of two high-fat diets supplemented with either high amylose maize starch (HIMA) or fructose on the fecal metabolome and microbiota composition. These effects were examined in obese Göttingen Minipigs as a model for human metabolic disorders. Thirty Göttingen Minipigs were fed with two experimental diets for five months, and fecal material was sampled three times during the dietary intervention. Non-targeted liquid chromatography-mass spectrometry was used to explore metabolite differences, short-chain fatty acids (SCFAs) were quantified by gas chromatography, and bacterial diversity was assessed by 16S rRNA gene sequencing. Sequencing data were analyzed using QIIME 2, and together with metabolomics and SCFAs data a sparse multi-block partial least squares regression model was calculated. Higher proportions of Bacteroidetes were found in the fecal contents of animals fed HIMA compared to that fed fructose together with higher quantities of total organic acids and total SCFAs. Furthermore, concentrations of acetate and acetate+propionate+butyrate were positively associated with the Ruminococcus genus and increased levels of several dicarboxylic acids (2,4-dimethyladipic acid, 3,3-methylglutaric acid) and pantothenic acid. Minipigs fed fructose contained higher butyrate levels, which were closely associated with enrichments in Roseburia, Clostridiaceae, Turicibacter, and Ruminococcus (Lachnospiraceae family). Bacteria from the Coprococcus, Blautia or Coprococcus genera were associated with a higher intensity of enterolactone (m/z 297.1135; [M+Cl] m/z 333.0902). Overall, we observed that different carbohydrate sources induced a contrasting stimulation of the microbial taxa which further reflected in a divergent fecal metabolome profile.

P-192

Analysis of bile acid profiles in patients with nonalcoholic fatty liver disease (NAFLD) depending on NAFLD severity

PRESENTING AUTHOR: Youngae Jung, KBSI, South Korea

CO-AUTHORS: Won Kim, Geum-Sook Hwang

Nonalcoholic fatty liver disease (NAFLD) is one of common causes of chronic liver disease that involve hepatic accumulation of triglycerides. NAFLD is progressed from a nonalcoholic fatty liver (NAFL) to nonalcoholic steatohepatitis (NASH). Bile acids has been considered to be related with pathogenesis of NAFLD. In this study, we analyzed the profile of bile acids in NAFLD patients using LC-MS system. Sera acquired from 234 Asian subjects with biopsy-proven NAFLD and metabolically healthy controls were used for analysis of bile acids. Bile acid profiles included total 13 bile acids such as cholic acid (CA), chenodeoxycholic acid (CDCA), deoxycholic acid (DCA), hyodeoxycholic acid (HDCA), ursodeoxycholic acid (UDCA), glycocholic acid (GCA), glycochenodeoxycholic acid (GCDCA), glycodeoxycholic acid (GDCA), taurodeoxycholic acid (TCA), taurochenodeoxycholic acid (TCDCA), taurodeoxycholic acid (TDCA), taurohyodeoxycholic acid (THDCA), taurosodeoxycholic acid (TUDCA). Most of bile acids showed the significant increases in NASH compared to NAFL, whiles didn't show the significant changes in NAFL compared to No-NAFLD. And primary bile acids were mainly increased in NASH. Our results showed that the bile acid profiles in NAFLD patients were changed depending on NAFLD severity and also presented to differently be changed according to types of bile acids.

P-193*

Plasma metabolomics identifies potentials biomarkers in different stages of Non-alcoholic fatty liver disease

PRESENTING AUTHOR: XIANGPING LIN, Laboratoire CSPBAT - Paris 13 University - UFR SMBH, France

CO-AUTHORS: Xinyu LIU, Mohamed TRIBA, Zhicheng LIU, Laurence LE MOYEC, Marianne ZIOL, Nada HELMY, Corinne VONS, Nadia BOUCHEMAL, Guowang XU, Carina Prip-buus, Philippe SAVARIN

Non-alcoholic fatty liver disease (NAFLD), characterized by abnormal accumulation of triglycerides (TGs) in the liver, which is becoming the most common chronic liver diseases in the global adult population. A previous study found that plasma metabolome was a better predictor for steatosis (80%) than noninvasive basal clinical data (58%). In this study, we investigate whether High-resolution mass spectrometry-based Plasma metabolomics could identify potentials biomarkers in different stages of the disease, and explore the further molecular mechanism involved in the progression of NAFLD. Plasma were collected from obese subjects with Normal liver (n=19), Steatosis (n=39) and NASH (n=24). To cover polar and non-polar metabolites, samples analysis were performed using two separated ultra-performance liquid chromatography High-resolution mass spectrometry-based Metabolomic and lipidomic platform, respectively. Compared with Normal liver obese, Steatosis or NASH subjects had abnormal liver function, high plasma TGs concentrations and were insulino-resistance. Binary Logistic Regression analysis found each a panel of potentials biomarkers for Steatosis, NASH and Steatosis from NASH, the regression models were evaluated by internal validation with Area under the ROC Curve around 0.7 (mean for validation). The small sample size may be a limit of this study, still, our results suggested that High-resolution mass spectrometry-based Plasma metabolomics may be a promising way to identify plasma biomarkers of NAFLD, the external validation is in progress, after validated in other independent studies, these biomarkers might help to diagnosis Steatosis from NASH, monitor development of NAFLD and shed light further on molecular mechanism involved in disease progression.
**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

**POSTER SESSIONS 3 AND 4 – Wednesday and Thursday –** all even number posters will be on display.

*AWARD WINNERS

BIOMEDICAL

P-194

Metabolomics approaches to metabolic changes after the intermittent fasting diet

PRESENTING AUTHOR: Jeongae Lee, Reaserch Scientist, South Korea

CO-AUTHORS: Yoon Hwan Kim, Kyoung Heon Kim, Bong Chul Chung, Young-Ran Yoon, Ji-Won Lee

Human serum with intermittent fasting was analyzed by metabolomics and multivariate analysis, and increased of 2-hydroxyphenylacetate and decreased of phenylpyruvate were observed in phenylalanine metabolism.

P-195

The impact of ulcers on the metabolomic profile of serum of diabetic patients

PRESENTING AUTHOR: Arsenty Melnikov, International Tomography Center SB RAS / Novosibirsk State University, Russian Federation

CO-AUTHORS: Yuriy Tsentalovich, Vadim Yanshole

It is believed that the alternations in lipid and protein catabolism in patients with diabetes mellitus (DM) leads to various complications, such as diabetic ulcer. Despite the fact that diabetic ulcer is one of the most common complication of DM, it is still poorly studied. Current study is devoted to the comparative metabolomic analysis of serum obtained from patients with type 2 DM with and without ulcers. To obtain the serum metabolomic profiles, two high-performance liquid chromatography methods (HILIC and RPLC) in conjunction with high-resolution ESI-q-TOF mass-spectrometric detection were used. This approach made possible to register more than 1700 features (in both HILIC and RPLC data), yielding more than 250 metabolites and lipids. For the majority of metabolites, their levels in the serum of diabetic patients with and without ulcers are similar, with few exceptions including 1,5-anhydroglucitol (fold change = 3.2, AUC = 0.96 ± 0.02), bilirubin (fold change = -3.1, AUC = 0.89 ± 0.03) and a group of phospholipids with the mean fold change equals to -1.48 ± 0.02. For all listed differences the p-value is lower than 0.05. This study shows that the development of diabetic ulcer is accompanied by the decrease in the concentrations of phospholipids and some lipid-related metabolites. Acknowledgements: Supported by the RFBR (18-33-20097) and by RFBR and NSO (18-415-543006).

P-196

Fast detection of drugs and metabolites in urine by Flow Injection Analysis coupled to Magnetic Resonance Mass Spectrometry

PRESENTING AUTHOR: Christopher Thompson, Bruker Daltonics Inc., United States

CO-AUTHORS: Matthias Witt, Aiko Barsch, Markus Godejohann

A fast method for detection of drugs and their metabolites in urine using flow injection analysis (FIA) and magnetic resonance mass spectrometry (MRMS) is presented. Roughly 250 samples can be measured in 24h using this technique. Pooled urine samples were purified by SPE using Merck LiChrolutEN SPE cartridges. Samples were extracted with methanol from SPE cartridges and diluted in Methanol for FIA. Each sample was analyzed in 5 minutes by FIA-MRMS in ESI in positive and negative ion mode. Spectra were acquired in a mass range m/z 107 – 3000 in quadrupolar detection mode with a resolving power of 1,350,000 at m/z 200. For the final mass spectrum 28 single scans were averaged. Analysis of data was performed with MetaboScape 4.0. The data of the ESI(+) and ESI(-) were combined for feature analysis. More than 2100 features were found for the pooled urine samples. More than 90% of the detected features could be assigned with a molecular formula. 300 drug candidates were annotated in the urine samples by matching a target list of expected compounds derived from HMDB (http://www.hmdb.ca/) with a mass error tolerance of only 0.5 ppm. The detected drugs were compared with the medication of the patients. Several drugs were found only in specific pooled urine samples. By comparing the relative abundances of features of all samples, possible metabolites of drugs could be identified.

P-197

Improved identification of human CSF metabolites using workflow-based approaches allowing integration of multiple search engines

PRESENTING AUTHOR: Kim Kultima, Uppsala University, Sweden

CO-AUTHORS: Payam Emami Khoonsari, Christoph Ruttkies, Stephanie Herman, Kristian Peters, Ralf Weber, Thomas Lawson, Joachim Burman, Ola Spjuth, Steffen Neumann and Kim Kultima and members of the ELIXIR Implementation Study on Metabolite Identification

Untargeted metabolomics as a high-throughput molecular phenotyping technique is growing across all domains in the life sciences. The data processing and analysis is often performed using different tools with little standardization and automation for interoperable and reproducible research. We have previously developed a comprehensive and interoperable workflow that integrates all necessary components for doing mass spectrometry-based identification and quantification of metabolites as well as the downstream analysis1–3. To improve metabolite identification we have now developed a workflow approach that allows integration of MS1 and MS2 information, resulting in improvement in statistical scoring of the identified compounds as well as a substantial decrease in computation time. The workflow includes multiple search engines such as CSI:FingerID and MetFrag. The workflow presented can directly connect to the MetaboLights data repository, and performs identification as well as quantification, alignment, and matching using a large collection of available tools. Together, we achieved a complete integration of several major metabolomics software suites resulting in a turn-key workflow for mass spectrometry-based metabolomics. This approach vastly improved identification of metabolites expressed in human cerebrospinal fluid (CSF). The workflow is available on Galaxy, Nextflow and Pachyderm on PhenoMeNal. This study is part of the European collaborative ELIXIR Implementation Study on Metabolite Identification.
**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

**POSTER SESSIONS 3 AND 4 – Wednesday and Thursday –** all even number posters will be on display.

*AWARD WINNERS

BIOMEDICAL

P-198

DecoMetDIA: Deconvolution of Multiplexed MS/MS Spectra for Metabolite Identification in SWATH-MS based
Untargeted Metabolomics

PRESENTING AUTHOR: Yandong Yin, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, China

CO-AUTHORS: Zheng-jiang Zhu

With recent advances in mass spectrometry, there is an increased interest in data-independent acquisition (DIA) techniques for metabolomics. With DIA technique, all metabolite ions are sequentially selected and isolated using a wide window to generate multiplexed MS/MS spectra. DIA strategy enables a continuous and unbiased acquisition of all metabolites and increases the data dimensionality, but presents a challenge to data analysis due to the loss of the direct link between precursor ion and fragment ions. Currently, only few DIA data processing methods are developed for metabolomics application, including MS-DIAL and MetDIA. Nevertheless, there is still a lack of software tools to support the SWATH-MS based untargeted metabolomics. Here we presented a new software tool, namely, DecoMetDIA, to support metabolite identification in SWATH-MS based untargeted metabolomics with spectra deconvolution with extracted MS2 EICs to satisfy the need of data processing tools for SWATH-MS technique in metabolomics. With the large-scale metabolite annotation of the MS2 spectra deconvoluted by DecoMetDIA using MetDNA and Sirius/ CSI:FingerID, we found that about 70~80% features were annotated, which proved that the MS2 spectra deconvoluted by DecoMetDIA were of high accuracy and ready to be used for large scale metabolite identification.

