**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

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-353

Elucidating markers of strawberry consumption in smokers and non-smokers using metabolomics

PRESENTING AUTHOR: Jessica Cooperstone, The Ohio State University, United States

CO-AUTHORS: Matthew D. Teegarden, Jennifer H. Ahn-Jarvis, Thomas J. Knobloch, Christopher M. Weghorst, Yael Vodovotz, Steven J. Schwartz, Jessica L. Cooperstone

**Background:** Promising results from pre-clinical and clinical research support a potential role for berries and their associated phytochemicals in the prevention of oral cancer. Strawberries are a significant dietary source of berry phytochemicals. Our group recently conducted a randomized, cross-over, placebo-controlled clinical study, in which subjects consumed strawberries powder (24 g/day) and a placebo product, each for 7 days. **Objective/Hypothesis:** Our objective was to investigate key markers of strawberry exposure in the urine of healthy individuals and individuals at high risk for oral cancer (smokers) using metabolomics. **Methods:** Urine samples from 9 smokers and 10 non-smokers following placebo and strawberry interventions were volumetrically normalized according to osmolality and chemically profiled using UHPLC-QTOF-MS. Data were modeled using multilevel, multivariate analyses to capitalize on the cross-over design of the study. **Results:** A total of 41 markers were found, 26 of which were elevated following strawberry treatment and 15 following the placebo. Using ion fragmentation and comparison to authentic and synthesized standards, we identified metabolites elevated with the strawberry intervention including urolithin A (and phase II metabolites), dihydroxybenzoic acids, and phase II metabolites of strawberry aroma compounds. Markers of placebo consumption included phase II metabolites of flavorants and a metabolite of colorants included in the formulation. **Conclusions:** Distinct metabolic profiles pertaining to strawberry and placebo treatment arms were observed and provide targets for validation. These findings will inform future clinical trials with strawberry products with regard to potential markers of exposure and implications of study design on metabolomic profiles.

P-354

Identifying Natural Compounds as Inhibitors of Quorum Sensing Transcriptional Regulator (SdiA) of Klebsiella pneumoniae through Modelling, Docking and Molecular Dynamics Simulation

PRESENTING AUTHOR: Mohamed Alajmi, King Saud University, Saudi Arabia

CO-AUTHORS: MD Tabish Rehman

Bacteria can communicate amongst themselves through cell-cell signalling system; quorum sensing (QS). Gram-negative bacteria detect N-acylhomoserine lactones (AHLs) signalling molecules through LuxR transcriptional regulators. Certain bacteria do not produce signalling molecules but can detect the signals produced by other bacterial species through SdiA (suppressor of cell division inhibition). SdiA is a homolog of