P-199

Automated microfluidic cell culture of stem cell derived dopaminergic neurons

PRESENTING AUTHOR: Edinson Lucumi Moreno, Leiden University, Netherlands

CO-AUTHORS: Khalid I. W. Kane, Siham Hachi, Moriz Walter, Javier Jarazo, Miguel A. P. Oliveira, Thomas Hankemeier, Paul Vulto, Jens C. Schwamborn, Martin Thoma, Ronan M. T. Fleming

Parkinson's disease is a slowly progressive neurodegenerative disease characterised by dysfunction and death of selectively vulnerable midbrain dopaminergic neurons and the development of human in vitro cellular models of the disease is a major challenge in Parkinson's disease research. We constructed an automated cell culture platform optimised for long-term maintenance and monitoring of different cells in three dimensional microfluidic cell culture devices. The system can be flexibly adapted to various experimental protocols and features time-lapse imaging microscopy for quality control and electrophysiology monitoring to assess cellular activity. Using this system, we continuously monitored the differentiation of Parkinson's disease patient derived human neuroepithelial stem cells into midbrain specific dopaminergic neurons. Calcium imaging confirmed the electrophysiological activity of differentiated neurons and immunostaining confirmed the efficiency of the differentiation protocol. This system is the first example of an automated Organ-on-a-Chip culture and has the potential to enable a versatile array of in vitro experiments for patient-specific disease modelling.

P-200

Comprehensive Analysis of Metabolites Using GC-MS/MS and LC-MS/MS -An Application to the Research of the
Intestinal Environment

PRESENTING AUTHOR: Akihiro Kunisawa, Shimadzu Corporation, Japan

CO-AUTHORS: Akihiro Kunisawa, Takanari Hattori, Shuichi Kawana, Shin-ichi Kawano, Yoshihiro Hayakawa, Junko Iida, Eiichiro Fukusaki, Mitsuharu Matsumoto.

A part of the metabolites produced by intestinal microbiota are absorbed constantly from the intestinal lumen and carried to systemic circulation; and play a direct role in health and disease. Recently, increasing attention has been devoted to the relationship among intestinal microbiota, the metabolites, and health and disease. There are limited reports concerning the function of metabolites produced by intestinal microbiome. Furthermore, these studies are targeting specific metabolites such as short-chain fatty acids but not global metabolites (metabolome). In this study, we analyzed metabolites in mice feces using GC-MS/MS and LC-MS/MS for comprehensive analysis. As results of GC-MS/MS analysis for the extracts of the mice feces, 100 metabolites were detected. Main compounds of 100 metabolites were short-chain fatty acids, organic acids and sugars. Seventeen sugars that are difficult to analyze using LC-MS/MS were detected. As results of ion-pairing LC-MS/MS analysis, 17 metabolites which were mainly amino acids were detected. As results of non-ion-pairing LC-MS/MS analysis, 75 metabolites were detected. Main compounds of 75 metabolites were amino acids, nucleotides, nucleosides and organic acids. These results indicated that comprehensive analysis of metabolites using GC-MS/MS and LC-MS/MS is effective for the research of the intestinal environment. In addition, we also report on the interpretation of the metabolome data using the Shimadzu Multi-omics Data Analysis Pack, which will facilitate the examination of the results.
**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

**POSTER SESSIONS 3 AND 4 – Wednesday and Thursday –** all even number posters will be on display.

*AWARD WINNERS

BIOMEDICAL

P-201

Metabolomic characteristics of the facial skin in young and aged cohorts from a Caucasian female population

PRESENTING AUTHOR: Philipp Ternes, BASF Metabolome Solutions GmbH, Germany

CO-AUTHORS: Allison Guinta, Nicolas Del Bene, Neethu Abraham, Lauren Junker, Wendy Chan, Hunter Cameron, Anita Samuga, Nengyi Zhang, George Kritikos, Vimal Rawat, Girish Srinivas, Sabrina Letoy-Okombi, Valérie André

An in-depth understanding of the changes which are occurring on multiple levels in the human skin during aging is of major interest for the development of novel cosmetic ingredients. Aging of the facial skin is known to be accompanied by profound changes in the skin microbiota and in biophysical skin quality parameters (e.g. Jugé et al. 2018, J. Appl. Microbiol. 125, 907; Zouboulis et al. 2018, Mech. Ageing Dev. 170, 98). To gain a better understanding of the interactions between the skin and its microbiota, we have conducted a simultaneous analysis of the skin microbiome and metabolome in the crow's feet/undereye area in cohorts of ≈ 50 young (age 18–35 years, wrinkle grade 0–1) and ≈ 50 aged (age > 55 years, wrinkle grade 5–6) healthy Caucasian women. This poster presentation will focus of the metabolomic characteristics of these two cohorts.

P-202

Simultaneous quantification of tryptophan pathway metabolites in serum, plasma and faeces using UHPLC-MS/MS method

PRESENTING AUTHOR: Dai Long Vu, Max Delbruck Center for Molecular Medicines, Germany

CO-AUTHORS: Andras Balogh, Hendrik Bartolomaeus, Jennifer Kirwan

Tryptophan and its metabolites play key roles in heart and brain health. The eventual fate of tryptophan involves an interplay between host and gut microbiome. Quantification of the tryptophan pathway metabolites provides important information about the eventual fate of tryptophan in the body which may be important for the study of certain diseases. Although current methods exist for the quantification of a subset of these metabolites, methods for simultaneous quantification of most of these compounds are still missing. We demonstrate a robust, accurate, reproducible method for the quantification of 30 compounds related to the tryptophan pathway using an ultrahigh performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) technique. The analytical method was fully validated for serum, plasma and faeces, following European Medicines Agency guidelines. We report on the analytical linearity, limit of detection, limit of quantification, matrix effect, recovery, accuracy and precision of the method. In addition, stability of standards and samples were also investigated. Except for tryptophan which was degraded after 24 hours at 4 °C, all other tested compounds were stable up to 48 hours, exceeding the run length of a typical batch. Our study also offers a high throughput analytical method as the separation in a reversed phase system can separate the analytes in a 10 minute gradient, fostering the analysis of batches with high number of samples.

P-203

Effect of nutrition on host-gut microbiota metabolic interactions of preterm infants fed with own mother's milk and donated mother milk

PRESENTING AUTHOR: José david Piñeiro, Neonatal Research Group, Spain

CO-AUTHORS: Anna Parra-Llorca, María Gormaz, María Cernada, Guillermo Quintás, María Carmen Collado, Máximo Vento, Julia Kuligowski

Breastfeeding is the gold standard for nutrition of the term and preterm infant. For preterm infants whose mothers are unable to provide of own mother's milk (OMM), pasteurized donor human milk (DHM) is the preferred alternative. Here, the metabolic signature of preterm infants has been studied to unscramble the effect of nutrition on their metabolism. Urine samples of 40 preterm infants fed with OMM (N=20) and DHM (N=20) aged between 2 and 4 weeks of life were collected and analysed employing Liquid Chromatography-High Resolution Mass Spectrometry. A peak table was extracted with XCMS and a total of 2241 m/z features were retained after data pre-processing and filtering. Pathway analysis (MetaboAnalyst) revealed a significant alteration of three pathways related to the biosynthesis of hormones. Significant differences were found between urinary fingerprints of preterm infants receiving OMM and DHM (Partial Least Squares Discriminant Analysis, p-value=0.009). 73 features were annotated corresponding to a total of 25 metabolites. Seven putatively annotated metabolites were related to the activity of the gut microbiome. This is the first study of the urinary metabolome of preterm infants fed with DHM vs. OMM. Observed changes could be potentially attributed to i) diet-dependent changes in the colonization pattern of preterm infants that might affect hormone production, ii) changes in the hormone levels in human milk and/or iii) the effect of pasteurization on DHM hormone levels. Based on the results, future studies for assessing the impact of maternal nutrition as well as treatment and storage on human milk composition are encouraged.
**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

**POSTER SESSIONS 3 AND 4 – Wednesday and Thursday –** all even number posters will be on display.

*AWARD WINNERS

BIOMEDICAL

P-204
**Metabolomic evaluation of probiotic beneficial effect on non-alcoholic fatty liver diseases in mouse model using LC-Orbitrap MS and GC-TOF MS**

PRESENTING AUTHOR: Jeong Seok Yu, Kookmin University, South Korea

CO-AUTHORS: Dae Hee Han, Ki-Tae Suk and Do Yup Lee

Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver diseases caused by numerous factors such as genetics, dysbiosis, and diet. NAFLD is defined as from mild liver steatosis to non-alcoholic steatohepatitis (NASH). NASH may lead to liver cirrhosis and hepatocellular carcinoma. Recent studies have proposed dysbiosis as the critical factor for NAFLD. Dysbiosis causes endothelial barrier dysfunction and leads to leaky gut. Compromised gut barrier allows the translocation of bacteria and bacteria-derived molecules to liver through portal vein. Colonic homeostasis is closely related with intestinal short chain fatty acids (SCFAs) which are the fermentation end-products by gut microbiota. Especially, butyrate is the most important energy source for enterocytes and contributes to colonic homeostasis. In current study, we applied four different combination of mass-spectrometric platforms to comprehensively measure a range of metabolic features and some key known factors. We first evaluated pharabiotic effect on the mice fed with western-diet by oral administration of 6 probiotics. Indeed, we observed the coordinated changes in SCFAs levels from cecal samples. Based on multivariate statistical model, we putatively identified metabolites indicating strong positive correlation with the SCFA alteration. Finally, we monitored the modulation of primary and secondary bile acids that may provide detailed mechanistic in molecular dynamics between gut and liver.

P-205
**Paroxetine treatment in mice increases fecal bile acid levels that correlate with behavior**

PRESENTING AUTHOR: Frederik Dethloff, MPI of Psychiatry, Germany

CO-AUTHORS: *Elijah Emmanuel, Vargas Fernando, Quinn Robert, Herzog David, Mueller Marianne, Dorrestein Pieter, Turck Christoph*

Selective Serotonin Reuptake Inhibitors (SSRIs) are commonly used drugs to treat major depressive disorder. Since SSRIs are drugs that are taken orally it is conceivable that they can affect gut microbiota and their functions and thereby impact other body organs. In this regard it has now become increasingly clear that microbiota communicate with the central nervous system through the gut-brain-axis. Several reports have shown that a distorted gut microbiota composition that is mediated by its secreted molecular constituents can impact the central nervous system and contribute to the development of psychiatric disorders. We treated DBA/2J mice for 2 weeks with the SSRI paroxetine and assessed their behavioral response with the forced swim test (FST) and the light-dark box test (LDB). In order to investigate paroxetine's effects on the gut microbiota we analyzed fecal pellets as a proxy. In addition to 16S rRNA we interrogated by tandem mass spectrometry the metabolite profiles of fecal pellet extracts. Untargeted metabolite profiling of drug- and vehicle-treated mice revealed altered levels of features that belong to the bile acid class and have a strong correlation with behavioral measures. No significant alterations of the taxa occured in response to the two week paroxetine treatment. Microbiota-derived metabolites may constitute biomarkers able to predict paroxetine treatment response.

P-206
**'Functional Microbiomics' – Assessing Nutrition-Microbiome-Host Interaction in Blood and Feces**

PRESENTING AUTHOR: Barbara Wolf, BIOCRATES Life Sciences AG, Austria

CO-AUTHORS: Hai Pham Tuan, Ulf Sommer, Svenja Heischmann, Doreen Kirchberg, Xenia Iwanowa, Radu Talmazan, Martin Buratti, Therese Koal

In recent years, microbiome research has dramatically reshaped our understanding of how microbes impact on a multitude of (patho-)physiological processes in the host. However, causal links are still lacking to a large extent. Metabolomics allows the investigation of microbial metabolic activities, and is thus the ideal technology to assess functional nutrition-microbiota-host crosstalk. Here, we discuss the application of a newly developed targeted assay for the quantification of endogenous and microbiota-derived metabolites. 10 µl human plasma and fecal samples were analyzed by a standardized, quantitative assay in kit format allowing for the analysis of 630 metabolites. 106 small molecules from 13 compound classes are analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS), hexoses and 523 lipids from 12 lipid classes by flow-injection analysis-tandem mass spectrometry (FIA-MS/MS) on an Agilent 1290 Infinity UHPLC – SCIEX QTRAP® 5500. MetIDQ™ software was used for the entire automated workflow. In plasma, more than 455 metabolites were quantified above LOD with high, and more than 120 metabolites in fecal samples. To a large extent, the small molecules and lipids quantified in feces overlap with those in plasma. A higher number of lipids, especially phosphatidylcholines and triglycerides, were quantified in plasma compared to fecal samples. In addition to endogenous metabolites, a multitude of microbiota-derived metabolites were quantified. The capability to quantify microbiota-derived metabolites in blood and fecal samples allows for correlation studies also with data from other omics technologies to investigate functional nutrition-microbiome-host interplay for uncovering causal links to pathophysiological processes, disease development, and response to drug treatment.
**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

**POSTER SESSIONS 3 AND 4 – Wednesday and Thursday –** all even number posters will be on display.

*AWARD WINNERS

BIOMEDICAL

P-207 Developing an optimal fecal extraction protocol to maximize metabolite coverage for metabolomics studies

PRESENTING AUTHOR: Mary Boyce, Edith Cowan University, Australia

CO-AUTHORS: Jessica Pandohee, Claus T Christopherson, David I Broadhurst

**Background:** Metabolomics is the analysis and unbiased relative quantification of all metabolites in a biological sample. In order to study the entire metabolome of a biological system, it is essential that most, if not all, of the metabolites present in the system in question are extracted. Human stool is a complex solid sample making reproducible extraction of metabolites challenging. The aim of this work was to develop extraction methods that maximize the metabolite coverage for untargeted and semi-targeted metabolomics studies. **Methods:** To investigate the effect of solvent polarity on the metabolite extraction coverage, homogenised fecal samples (100 mg) were individually extracted with 200 µL solvent. The supernatant was dried and the residue derivatised using methoxyamine in pyridine followed by MSTFA. The samples were analysed on a ThermoFisher Q-Exactive GC-MS. The metabolites were identified using both our in-house library of 400 metabolites and NIST. **Results:** Of the 16 solvents tested, water (pH 2), methanol and acetone extracted 91, 102 and 91 respectively of the 400 analytes in our in-house library with RSD <30%. The solvents MTBE and chloroform extracted 28 and 30 analytes, respectively. When performing a biphasic extraction using water (pH 2):methanol:acetone (40:30:30) as aqueous layer and MTBE as non-aqueous layer, 99 and 40 metabolites with an RSD < 30% are identified respectively. On the other hand, 54 (non-aqueous) and 130 (aqueous) metabolites are identified in a sequential extraction. **Conclusions:** A sequential extraction involving MTBE followed by a methanol/acetone/water phase provides optimal coverage of feces for untargeted or semi-targeted studies.

P-208 Can probiotics reduce urogenital malodour in women? – A targeted metabolomics approach

PRESENTING AUTHOR: Scarlett Puebla-Barragan, The University of Western Ontario, Canada

CO-AUTHORS: Justin Renaud, Mark Sumarah, Gregor Reid

Millions of women around the world seek medical attention due to urogenital malodour, which negatively impacts their quality of life. The main causes are urinary tract infections (UTIs) and bacterial vaginosis (BV), which can occur as a result of vaginal dysbiosis, defined as a disruption of the vaginal microbiota. Unfortunately, the standard treatment for these conditions is a course of antibiotics, which is not always effective and takes several days to improve the symptoms; therefore, a more efficient way to reduce malodour is required. Previously, it was reported that the biogenic amines: putrescine, cadaverine, trimethylamine, and tyramine, are responsible of malodour in BV. However, it is not clear if these are also present in higher concentrations during a UTI. Using HILIC-MS analysis and a targeted metabolomics approach, we identified that uropathogenic strains of E. coli (which cause around 80% of UTIs) can significantly produce putrescine, cadaverine, and trimethylamine when using female urine as substrate in vitro. These results were confirmed analyzing 30 clinical samples. This demonstrates that there is an overlap in the causes of malodour during a UTI and BV. With the same analytical approach, we demonstrated that certain lactobacilli strains can degrade these malodorous compounds in conditions that simulate the vaginal environment in vitro. Some lactobacilli probiotics are already known to help in restoring the vaginal microbiota. In addition to this benefit, our evidence suggests that some lactobacilli strains could also target urogenital malodour – making them a more efficient alternative to the current treatment.

P-210 Analytical methods to measure kinetics of fermentation of non-digestible carbohydrates inside the human gut using novel gastrointestinal sampling capsules

PRESENTING AUTHOR: Melany Rios-Morales, University Medical Center Groningen, Netherlands

CO-AUTHORS: Mara van Trijp, Theo Boer, Rebecca Heiner-Fokkema, Albert Gerding, Fjodor Sluijs, Guido Hooiveld, Barbara Bakker, Dirk-Jan Reijingoud

Consumption of non-digestible carbohydrates (NDC) has been linked to many health benefits. However, detailed knowledge of the exact fate and impact of NDC in the intestinal tract is lacking. Gut microbiota use NDC as substrates to produce short chain fatty acids (SCFAs) and other metabolites. In mice after feeding NDC, only the uptake flux of SCFAs from the intestine correlated with improvements of the metabolic syndrome. In humans, access to this inner world is now possible by an orally swallowable gastrointestinal capsule, that can deliver or sample at an exact location in the gastrointestinal tract. Specific challenges of using the gastrointestinal capsules are: 1) The small volumes of sample obtained. To overcome this, we optimized the analytical protocols to measure NDC breakdown, gut microbiota composition, and bacterial fermentation products (notably SCFAs) in the same small and complex samples as we expect to obtain from the capsule; and 2) the capsule with sample remains in the body until its excretion. Hence, we developed a stabilizing solution to be preloaded in the capsule that can efficiently stop fermentation for up to 2 days. This will allow us to collect a representative sample of the human colon. Furthermore, we verified that the quench solution was not a source of variation in the analytical analyses mentioned above. This analytical toolbox to use gastrointestinal sampling capsule systems will allow us to study fibre intake, SCFAs uptake fluxes and improvements of the metabolic syndrome markers in human subjects.
**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

**POSTER SESSIONS 3 AND 4 – Wednesday and Thursday –** all even number posters will be on display.

*AWARD WINNERS

BIOMEDICAL

P-211 **Metabolomics of Intestinal Bacteria for Novel Kidney Stone Therapeutics**

**PRESENTING AUTHOR:** Casey Chamberlain, University of Florida, United States

**CO-AUTHORS:** Marguerite Hatch, Timothy J. Garrett

Kidney stone disease afflicts 1 in 11 people in the U.S. and burdens the economy with an annual treatment cost of over $10 billion. Nearly 80% of kidney stones are composed of calcium oxalate (CaOx) and form when humans are exposed to oxalate through the diet and endogenous glyoxylate metabolism in the liver. Recently, significant clinical interest has turned to the intestinal microbiome, specifically to a class of "oxalotrophic" bacteria which degrade dietary oxalate. These bacteria reduce the oxalate absorbed by the body, which directly influences CaOx stone formation risk. The commensal Oxalobacter formigenes (Oxf) utilizes oxalate as its sole energy source and has the exclusive ability among oxalotrophs to draw endogenous circulating oxalate from the bloodstream into the intestine to increase oxalate availability, allowing it to feed on both metabolic and dietary oxalate. Hence, its use as a probiotic therapy for kidney stone disease is of key interest. Compelling evidence demonstrates that Oxf produces an unidentified secreted bioactive compound to regulate oxalate transport proteins in the intestinal wall to bring oxalate across the epithelium. This compound, if identified, could potentially serve as a future therapy for kidney stones and other oxalate-derived conditions. This study seeks to identify this secreted factor and other oxalate disease-relevant metabolic products through a UHPLC-HRMS-based characterization and comparison of the metabolome and lipidome of several strains of Oxf. We also present a related analytical investigation of Lactobacillus acidophilus and Lactobacillus gasseri, two additional intestinal oxalotrophs that also show promise as future oxalate-degrading probiotic therapies.

P-212 **Comparing metabolomic profiles of plasma, urine, feces and food in monkey study**

**PRESENTING AUTHOR:** Suzumi M Tokuoka, The University of Tokyo, Japan

**CO-AUTHORS:** Megumi Ishibashi, Fumie Hamano, Yoshihiro Kita, Yoshiya Oda

Human metabolome profiles are affected by environment factors, especially lifestyle such as eating habits and exercise. We examined systematically whether metabolomic profiles would be similar in the model animals under the same environmental conditions even though genetic backgrounds are different. We chose monkey as a model animal that seems to be closer to humans than rodents. The mass spectrometric measurements of hydrophilic and hydrophobic metabolites were performed using both non-targeted and targeted approaches on 20 cynomolgus monkeys, which were composed of four groups in each five animals according to birthplace, age and health condition. The metabolomic profiles of serum, urine and feces were differed widely among individuals rather than the groups despite being kept for a long period under the same environment and food. No marked difference in profiles was found between chronically sick group (with diarrhea and vomiting) and others, but some metabolite levels may be changed in sick monkeys. In spite of the food does not contain arachidonic acid and contains very few docosahexaenoic acid, significant amount of them were detected in plasma. Those levels were different among monkeys and fatty acid profiles in plasma may vary by a country of origin. Phospholipid profile was characteristic among plasma, urine and feces. Plasma contains more phosphatidylcholine with unsaturated longer fatty chain, but urine contains shorter fatty chain and lysophospholipid rather than diacyl phospholipids. Feces have more sphingomyelin and phosphatidylethanolamine. We are now looking for relationships of metabolic profiles among plasma, urine, feces and food.

P-213 **Immunomodulatory Function of Human Gut Microbiota during Early Life Development**

**PRESENTING AUTHOR:** Zdeněk Spacil, Masaryk University, Czech Republic

**CO-AUTHORS:** Veronika Vidová, Eliška Stuchlíková, Anne-Christine Aust, Jana Klanová, Zdeněk Spáčil

The exposome paradigm represents a complement to heritable factors in the understanding of health and disease. While the human gut microbiota is considered a pivotal external factor affecting the metabolism and overall health of the host through immunomodulation. The early life stimulation shaping the innate immune system may be the initial mechanism behind the development of some diseases. An appropriate multi-omics toolbox is required to explore immune-metabolic viewpoint of microbial immunomodulation and disease. We applied tandem mass spectrometry assays using a triple quadrupole mass analyzer (selected reaction monitoring – SRM) for metabolic profiling of specific catabolites of tryptophan and kynurenine pathways in biofluids and cell culture material. SRM-proteomics assays were used to determine circulating and local inflammatory markers. High resolution/accurate mass (HR/AM) exploratory mass spectrometry (Orbitrap Fusion, Thermo Scientific) was utilized for metabolic screening. Results demonstrated microbial metabolites clinically relevant markers of inflammation, dysbiosis or a disease condition, documenting a tight connection between microbial colonization and the host immune system. We have mapped the distribution of microbiota-associated metabolites within biofluids in adults, pregnant women, and neonates and linked them to levels of acute phase proteins. The study is pioneering the functional characterization of microbiota, mediated via stimulation of the immune system and essential to understanding its role in human health. This is consistent with age-dependent changes, underlying emerging concepts of exposome and inflammingaging. Funding: GACR project (17-24592Y), GAMU project (MUNI/G/1131/2017), CETOCOEN PLUS (MEYS, CZ.02.1.01/0.0/0.0/15_003/0000469), the RECETOX research infrastructure (MEYS, LM2015051; CZ.02.1.01/0.0/0.0/16_013/0001761).
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**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

**POSTER SESSIONS 3 AND 4 – Wednesday and Thursday –** all even number posters will be on display.

*AWARD WINNERS

BIOMEDICAL

P-214

Surveying the spectrum of bacterial sialic acids using high resolution mass spectrometry

PRESENTING AUTHOR: **Hugo Kleikamp, TU Delft, Applied Sciences, Biotechnology, Netherlands**

CO-AUTHORS: **Yuemei Lin, Mark van Loosdrecht, Martin Pabst**

Over the past decades nonulosonic acids have been increasingly found in (pathogenic) prokaryotes, decorating the contact surface to their external environments. A recent phylogenetic large-scale study discovered a surprisingly wide distribution of an ancient core nonulosonic acid biosynthetic pathway. However, predicted ORFs often do not encode for functional entities, and most importantly, genomics alone does not reveal the important post synthesis processing events, which make nonulosonic acids so unique. There are many open questions surrounding the evolution and utilisation of nonulosonic acids in prokaryotes, as well as their role in diseases. Unfortunately, their analysis with conventional methods is a laborious process. Therefore, we developed a fast but highly specific assay for the screening of existing nonulosonic acids, and the discovery of novel nonulosonic acid variants, using high-resolution mass spectrometry, in combination with a specifically developed data processing tool. The high specificity of the assay allows for direct analysis from crude cell lysates, and analysis of low abundant species in complex communities. The application of high-resolution mass spectrometry also enables stable isotope incorporation experiments to follow novel biosynthetic routes in bacteria. Using this newly developed assay, we provide a comparative large-scale study on the diversity and distribution of different classes of nonulosonic acids across a variety of species, including purely environmental samples. The developed assay serves as a tool to explore complex samples and communities to study the broader ecological context of this prominent, but still poorly analysed class of sugars.

P-215

Metabolic insights from plasma samples of Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) patients

PRESENTING AUTHOR: **Arnaud Germain, Cornell University, United States**

CO-AUTHORS: **Arnaud Germain, David Ruppert, Susan M. Levine, Maureen R. Hanson**

The well-documented constellation of debilitating symptoms endured by Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) patients implicates a severe disruption of homeostasis of metabolic networks. Here we reflect on the knowledge generated by the plasma datasets issued from our three cohorts totaling 113 distinct female subjects, 47 controls and 66 patients. The chemical fingerprint of our first cohort consisted of 361 metabolites, and conclusions from our quantitative analysis focus on disturbances in fatty acid and lipid metabolism. For our second cohort, the Metabolon® platform expanded our vision to 832 data points divided in eight super-pathways and imply a redox imbalance in disease symptomatology. Our latest dataset consists of 768 metabolic measurements from Metabolon®'s global metabolomics technology, while their complex lipid panel quantified 1,022 lipids. The concentrations of a few dipeptides were negatively affected in the patient cohort, indicating a potential disruption in protein catabolism, while differences in abundances of certain triacylglycerols (TAGs) were also detected. These molecules are fatty acids, which are required for various bodily functions, acting as the most important caloric source of energy homeostasis in mammals, serving as a flexible on-demand energy depot. TAGs are also an integral part of the bloodstream where they facilitate absorption of nutrient molecules such as fat-soluble vitamins. Abnormally high levels have, however, been linked to a wide range of cardiovascular problems. Our studies represent valuable steps in understanding the underlying problems that manifest in patients affected by this disease of unknown cause and poor prognosis.

P-216

Effects of dipeptidyl peptidase 4 inhibitor on the human urinary metabolomic profiling

PRESENTING AUTHOR: **Jihyun Kang, Seoul National University, South Korea**

CO-AUTHORS: **Bora Kim, Yun Kim, SeungHwan Lee, Kyung-Sang Yu, In-Jin Jang, Joo-Youn Cho**

Dipeptidyl-peptidase IV inhibitor (DPP-4) is oral medication for type 2 diabetes mellitus that stimulates insulin secretion by inhibiting degradation of the incretin, glucagon-like peptide-1 and glucose-dependent insulinotropic peptide. Therefore, DPP-4 inhibitors maintain blood glucose homeostasis by suppressing glucagon secretion and slowing gastric emptying. In this study, we aimed to discover sets of biomarkers for effects of DPP-4 inhibitors in urine samples from healthy volunteers and explore its underlying mechanism of action using metabolomics approach. Urine samples from 34 healthy subjects treated with DPP-4 inhibitor were collected at baseline and after 7 days treatment period and analyzed using Ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS) metabolomics platform. A total of 4775 metabolic features were detected through UPLC-QTOF-MS and putative markers were identified (FDR adjusted P<0.0005, power >0.9995). After treatment of DPP-4 inhibitor, healthy subjects showed relative intensity increase in 9 endogenous metabolites, dihydroxyfumaric acid, tyrosylthreonine, glutamine, leucylproline, ethylglycine, threonic acid, n-methyl-l-glutamic acid, methionylalanine, and kynurenic acid. 5 endogenous metabolites, including histidinal, arabinonic acid, phenylalanylalanine, alanylhydroxyproline, and capryloylglycine were significantly down regulated after treatment. Among the putative markers, 5 dipeptides showed differences after treatment. DPP-4 preferentially cleaves Xaa-Pro or Xaa-Ala dipeptides from the N-terminus of polypeptides and preventing the degradation of incretin hormones, by DPP-4 inhibition, speculated relative intensity of dipeptides change in postdose. In this study, pharmacometabolomics provides powerful tools for identifying effects of global metabolomics profiling by DPP-4 inhibitor treatment.
<figure><img src="image_66.png" /></figure>

**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

**POSTER SESSIONS 3 AND 4 – Wednesday and Thursday –** all even number posters will be on display.

*AWARD WINNERS

BIOMEDICAL

P-217

Determination of metabolic profiles of serum samples from women with PCOS based on LC-MS/GC-MS techniques

PRESENTING AUTHOR: Anna Stefaniak, Medical University of Gdańsk, Department of Biopharmacy and Pharmacodynamics, Poland

CO-AUTHORS: Buszewska-Forajta Magdalena, Szybiak Aleksandra, Kowalewska Agnieszka, Rachon Dominik, Markuszewski Michał Jan

Despite such a high incidence, the pathogenesis of polycystic ovary syndrome still remains unexplained. PCOS is multifactorial disorder, which is diagnosed in about 10% of women of reproductive age. It is connected with ovulatory dysfunction and polycystic ovaries appearance and also with higher level of androgens concentration. Therefore, PCOS is one of the main reasons of female infertility. Besides to endocrine dysfunction, it may cause the development of different metabolic disorders. The use of the metabolomic approach in a field of PCOS aims to identify specific metabolic pathways potentially involved in the pathophysiology of this frequent disorder. Determination and comparison of the metabolic profiles of serum samples obtained from women with PCOS and healthy controls was the main aim of this study. Untargeted metabolomic analysis was conducted with the use two complementary analytical techniques: liquid chromatography and gas chromatography coupled with mass spectrometry. Next step was identification metabolites specific for biochemical pathways disturbed in PCOS. Univariate and multivariate statistical analysis showed the difference of metabolic profiles in studied groups of women. During the untargeted metabolomic analysis of serum samples, a few metabolites connected with metabolic disorders of amino acids, carbohydrates, steroid hormones, lipids, purines and citric acid cycle were identified. Understanding the pathomechanism of this syndrome and the identification of new potential biomarkers of PCOS will contribute to its early recognition and effective treatment.

P-218

Metabolic characterization and predictive model construction for partial hepatectomy mortality in porcine model

PRESENTING AUTHOR: Hyung Do Kwon, Kookmin University, South Korea

CO-AUTHORS: Do Yup Lee

The molecular and cellular mechanism in liver regeneration process following partial hepatectomy has been well known; however, hepatectomy mortality still remains significant problem in clinics. Therefore, we explored metabolic features reflected in serum metabolite after partial hepatectomy in porcine model. Pig shares the similarity in anatomy and physiology with human so it has been regarded as a proper model for liver study. In this study, based on gas chromatography time-of-flight mass spectrometry (GC-TOF-MS), primary metabolite profile was acquired from three groups, sham, 70%, and 90% liver resection groups. Un-supervised multivariate statistics revealed distinctive profiles of the three groups. And subsequent modeling based on orthogonal projection in latent structure-discriminant analysis (OPLS-DA) proposed key molecular features that discriminated hepatectomy mortality. The linear combination with malic acid and methionine showed the best predictive power in which the area under the curve (AUC) ranged from 0.939 to 0.989. The current study provides the understanding of liver regeneration and metabolic capacity impaired by hepatectomy.

P-219

Metabolomic profiling of orbital lymphoproliferative diseases

PRESENTING AUTHOR: Eri yamaguchi, Tokyo Medical University, Japan

CO-AUTHORS: Eri Yamaguchi, Hiroyuki Shimizu, Yasuko Aita, Atsumi Tomita, Yoshihiko Usui, Hiroshi Goto, Masahiro Sugimoto.

Orbital lymphoproliferative diseases, particularly mucosa-associated lymphoid tissue (MALT) lymphoma and IgG4-related ocular disease (IgG4-ROD), have similar clinical and also histopathological features, and are therefore often difficult to differentiate. In this study, we conducted metabolomic profiling of IgG4-ROD and MALT lymphoma tissue samples. Six samples of orbital MALT lymphoma (mean age 62.3 years; 4 males and 2 females) and 11 samples of IgG4-ROD (mean age 64.8 years, 4 males and 7 females) and their matched controls samples were included. Liquid chromatography with time-of-flight mass spectrometry (LC-TOF-MS) was used for profiling lipid soluble metabolites. Principal component analysis clearly showed four groups: adipose tissue, tumor tissue, IgG4-ROD, and MALT lymphoma. Comparison between orbital adipose tissues and matched samples showed significant differences in 174 and 132 metabolites in IgG4-ROD and MALT lymphoma, respectively. Comparison between IgG4-ROD and MALT lymphoma showed significant differences in 12 metabolites. These tissue-specific difference would help understanding the pathogenesis of these diseases.
<figure><img src="image_67.png" /></figure>

**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

**POSTER SESSIONS 3 AND 4 – Wednesday and Thursday –** all even number posters will be on display.

*AWARD WINNERS

BIOMEDICAL

P-220

Metabolomics in Nonhuman Primate Models for Radiation Biodosimetry in Emergency Preparedness

PRESENTING AUTHOR: Albert Fornace, Jr., Georgetown University, United States

CO-AUTHORS: Evan L. Pannkuk, Evagelia C. Laiakis, Kirandeep Gill, Shreyans K. Jain, Khyati Y. Mehta, Denise Nishita, Kim Bujold, James Bakke, Janet Gahagen, Simon Authier, Polly Chang, Albert J. Fornace Jr.

Rapid assessment of radiation signatures in noninvasive biofluids may aid in assigning proper medical treatments for acute radiation syndrome (ARS) and delegating limited resources after a nuclear disaster. Metabolomic platforms allow for rapid screening of biofluid signatures and show promise in differentiating radiation dose and time post-exposure. Here, we use global metabolomics to differentiate temporal effects found in nonhuman primate (NHP) urine and serum small molecule signatures after a 4 Gy total body gamma irradiation. Random Forests analysis show days (1 – 60) after a 4 Gy exposure can be readily separated and provide critical information for ARS treatment. Eight compounds involved in protein metabolism, fatty acid β-oxidation, DNA base deamination, and general energy metabolism were identified in each biofluid. The greatest perturbations were seen at 1 d in urine highlighted by increases in carnitine, acylcarnitines, xanthine, creatine, 7-methylguanine, and dimethylarginine. Changes in serum metabolite concentrations were more dynamic (1 – 21 d) including carnitine, acylcarnitines, guanine, oleamide, and hypoxanthine. We also developed a targeted multiplex assay validating a six compound panel (hypoxanthine, carnitine, acetylcarnitine, proline, taurine, and citrulline) previously identified in a training cohort at 7 d after a 4 Gy exposure. Receiver operating characteristic (ROC) curve analysis indicated the highest sensitivity and specificity for carnitine and acetylcarnitine (urine, area under the curve [AUC] = 0.99) and taurine, carnitine, and hypoxanthine (serum, AUC = 0.95). These results highlight the utility of mass spectrometry (MS) platforms to differentiate time post-exposure and acquire reliable quantitative biomarker panels for classifying exposed individuals.

P-221

Effect of repeated freeze-thaw cycles on NMR measured lipoproteins and metabolites in biofluids

PRESENTING AUTHOR: Guro Giskeødegård, NTNU, Norway

CO-AUTHORS: Feng Wang, Julia Debik, Trygve Andreassen, Leslie R. Euceda, Tonje H. Haukaas, Hartmut Schäfer, Tone F. Bathen

Metabolic profiling of biofluids by Nuclear Magnetic Resonance (NMR) spectroscopy serves as an important tool in disease characterization, and its accuracy largely depends on the quality of the samples. We aimed to explore possible effects of repeated freeze and thaw cycles (FTCs) on concentrations of lipoprotein parameters in serum and metabolite concentrations in serum and urine samples. Serum and urine samples were collected from two sets of 20 healthy donors, and aliquots were subjected to 1–5 FTCs. Samples were subsequently analyzed by NMR spectroscopy, and lipoprotein parameters and metabolites were quantified using a commercial analytical platform. In total, 112 lipoprotein parameters, and 20 serum and 35 urine metabolites were quantified. Principal component analysis showed no systematic changes related to FTCs. Samples from the same donor were closely clustered, showing a higher between-subject variation than within-subject variation. The coefficients of variation were small (<2.1%, <1.4% and <2.2% for lipoprotein parameters, and serum and urine metabolites, respectively), showing a low variation in samples from the same donor. Systematic effects of FTCs on the concentrations were tested by Wilcoxon tests between consecutive FTCs. Only five lipoprotein parameters (VLCH, VLFC, V1CH, V4CH, and V3FC) differed significantly between FTC 3 and 4 (p-values<0.05), while other lipoprotein and metabolite concentrations showed no significant change. In conclusion, five FTCs did not significantly alter the concentrations of serum and urine metabolites, and introduced only minimal changes to serum lipoprotein parameters evaluated by the NMR- based platform.

P-222

Lipidomic analysis of acellular cod skin xenograft versus traditional dermal substitutes

PRESENTING AUTHOR: Aristotelis Kotronoulas, Center for Systems Biology, University of Iceland, Iceland

CO-AUTHORS: H. S. Jónasdóttir, R. S. Sigurðardóttir S. Halldórsson, G. G. Haraldsson, Óttar Rolfsson

Acellular fish skin (ACS) has emerged as a dermal substitute used to promote wound healing. The effects of ACS have been associated to the poly-unsaturated fatty acids (PUFA) although their presence in processed ACS has not yet been determined. Here, we compared the total free fatty acid (FFA) and lipid profiles of ACS to that of human cadaver skin (HCS) and two grafts of bovine origin. FFA quantification was performed by GC-FID and the lipid profile analysis by untargeted UPLC-MS/MS. Our results showed that ACS contained significantly higher omega3 FAs and PUFA than all other tested grafts, with percentages that exceeded 20% and 30% of the total FAs content respectively. The majority of the PUFAs found in HCS were omega-6 while the two grafts of bovine origin presented insignificant PUFA amounts. Regarding the grafts' lipidomic profile, both unsupervised PCA and supervised PLS analysis demonstrated that all analyzed materials had very different lipid composition. Based on the heatmap and the hierarchical clustering, approximately 50 lipids from positive and negative ESI mode were identified as the most distinctive between the four materials and their possible structure was annotated. Further analysis of the lipid profiles demonstrated that phosphatidylcholine containing 20:5 and 22:6 omega3 PUFAs were characteristic for the ACS grafts. Our study showed that the lipid and fatty acid content of the ACS is different to other wound grafting materials. The high omega3 PUFA found may be associated with a range of positive wound healing mechanisms including anti-inflammatory activity, cell migration and stimulation.
**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

**POSTER SESSIONS 3 AND 4 – Wednesday and Thursday –** all even number posters will be on display.

*AWARD WINNERS

BIOMEDICAL

P-223 Improving the Diagnosis, Treatment, and Prevention of Chronic Kidney Disease by Establishing a Mass Spectrometric Reference Measurement Procedure for Parathyroid Hormone

PRESENTING AUTHOR: Candice Ulmer, Centers for Disease Control and Prevention, United States

CO-AUTHORS: Hubert Vesper

Parathyroid hormone (PTH) is a key biomarker for chronic kidney diseases, one of the leading health conditions in the USA. In addition, the measurement of parathyroid hormone (PTH) along with the respective fragmented versions of this hormone in serum and/or plasma is necessary for the detection, diagnosis, and prevention of calcium, phosphate, and vitamin D disorders. Current quantitation approaches suffer from sensitivity, PTH instability, and method measurement variability. CDC's PTH standardization program is improving the accuracy and reliability of measurements performed in patient care. A reference measurement procedure was developed for the quantitation of intact PTH (1-84) and select PTH fragments in serum. Stable isotope labeled internal standards were spiked prior to sample preparation. PTH isolation was optimized using immunocapture methodologies with monoclonal antibodies. The UHPLC-HRMS-based reference procedure allows for the highly specific detection and quantitation of intact PTH (z=6) without isotopic interference from the labeled internal standard. Preliminary results demonstrate the ability to quantify intact PTH and PTH fragments within a 5-ppm mass accuracy window without enzymatic digestion. Calibration curves using stable isotope labeled internal standards were generated for iPTH, N-terminal PTH fragments, C-terminal PTH fragments, and mid-region PTH fragments. Initial assessments demonstrate excellent linearity (R2 = 0.991-0.997), reproducibility, and sufficient sensitivity of the UHPLC-HRMS system with a linear range of 15-3000 pg/mL PTH. This method was applied to samples from healthy patients and patients with different CKD stages. PTH and its fragments were detectable in these samples.

P-224 Metabolic profiling of renal cystic fluids in autosomal dominant polycystic kidney disease patients

PRESENTING AUTHOR: Woori Chae, Seoul National University, South Korea

CO-AUTHORS: Hyunjin Ryu, Seung Seok Han, Joo-Youn Cho

Autosomal dominant polycystic kidney disease (ADPKD) is a genetic disorder resulted from mutations in PKD1 and PKD2 genes. It occurs in about 1 in 400 to 1000 and is the fourth leading cause of end-stage renal disease. Tolvaptan, a vasopressin receptor antagonist, is the first line treatment for ADPKD but there are various known side effects. Currently, no drug has been developed to inhibit the formation and development of polycysts. In this study, we compared the metabolic profiles of renal cyst fluids obtained from patients with ADPKD and those with simple cysts as a control to find a new therapeutic target. Differences in metabolite profiles according to various clinical parameters were also compared in ADPKD patients. Fifteen cyst fluids from ADPKD patients and twenty-seven cyst fluid from patients with simple cysts were collected and analyzed with both reverse phase and hydrophilic interaction liquid chromatography column using ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry. Though we did not find any significant differences between ADPKD and simple cyst groups, several statistically significant metabolites were found in ADPKD patients according to the incidence of advanced chronic kidney disease. It was the first report of comparative human metabolome profile of ADPKD and simple cyst fluids, which may contribute to understanding the pathogenesis of ADPKD.

P-225 Analysis of low-molecular-weight fraction of honeybee venom using mass spectrometry-based methodologies

PRESENTING AUTHOR: Agnieszka Klupczynska, Department of Inorganic and Analytical Chemistry, Poznan University of Medical Sciences, Poland

CO-AUTHORS: Klupczynska Agnieszka, Dereziński Paweł, Plewa Szymon, Pawlak Magdalena, Garrett Timothy J., Rubio Vanessa Y., Kokot Zenon J., Matysiak Jan

Honeybee venom is a natural product that pose danger to human health, but also have therapeutic properties. Peptides and proteins constitute the majority of its dry mass. However, there are also multiple low-molecular-weight compounds present in honeybee venom, which have various biological functions. The aim of the study was to identify and quantify the metabolites present in that bee product. A quadrupole-Orbitrap mass spectrometer coupled to a liquid chromatograph was used for untargeted metabolomics. Moreover, two targeted methodologies were applied that utilized a triple quadrupole mass spectrometer coupled to a liquid chromatograph. Polar metabolites (amino acids, biogenic amines, organic acids) were determined with chromatographic separation, while lipid metabolites were analyzed using flow injection analysis. The untargeted research allowed to identify several dozens of metabolites present in honeybee venom, which belong to such classes as: amino acids, catecholamines, organic acids, fatty acids, purines, pyrimidines and saccharides. 42 metabolites were quantified using targeted platforms. Among metabolites determined in the highest amount were citric acid, glycine, proline, dopamine and histamine. The research revealed that honeybee venom is a highly complex matrix not only in relation to proteins, but also to metabolites. Moreover, the variability in the composition of venom was observed. Factors influencing the changes in venom composition need to be studied and the composition need to be controlled, i.a. in order to ensure the safety of using venom as a product for medical formulations. The project received support from the National Science Centre, Poland (grant number: 2016/23/D/NZ7/03949).
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**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

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P-226 Peripheral vs. Cord blood: assessment of inflammatory, growth factors and metabolomic profile for the production of serum eye drops

PRESENTING AUTHOR: Thelma Pertinhez, University of Parma, Dept. Medicine and Surgery, Italy

CO-AUTHORS: Manuela Guardi, Lucia Merolle, Chiara Marraccini, Roberto Baricchi

Dry Eye Disease is characterized by tear increased osmolality and inflammation. Serum eye drops have emerged as a therapy for these ocular surface disorders (1). The rationale for the topical use of serum drops is based on their composition similarity to tears, due to the presence of growth factors, cytokines and vitamins, essential to maintain corneal homeostasis and wellness. Given the lack of guidelines for serum drops production, our aim was to compare the inflammatory, growth factors and metabolomic profiles from two homologous blood sources: cord blood serum (CBS) and adult peripheral blood serum (PBS), in order to pave the way for a standardization preparations procedure. This study involved S. Orsola-Malpighi Teaching Hospital (Bologna) and Azienda USL-IRCCS di Reggio Emilia. Metabolomics was performed by Nuclear Magnetic Resonance and Luminex Assay was used to measure cytokines and growth factors. Over 40 metabolites were identified and quantified. We found significant differences between the two blood-derived serum drops, with higher concentration of metabolites in CBS than PBS. The main differences were in choline, β-hydroxybutyrate, acetoacetate and glucose levels. As expected CBS showed higher concentration of TGF-α, β-NGF, FGF levels with respect to PBS. Inflammation pathway was not stimulated in CBS and nor in PBS. These results indicate that both products are safe, although the high level of TGF-α might be dangerous for patient with neoplastic disease. 1. Hessen M, Akpek EK. J Ophthalmic Vis Res. 2014, 9:240-50. Supported by Centro Regional Sangue, RER, Italy.

P-227 LC-MS/MS Method Development and Validation for Potential Early Biomarkers in Parkinson's Disease

PRESENTING AUTHOR: Jorge Sáiz, Universidad San Pablo CEU, Spain

CO-AUTHORS: Marcela Konjevod, Alberto Bergareche, Pilar Amiano, Fernando Goñi, Eva Ardanaz, José María Huerta, Coral Barbas

Parkinson's disease (PD) is an age-related, debilitating, severe neurodegenerative disorder characterized by degeneration of dopaminergic neurons. The identification of biomarkers for the early diagnosis of PD could improve treatment and decrease misdiagnosis. The aim of this study was to develop and validate LC-MS/MS methods that will allow to find compounds that could represent potential biomarkers. The study enrolled plasma samples as part of "The European Prospective Investigation into Cancer and Nutrition" study that were collected from healthy donors, who were followed up for 15 years. Among them, some developed PD in the following years. These samples are of great value for the discovery of biomarkers for the early diagnosis of PD and makes this study a high risk project involving the validation of biomarkers in healthy subjects. The analytes were chosen according to findings in previous studies on Parkinson's disease to be measured in target. 5 methods were developed in LC-QqQ-MS for the quantitation of groups according to their chemical characteristics. The methods were validated in terms of linearity, LOQ, repeatability, intermediate precision and stability. The LOQ for most of the compounds were below 500 ppb. The results showed good intermediate precision (1.8-32.4 %RSD), repeatability (0.3-10.2 %RSD) and stability at 4º C, as well as at room temperature for the analyzed compounds. Recoveries were found to be between 40 and 100 %. Statistically significant compounds belonged to the groups of fatty acids, sugars, alcohols and organic acids.

P-228 A Novel Metabolic Signature To Predict the Requirement of Dialysis or Renal Transplantation in Patients with Chronic Kidney Disease

PRESENTING AUTHOR: Helena U. Zacharias, University Medicine Greifswald, Germany

CO-AUTHORS: Michael Altenbuchinger, Ulla T. Schultheiss, Claudia Samol, Fruzsina Kotsis, Inga Poguntke, Peggy Sekula, Jan Krumsiek, Anna Köttgen, Rainer Spang, Peter J. Oefner, Wolfram Gronwald

Identification of chronic kidney disease patients at risk of progressing to end-stage renal disease (ESRD) is essential for treatment decision-making and clinical trial design. Here [1], we explored whether proton nuclear magnetic resonance (NMR) spectroscopy of blood plasma improves the currently best performing kidney failure risk equation, the so-called Tangri score. Our study cohort comprised 4640 participants from the German Chronic Kidney Disease (GCKD) study, of whom 185 (3.99%) progressed over a mean observation time of 3.70 ± 0.88 years to ESRD requiring either dialysis or transplantation. The original four-variable Tangri risk equation yielded a C statistic of 0.863 (95% CI, 0.831–0.900). Upon inclusion of NMR features by state-of-the-art machine learning methods, the C statistic improved to 0.875 (95% CI, 0.850–0.911), thereby outperforming the Tangri score in 94 out of 100 subsampling rounds. Of the 24 NMR features included in the model, creatinine, high-density lipoprotein, valine, acetyl groups of glycoproteins, and Ca2+-EDTA carried the highest weights. In conclusion, proton NMR-based plasma fingerprinting improved markedly the detection of patients at risk of developing ESRD, thus enabling enhanced patient treatment. [1] Zacharias, et al., A Novel Metabolic Signature To Predict the Requirement of Dialysis or Renal Transplantation in Patients with Chronic Kidney Disease, Journal of Proteome Research, 2019, DOI: 10.1021/acs.jproteome.8b00983.
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P-229

DAC-Met: Exploring the metabolic fate of the herbal components in "maoto" decoction through a differential annotation strategy

PRESENTING AUTHOR: Katsuya Ohbuchi, Tsumura & Co., Japan

CO-AUTHORS: Nozomu Sakurai, Hiroyuki Kitagawa, Hirotaka Kushida, Akinori Nishi, Masahiro Yamamoto, Kazuhiro Hanazaki, Masanori Arita

Natural products are attractive as drug leads. Traditional herbal medicine (THM) is expected to overcome the limitations of a single chemical agent, and is prescribed by over 80% of medical doctors in Japan together with Western drugs. To elucidate their molecular action, comprehensive profiling of the THM components and their bio-conversion is essential. We present a data-independent (non-targeted) strategy, named DAC-Met (Differential Annotation of Converted Metabolites), to explore metabolized plasma components using high-resolution mass spectrometry. As a showcase, we chose the effect of "maoto", a mixture of ephedra herb, apricot kernel, cinnamon bark, and glycyrrhiza root, widely prescribed for febrile symptoms including fever and influenza in Asian countries. Maoto was administered to four healthy volunteers. Blood samples were collected before and after the maoto treatment, 7-time points in total. We performed plasma metabolome analyses and found maoto-derived metabolites, including known major components (ephedrine, prunasin, cinnamic acid, and glycyrrhetinic acid). Using the mass difference of major bio-conversions, we additionally estimated the structure of 11 unidentified metabolites, and 4 were later confirmed with authentic standards. With literature knowledge, we also reconstructed the metabolic pathway of maoto components. From time-resolved measurements, their kinetic profiles became clear too: many showed rapid absorption and elimination but glycyrrhetinic acid and related conjugates became prominent 4 hours after administration. The converted metabolites and their diverse kinetics revealed in this study exemplify the complex mechanism of THM action. This talk covers detailed strategy of our differential analysis and the future prospect of natural product research.

P-230

Lipidome modules underlying subclinical osteoporosis and atherosclerosis comorbidity: The Cardiovascular Risk in Young Finns Study

PRESENTING AUTHOR: Binisha Hamal Mishra, Tampere University, Finland

CO-AUTHORS: Pashupati P. Mishra, Nina Mononen, Mika Kähönen Olli T. Raitakari, Reijo Laaksonen, Terho Lehtimäki

**Background:** Evidences suggest that atherosclerosis and osteoporosis are comorbid conditions. However, the underlying mechanism is largely unknown. Subjects and methods: System-level analysis of untargeted serum lipidome was performed to identify networks of lipid species underlying atherosclerosis and osteoporosis comorbidity. The study is based on Young Finns Study cohort from 2007 follow-up (number:1194, age:30-45 years, women:59.30%). Carotid and bulbus intima media thickness was used as proxy for subclinical atherosclerosis. Total mineral density and area of heel bone, distal radius, distal radius spongy bone, radial shaft cortical bone, distal tibia, distal tibia spongy bone and tibial shaft cortical bone were used as proxy for subclinical osteoporosis. Co-expression network analysis was performed to identify modules of interconnected lipid species. Expression profiles in modules were summarized by first principal component termed as module eigenlipid. Lipid modules that were significantly correlated with both bone and artery variables were considered to be related to comorbidity. Results: A module with 124 lipid species was significantly associated with both artery (Pearson correlation: 0.16, p-value: 6e-08) and bone variables (Pearson correlation: 0.23, p-value: 1e-15). Majority of the lipid species in the module belonged to classes glycerolipid, glycerophospholipid and sphingolipid. Glycerolipids were found statistically most strongly associated with the comorbidity. The identified module contained high-risk ceramides that confirms previously suggested association between ceramides and cardiovascular outcomes, and suggests its potential role in subclinical osteoporosis. Conclusion: This study identified a lipid module associated with subclinical phase of atherosclerosis and osteoporosis comorbidity and provides potential biomarker for the comorbidity diagnosis.

P-231

LC-MS analysis of free fatty acids in acromegaly patients

PRESENTING AUTHOR: Bettina Gürtl, CeMM - Research Center for Molecular Medicine, Austria

CO-AUTHORS: Paul Fellinger, Yvonne Winhofer-Stöckl, Kristaps Klavins

Acromegaly is a rare disease resulting from an overproduction of growth hormone which leads to changes in the physical appearance and inner organs. Acromegaly patients also display an extremely low hepatocellular lipid content, despite pronounced insulin resistance. Therefore, studying acromegaly may help to elucidate underlying antisteatotic pathways in non-fatty liver disease. Fatty acids play an essential role in lipogenesis and hence free fatty acid levels and distributions could be used to reflect alterations in these pathways. In this work a comprehensive analysis of free fatty acids in plasma samples from acromegaly patients was performed by liquid chromatography-mass spectrometry (LC-MS). The analysis of fatty acids is often conducted by gas chromatography-mass spectrometry due to the low chromatographic performance and ionization efficiency of LC-MS. To overcome these challenges, a chemical derivatization reaction, based on 2-hydrazinoquinoline was established. The sample preparation was optimized by testing different extraction solvents, evaluating the influence of used consumables and investigating different durations of the derivatization reaction. For the LC-MS analysis a reversed phase chromatographic separation coupled to a high resolution mass spectrometer was used. The developed method provides a comprehensive coverage of 25 fatty acids including short- and long-chain fatty acids. Plasma samples from 15 patients with active acromegaly and 17 healthy controls, matched for age, BMI, gender and body composition (fat free mass), were analyzed using the established workflow. The obtained quantitative profiles of free fatty acids showed that the concentration of the majority of detected free fatty acids was lower in acromegaly patients.
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P-232

Fish mucus collection strategies for metabolomics: a distinct response to Aqui-S and benzocaine anaesthetics

PRESENTING AUTHOR: Lada Ivanova, Norwegian Veterinary Institute, Norway

CO-AUTHORS: Rangel-Huerta, O.D., Tartor, H., Gjessing, M.C., Dahle, M.K., Thoen, E., Uhlig, S.

Fish mucus has recently received significant interest in aquaculture. Mucus on accessible/reachable mucosal surfaces including skin and gills can be collected in a simple and non-invasive manner, providing the relevant information about the health status of the fish. However, the sampling approach may be challenging source of bias in comparative metabolomics of fish skin and gill mucus. Therefore, standardisation of sampling procedures, as well as increasing the understanding of potential sources of error are highly valuable. Different anaesthetics facilitate work with fish and reduce the level of stress, respecting the refining principle of using fish as laboratory animals. The addition of anaesthetics to the water is a common procedure prior to sampling in aquaculture. There is also a trend towards reducing the use of synthetic anaesthetics to avoid unwanted effects on the environment. Therefore, there is a growing interest in the use of natural products such as clove oil (Aqui-S) as a viable alternative to synthetic anaesthesia. This study aims to explore the effect of Aqui-S and benzocaine on the composition of the skin and gill mucus obtained after sedation of the fish. By using targeted and untargeted metabolomics, alterations in the metabolome of fish mucus induced by both anaesthetics have been evaluated. The results will be used further to improve experimental designs used in metabolomics, realising such alterations.

P-233

Evaluation of inhibitory effect of grapefruit juice on intestinal CYP3A activity

PRESENTING AUTHOR: Yujin Lee, Seoul National University, South Korea

CO-AUTHORS: Soyoung Lee, Andrew HyoungJin Kim, Joo-Youn Cho

The cytochrome P450 (CYP) 3A enzyme is major phase I drug metabolizing enzyme which accounts for 40% and 80% of total CYP in the liver and intestine, respectively. Among the factors affecting CYP3A activity, grapefruit juice is known to inhibit intestinal CYP3A activities which may lead to undesired drug interaction. We aimed to evaluate the intestinal CYP3A inhibitory effect by grapefruit juice using midazolam and endogenous metabolic markers. Fifteen Korean healthy male subjects completed the study. Subjects received oral and intravenous midazolam as probe drug. CYP3A activity was evaluated using midazolam clearance and endogenous metabolic markers. To inhibit intestinal and total (both hepatic and intestinal) CYP3A activities, grapefruit juice and clarithromycin were used, respectively. Urinary and plasma steroids were quantified by using GC-MS and the concentration of midazolam and its metabolites were quantified using LC-MS. In total CYP3A inhibition phase, urinary markers, 6β-OH-cortisone/cortisone and 6β-OH-cortisol/cortisol, and the concentration of plasma marker, 4β-OH-cholesterol, decreased 0.4, 0.5 and 0.3-fold, respectively. Midazolam clearance was also significantly decreased 0.3-fold compared to that of control. However, in intestinal CYP3A inhibition phase, both midazolam clearance and endogenous metabolic markers were not significantly decreased except for 6β-OH-cortisone/cortisone ratio was significantly decreased 0.6-fold compared to that of control. In conclusion, the inhibitory effect of clarithromycin on CYP3A activity was significant. However, the inhibitory effect of grapefruit juice was negligible. Therefore, we speculate that there will be no significant interaction between grapefruit juice and CYP3A substrate drugs.

P-234

Central and peripheral metabolomics in a model of depression: biosignatures of drug responder and non-responder mice

PRESENTING AUTHOR: Sylvie Mavel, University of Tours, France

CO-AUTHORS: Sylvie Mavel, Antoine Lefèvre, Marc Legrand, Laurent Galineau, Lydie Nadal-Desbarats, Catherine Belzung, Patrick Emond

Major depressive disorder is a heterogeneous disorder with a wide spectrum of symptoms leading to disability, suicide and physical disorders. Selective serotonin reuptake inhibitors (SSRIs) are ones of the most commonly used drugs of treatment of depression. For unknown reasons, a great number of patients do not show any improvement during drug treatment (30-40%). The underlying molecular mechanisms of depression remain unknown. To improve knowledge of pathophysiology and to search biomarkers to help prognostic of antidepressant treatment response in patient, we employed a LC-HRMS metabolomics approach in a BALB/c induced mouse model of depression to investigate metabolic changes in 6 specific brain regions, after 5 weeks with a fluoxetine treatment. From 770 cumulated targeted metabolites, 68 discriminant Variable Importance in Projection (VIP) were shown to discriminate vehicle-treated group from drug responder and non-responder mice, after OPLS-DA. This central metabolomics approach is completed by a peripheral analysis through plasma analysis. A less robust OPS-DA model was obtained, but 27 discriminant metabolites (over 197 robustly analyzed) were altered in plasma. Comparison of central VIP with peripheral VIP leads to 7 common metabolites. This study shows the complementary of these two compartments, i.e. cerebral and plasma metabolome. From this, specific brain regions shown particular metabolism which could explain some aspects of the depressive pathophysiology and some biomarkers could be proposed to anticipate antidepressant treatment response.
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P-235 Differences in metabolic profiles of venous and arterial umbilical cord blood

PRESENTING AUTHOR: Olle Hartvigsson, Chalmers University of Technology, Sweden

CO-AUTHORS: Alastair Ross, Carl Brunius, Malin Barman, Ann-Sofie Sandberg

Analysing umbilical cord blood is important for assessing neonatal health and development. The easiest and most common collection procedure is to squeeze blood out of the cord after it is severed. Hence a mixture of venous (from mother to child) and arterial (from child to mother) blood is collected. This study aimed to determine key differences in metabolite profiles between venous and arterial cord blood plasma. The metabolome of venous, arterial and mixed squeezed umbilical cord blood was analysed from 52 children using a combination of targeted and untargeted GC-MS/ MS. Data was analysed by multilevel Random Forest (ML-RF) in a repeated double cross validation framework incorporated with unbiased variable selection. In pairwise analysis of arterial and venous blood, approximately 75% of samples were correctly classified (p=0.0078). Arterial blood had higher concentrations of glucose, sorbose and galactose than venous blood which contained higher levels of α-ketoglutaric acid, L-glutamic acid, homocysteine and several other metabolites. Mixed blood had a metabolic profile that was in-between the arterial and venous blood, but could not be classified properly by multivariate models. Our results highlight that cord blood sampling with non-systematic mixing of arterial and venous blood induces undesirable variability in metabolomics analyses. We therefore conclude that control of the sampling procedure is imperative during metabolomics analyses, especially when energy or amino acid metabolism are relevant for the research question.

P-236 Comparison of plasma metabolic profiling in allergic rhinitis and asthma by 1H NMR in early childhood

PRESENTING AUTHOR: Meng-Han Chiang, Research Fellow, Taiwan

CO-AUTHORS: Chih-Yung Chiu, Gigin Lin

Allergic rhinitis and asthma are both chronic heterogeneous disorders with the high prevalence of coexistence. Clinically, house dust mite sensitization with elevated serum immunoglobulin E (IgE) levels is a significant factor in relation to both allergic rhinitis and asthma. However, few studies have addressed the potential molecular mechanisms between allergic rhinitis and asthma, especially in children. Metabolomics provides a complementary functional approach to the molecular functions interacted with the host, offering the opportunity to characterize diseases effectively. The aim of this study was to investigate plasma metabolic profiling in children with allergic rhinitis and asthma. Plasma samples were collected and metabolites were analyzed by 1H-nuclear magnetic resonance spectroscopy (NMR). NMR spectra were processed by an open source software NMRProcFlow and subsequently analyzed by MetaboAnalyst. In this study, a total of 111 children with mean age of 4.9 ± 0.5 years were enrolled. Among them, there were 27 asthmatics, 44 allergic rhinitis, and 40 healthy controls. After analysis, total 104 buckets refer to 50 known metabolites were identified by Chenomx software. Compared to healthy controls, six and five metabolites were significantly different in children with allergic rhinitis and asthma respectively. Among them, allergic rhinitis associated hypoxanthine, glutamate, and phenylalanine were positively changed with mite-allergen specific IgE levels, whereas asthma associated glutamine and arabinose were inversely changed in opposing directions. Clinically, a comprehensive analysis of functions studies related to these 8 metabolites could potentially provide further prevention and treatment in children with allergic rhinitis and asthma.

P-237 Endometrial Receptivity in Women with Dormant Genital Tuberculosis: A Molecular and Metabolomic Approach

PRESENTING AUTHOR: Koel Chaudhury, School of Medical Science and Technology, Indian Institute of Technology Kharagpur, India

CO-AUTHORS: Elavarasan Subramani, Baidyanath Chakravarty

Genital tuberculosis (GTB) in women is one of the common causes of infertility in emerging countries. The important problem in diagnosing and treating GTB is that it often exists in a dormant form without any clinical symptoms. There is a general consensus that GTB affects implantation due to ovarian and endometrial involvement. Behavior of the endometrium during the window of implantation in asymptomatic infertile women having dormant GTB is not well understood. The possible molecular association between dormant GTB and endometrial receptivity is, therefore, explored. Reduced levels of endometrial receptivity markers suggest impairment in the receptive status of the endometrium in dormant GTB women. Also, alterations in endometrial thickness, blood flow parameters and vascular endothelial growth factor indicate poor angiogenesis during implantation window. Compromised LIF-STAT3 signaling pathway could possibly be responsible for repeated implantation failure. Metabolomic studies indicate a clear metabolic differentiation in the metabolic profile of endometrium and serum between women with dormant GTB and controls. The significantly altered endometrial tissue and serum metabolites could be largely related to energy metabolism and amino acid biosynthesis. This underlines the metabolic influence of tubercle infection, even in its dormant form. Expression of several serum metabolites followed a similar pattern as that observed in the endometrium of dormant GTB women. The potential of these metabolites as putative diagnostic markers of dormant GTB seems promising.
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P-238

Study of fetal pig maturity in relation with neonatal survival using a multi-fluids metabolomic approach

PRESENTING AUTHOR: Laurence Liaubet, INRA-GenPhySE, France

CO-AUTHORS: Gaëlle Lefort, Nathalie Vialaneix, Hélène Quesnel, Marie-Christine Père, Yvon Billon, Laurianne Canario, Nathalie Iannuccelli, Cécile Canlet, Alain Paris, Rémi Servien, Laurence Liaubet

Selection for prolificacy and lean growth in swine has been associated with a substantial increase in piglet mortality. The first 24-48 hours after birth represent the most critical period. A major determinant for early survival is piglet maturity at birth, which strongly relies on the process of maturation during the last month of gestation. The objective of the current study was to compare progeny from Large White and Meishan pigs, which differ for piglet survival and vitality, to describe the metabolic status of the piglets. Thirty-nine sows were anesthetized at 90 or 110 days after conception (average gestation term: 114 days). Their fetuses (on average 15.7 per sow) were quickly obtained by caesarean section and this study focuses on the plasmatic, the urinary and the amniotic liquid metabolomes (1H-NMR) of 507 to 604 fetuses. Raw 1D Bruker spectral data files were treated with ASICS, an R package available on Bioconductor. ASICS allowed the direct identification and quantification of about 90 metabolites from a library of 190 metabolites, with about 60 metabolites per fluid, among which 39 were in common in the three fluids. A multivariate analysis and a mixed model were applied to the quantification results to explain the differences between the gestational stages and the breeds. The results provide new insights in the biological pathways involved in piglet maturation. Grants: ANR ANR-09-GENM005 (PORCINET) and ANR-16-CONV-0004 (#DigitAg), INRA (divisions GA, SA, MIA).

P-239

Neonatal hair profiling of monochorionic twins can be implicated to reflect a longitudinal metabolic phenotype of compromised fetal growth rate and abnormal umbilical artery flow

PRESENTING AUTHOR: Ting-Li Han, First affiliated Hopstial of Chonqing Medical University, China

CO-AUTHORS: Jing Yang, Yuan Wei, Yang Yang, Zhang Hua, Yangyu Zhao

Selective intrauterine fetal growth restriction (sIUGR) in monochronic diamniotic twins, especially types 2&3 with abnormal umbilical artery Doppler, results in increased risks of fetal/perinatal mortality and postnatal disability. Hair metabolome profiles of neonates were characterized to reflect the long-term pathophysiological changes across different clinical forms of sIUGR twins. Hair samples were collected at delivery from 10 pairs of type 1 sIUGR twins, 8 pairs of types 2&3 sIUGR twins, and 11 pairs of uncomplicated twins. The hair metabolome was characterized by gas chromatography-mass spectrometry. PLS-DA analysis and ROC curves were applied to screen for metabolites between/within twin pairs in different sIUGR and uncomplicated twin subgroups. A generalized estimating equation (GEE) model was implemented to correlate hair metabolites to birthweight and fetal growth rate within/between sIUGR co-twins. Metabolic pathway analysis and metabolic network reconstructions were performed using the KEGG database. Our results demonstrated that the hair metabolite profiles of different sIUGR subclinical forms were associated with the averaged fetal growth rate after 28 weeks of gestation but not birthweight. The hair profiles were capable of discriminating type2&3 sIUGR twins from uncomplicated twins. In particular, the metabolites 2-aminobutyric acid, cysteine, alanine, proline, valine, and tyrosine areas under the ROC curve were above 0.9. The metabolic pathway analysis highlighted the associations of sIUGR twins with abnormal umbilical artery flow including nutrient depletion pathway, glutathione metabolism, and attenuated nerve development. This study offers novel insight into evaluating the severity of intrauterine ischemia and hypoxia for T2&3 sIUGR twins, through the neonatal hair metabolome.

P-240

Early phenotypic alterations of newborns suffering from Hypoxic-Ischemic Encephalopathy with pathologic cerebral magnetic resonance imaging

PRESENTING AUTHOR: Julia Kuligowski, Health Research Institute La Fe, Spain

CO-AUTHORS: José David Piñeiro-Ramos, Antonio Núñez-Ramiro, Anna Parra-Llorca, Juan Martínez-Rodilla, Roberto Llorens, Ángel Sánchez-Ilana, Guillermo Quintás, Máximo Vento

Hypoxic-ischemic encephalopathy (HIE) the one of the major causes of neurodevelopmental impairment in the pediatric age. Therapeutic hypothermia (TH) initiated within 6 h of birth is now established as the standard of care for infants with moderate and severe HIE. TH has significantly reduced mortality and improved neurological outcome in survivors. However, no objective bedside test is available for an accurate and early diagnosis of HIE. This study elucidates early metabolic changes in newborns with HIE in order to select potential disease specific target biomarkers capable of predicting long-term adverse outcomes. We analyzed changes in the plasma metabolome employing gas-chromatography-mass spectrometry (GC-MS) and liquid chromatography-time-of-flight MS (LC-TOFMS) in a cohort of 55 asphyxiated infants who evolved to moderate/severe HIE. Hypoxia-ischemia induced brain injury was assessed employing magnetic resonance imaging (MRI) between days 4 and 8 post-partum. This study describes for the first time the evolution of the plasma metabolome of newborn infants with HIE covering the time period between birth and completion of TH. Results obtained allowed a critical assessment of the usefulness of lactate, pyruvate, and pyruvate/lactate for early outcome prediction. Besides, metabolomic profiling identified a dynamic perturbation of eleven metabolic pathways, including amino acid and purine metabolism, and the steroid hormone biosynthesis, in newborns with pathologic MRI outcomes. Future work will focus on the specific study of the steroid hormone biosynthesis pathway, which remained altered during the whole study period, to explore its potentials for biomarker discovery or adjuvant therapies to be combined with TH.
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P-241 Evaluation of Lipid Profile Differences between Neonatal and Adult Serum

PRESENTING AUTHOR: Richard Beger, NCTR, United States

CO-AUTHORS: Mustafa Akkoyunlu, Jiyeon Yang, Jinchun Sun

Introduction: While lipid profiles in terms of lipid classes including total cholesterol, triglycerides, high-density lipoprotein cholesterol and low-density lipoprotein cholesterol have been measured in newborn infants, little work has been conducted to show the lipid levels as lipid species in neonatal serum. Methods: Adult pooled sera (AS) were collected from 2 adult mice (8 to 12-week-old) for each replicate (n=5 replicates). Neonatal pooled sera (NS) were collected from 8-10 mice (1-week-old) for each replicate (n=5). An aliquot of 50 µL of the reconstituted sample was directly infused into a QTRAP 5500 MS with SelexION™ technology at a flow rate of 7 µL/min for targeted lipid profiling. Results: Lipidomics analysis results showed that total lipid amount (13 classes of lipids including neutral and polar lipids) present in NS was lower vs AS or in HINS vs HIAS, which is consistent with previous studies on newborns. Lipid classes of CE, DAG, LPC, LPE and TAG were significantly lower in NS vs AS; while lipid classes of PC, PE and SM were significantly higher in NS vs AS. The top fatty acid chain lipids differentiated NS from AS were the most abundant TAGs including (FA 16:0), (FA 16:1), (FA 18:1) and (FA 18:2). Interestingly, NS and HINS serum had higher levels of all lipid classes containing arachidonic acid (signaling molecule and a key inflammatory intermediate) chain vs AS and HIAS. Novel Aspects: Lipidomics analysis was conducted to evaluate the lipidome differences between neonatal and adult mouse sera at lipid species level.

P-242 LC-MS Based Targeted and Untargeted Metabolomics for the Identification of Potential Biomarkers in Plasma From Pediatrics With Chronic Kidney Disease

PRESENTING AUTHOR: Sandra Benito, University of the Basque Country, Spain

CO-AUTHORS: Nora Unceta, Alicia Sánchez-Ortega, M. Aránzazu Goicolea, Ramón J. Barrio

The assessment of chronic kidney disease (CKD) is carried out using glomerular filtration rate (GFR), which is calculated from serum creatinine concentration. However, serum creatinine concentration changes according to several factors, thus endangering CKD diagnosis, especially in early stages of the disease. In addition, pediatric CKD is related to malnutrition, development, metabolic and cardiovascular complications, leading to a 30 times higher mortality rate in paediatrics suffering from advanced CKD in comparison with their healthy counterparts. Regarding that the discovery of new biomarkers might allow earlier diagnosis as well as a better response, comprehensive research has been performed in plasma samples from paediatrics with CKD. Following targeted and untargeted metabolomics approaches 7 potential biomarkers have been found which could be useful for pediatric CKD by means of comparison of metabolic profiles: citrulline, S-adenosylmethionine, symmetric dimethylarginine, cis-4-decenoylcarnitine, n-butyrylcarnitine, bilirubin and sphingosine-1-phosphate. The use of these metabolites in addition to creatinine enables better differentiation of paediatrics with CKD and control paediatrics in comparison with the use of creatinine alone. Afterwards, these 7 biomarkers in addition to the classical biomarker creatinine have been gathered together into a routine analytical method. Aimed at evaluating the usefulness of this routine analysis method, a CKD pediatric population have been analyzed as well as a healthy pediatric population, being able to differentiate between them by means of data analysis.

P-243 Correlating plasma and liver metabolic profiles: an application to the study of organ maturation

PRESENTING AUTHOR: Oskar González, University of the Basque Country, Spain

CO-AUTHORS: Oihane E. Albóniga, Rosa M. Alonso, Yun Xu, Royston Goodacre

Liver plays a main role in drug metabolism, so studying the grade of maturation of this organ would help to develop more appropriate dosing regimens for paediatric population. Nevertheless, considering the invasive nature of liver analyses there are obvious ethical boundaries. In this work, we investigated the suitability of plasma as an alternative matrix to evaluate the biological age of liver. With this aim, we studied the correlation of plasma and liver metabolomic profiles obtained by UHPLC-TOF-MS for piglets of different ages. By means of Pearson correlation analysis we observed that 360 and 1784 pairs of features were significantly correlated in positive and negative ionization mode, respectively. Procrustes transformation was applied in order to assess the similarity of the clustering resulting from the data obtained from the two matrices. The dissimilarity values were low both for ESI+ (0.3753) and for ESI- (0.3673) and, hence, liver and plasma are expected to provide similar discriminatory information. Furthermore, Multiblock Principal Component analysis demonstrated to be a very suitable tool to combine the data obtained from both matrices and to better understand the clustering according to three groups: newborns, neonates and infant piglets. Considering all these results, plasma proved to be a matrix of great interest to provide insight into the maturation grade of liver. This could lead to less invasive procedures to study drug metabolism and it could be an important step forward towards an accurate dosing in paediatric population.
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**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

**POSTER SESSIONS 3 AND 4 – Wednesday and Thursday –** all even number posters will be on display.

*AWARD WINNERS

BIOMEDICAL

P-244 LC-MS based metabolomics: Biomarkers of organ maturation in paediatrics

**PRESENTING AUTHOR:** Oihane Elena Albóniga, Analytical Chemistry Department, Faculty of Science and Technology, University of the Basque Country, Spain

**CO-AUTHORS:** Oskar González, María E. Blanco, Rosa M. Alonso, Yun Xu, Royston Goodacre

Knowledge about drug metabolism in paediatric population is limited mainly due to ethical issues, scarce number of clinical trials and reduced sample size of the few paediatric studies available. In consequence, drug administration in children is often based on algorithms from allometric models optimised for adults, without considering the maturation state of key organs, such as liver, which during childhood are still in their development process. The invasiveness of liver sample collection requires the use of animal models, such as piglets, to study the paediatric population. In this study, metabolomic profiles of liver tissue, obtained from 36 piglets of different ages, were analysed by an HPLC-Q-TOF-MS system in positive and negative electrospray ionization (ESI) modes. The multivariate analysis by Principal Component Analysis (PCA) showed a perfect clustering of the three groups of piglets (newborns, neonates and infants). By univariate analysis, 91 features (m/z-tr) at ESI+ and 48 features at ESI- differentiating the three groups of piglets were found. The HPLC-MS/MS analysis of these significant features and their comparison with Metlin, HMDB, mzCloud and MyCompoundID databases allowed the identification of 13 acylcarnitines, such as acetylcarnitine, butyryl-L-carnitine, 2-methylbutyrol-carnitine or palmitoyl-L-carnitine, among others, suggesting the involvement of the fatty acids metabolism in the liver maturation state. These results provided an important contribution in paediatrics and demonstrated that metabolomics is a useful tool in this area.

P-245 Metabolomic prediction of adverse pregnancy outcomes

**PRESENTING AUTHOR:** Nancy McBride, MRC IEU, United Kingdom

**CO-AUTHORS:** Paul Yousefi, Matthew Suderman, Caroline Relton, Deborah Lawlor

Pregnancy-related disorders, such as gestational diabetes (GD), hypertensive disorders of pregnancy (HDP), small for gestational age (SGA), large for gestational age (LGA) and preterm birth (PTB), can have long-lasting adverse effects on maternal and offspring health and are a significant public health concern. Current risk factors rely on clinical predictors such as smoking, age, parity and body mass index (BMI). However, these are poor predictors. Metabolites may reflect dynamic changes in metabolic pathways of women with pregnancy-related disorders. We aim to see whether we can use metabolomics to improve prediction of adverse pregnancy outcomes. We used the Born in Bradford multi-ethnic cohort and the UK Pregnancies Better Eating and Activity Trial randomised control trial of obese pregnant women. Both have nuclear magnetic resonance (NMR) metabolite data from pregnant women at 24-28 weeks gestation. We used 159 NMR-derived metabolite measures and information on clinical predictors from 9,312 women to generate prediction models. Penalised regression was used to create predictive models with an elastic net penalty for HDP, GD, SGA and LGA from 90% of the cohort. Out-of-sample performance was conducted in a randomly allocated subset of 10% of observations that were withheld during training using receiver operating characteristic curves. We found that a model combining the metabolites and existing clinical predictors had very good discrimination for GD (AUC 0.85), good for HDP (0.76) and SGA (0.73), poorer for LGA (AUC 0.69). We will now compare this to the predictive purpose of the mass spectrometry (MS) metabolite platform.

P-246 Pre-symptomatic biomarker for preeclampsia in mid-trimester

**PRESENTING AUTHOR:** Yujin Kang, Kookmin University, South Korea

**CO-AUTHORS:** Seung Mi Lee, Joong Shin Park, Do Yup Lee

Preeclampsia (PE) is a hypertensive disease associated with pregnancy that has not yet been clearly elucidated. The disorder can affect fetus and pregnant women, which in turn necessitates special care. In particular, it may cause complications such as fetal growth retardation, premature birth, renal dysfunction, and chronic hypertension in pregnant women. Sometimes, preeclampsia is very similar to symptoms of normal pregnancy such as weight gain, swelling and headache and some mothers have no symptoms at all. Accordingly, it is very important to properly diagnose preeclampsia through periodic antenatal screening and early diagnosis. In our current study, metabolic profiling of maternal plasma samples in mid-trimester was performed using gas chromatography-time of flight mass spectrometry (GC-TOF MS) and liquid chromatography orbitrap mass spectrometry (LC-Orbitrap MS). A total of 330 metabolites were semi-quantitatively identified. Primarily, partial least squares-discriminant analysis (PLS-DA) demonstrated a clear discrimination between pregnant women with preeclampsia and healthy controls. Subsequent binary logistic regression analysis constructed a potential biomarker recomposite with 3 metabolites. The predictive model successfully discriminated preeclampsia group in the mid-trimester (AUC = 0.801).
**POSTER SESSIONS 1 AND 2 – Monday and Tuesday –** all odd number posters will be on display.

**POSTER SESSIONS 3 AND 4 – Wednesday and Thursday –** all even number posters will be on display.

*AWARD WINNERS

BIOMEDICAL

P-247 Investigation of Early Diagnosis of Neonatal Sepsis by Untargeted Metabolomics Approach

PRESENTING AUTHOR: Giuseppe Giordano, Padova University, Women's and Children's Health Department, Italy

CO-AUTHORS: Veronica Mardegan, Antonella Gucciardi, Matteo Stocchero, Paola Pirillo, Gabriele Poloniato, Mauro Naturale, Eugenio Baraldi.

Neonatal sepsis is a complex infection-induced systemic inflammatory response syndrome and it is a main cause of mortality and neurologic sequelae in newborns. An early and accurate detection of sepsis is mandatory in neonates, since the clinical course of the infectious process can be fulminant, leading to septic shock and death within hours after the first clinical symptoms. To date, there is still no reliable biochemical marker of neonatal sepsis. The aim was to compare the metabolic profile of urine collected within 24 hours of birth between preterm neonates affected by early onset sepsis (EOS) and healthy preterm infants, searching for a specific metabolic profile enabling the identification of preterm newborns prone to develop EOS. Infants who developed a septic episode within 72 hours of birth were enrolled as cases (9 cases and 10 controls who did not developed a septic episode). The urine samples underwent untargeted metabolomic analysis using LC-MS. The data were analyzed by multivariate and univariate data analysis tools. For each data set, a multivariate biomarker showing area under the ROC curve calculated by cross-validation greater than 0.80 was built. The annotated variables were submitted to over-representation pathway analysis that highlighted the perturbation of 6 metabolic pathways. Neonates with EOS showed a specific metabolic profile compared to those of newborns not affected by sepsis at the onset of infection. Results of this research support the effectiveness of metabolomics in exploring biochemical pathways of neonatal sepsis, potentially providing novel putative biomarkers for early diagnosis.

P-248 Metabolomic barometer of gestational and postpartum weight in overweight pregnant women

PRESENTING AUTHOR: Chung-Ho Lau, Imperial College London, United Kingdom

CO-AUTHORS: Victoria Taylor-Bateman, Panagiotis A Vorkas, Gonçalo Graça, Elena Chekmeneva, Timothy M.D. Ebbels, Linda Van Horn, Queenie Chan, Elaine Holmes

Background: Obesity amongst women of reproductive age is increasingly common in developed nations and has been shown to adversely affect childhood cardio-metabolic, respiratory and cognitive-related health outcomes in offspring. Metabolomic signatures of obesity are readily captured in biofluids samples and could potentially provide a molecular barometer for monitoring excessive gestational weight gain (GWG) and postpartum weight loss (WL) in overweight/obese pregnant women. Methods: Urine and blood plasma samples were collected from 114 overweight or obese ethnically diverse pregnant women from Chicago (USA), as part of a randomised diet and lifestyle intervention trial (Maternal Offspring Metabolics Family Intervention Trial; www.clinicaltrials.gov NCT01631747). Blood plasma lipids and urine samples at 15 weeks, 35 weeks of gestation, and at 1 year postpartum were respectively analysed by LC-MS and NMR. Results: Urinary 4-deoxyerythronic acid was found positively correlated to body mass index (BMI) and a broad spectrum of alterations in levels of blood plasma phosphatidylcholines, lysophospholipids, and sphingomyelins were associated with BMI, GWG, and WL. Specifically, several plasmanyl-/plasmenyl-phospholipids were negatively associated with GWG, and lysophosphatidylcholines (including LPC 20:4) were positively associated with WL. Multiple lipids with apparent 18:2 fatty-acid chains were significantly associated with GWG/ WL, suggesting linoleic acid, an essential nutrient, may play an important role in prenatal and postpartum weight management. Conclusions: Maternal obesity-related parameters are associated with urine and plasma metabolomic profiles, which could be further exploited to evaluate the beneficial effect of diet and lifestyle intervention in pregnancy.

P-249

A population-based resource for intergenerational metabolomics analyses in pregnant women and their children:
the Generation R Study

PRESENTING AUTHOR: Ellis Voerman, Erasmus University Medical Center, Netherlands

CO-AUTHORS: Vincent W.V. Jaddoe, Romy Gaillard

Adverse exposures during fetal and early postnatal life may predispose children to cardio-metabolic disease in later life. Early-life developmental adaptations of metabolic pathways may underlie these associations. Metabolomics may serve a tool to disentangle these potential metabolic adaptations. In a prospective population-based cohort study among pregnant women and their children from Rotterdam, the Netherlands, we used a targeted metabolomics approach to measure metabolite concentrations in maternal blood in early-pregnancy (n=825), cord blood at birth (n=932), and child's blood at age 10 (n=508). We used liquid chromatography mass spectrometry (LC-MS/MS) to measure blood concentrations of amino acids, acyl-carnitines, polar lipids (including diacyl-phosphatidylcholines, acyl-alkyl-phosphatidylcholines, sphingomyelines, acyl-lysophosphatidylcholines and alkyl-lysophosphatidylcholines), and non-esterified fatty acids. Preliminary exploratory analyses showed weak correlations between maternal concentrations of individual metabolites and concentrations of these metabolites in their children at birth (highest Spearman's Rho: 0.36) and at the age of 10 years (highest Spearman's Rho: 0.24), respectively. Child's metabolite concentrations at birth and at 10 years of age were also weakly correlated (highest Spearman's Rho: 0.36). Whether individual metabolite concentrations or metabolite patterns are related to maternal and child adverse exposures and health outcomes is subject of future analyses. These data are an important population-based resource for metabolomics analyses in pregnant women and their children to address the early origins of health and disease.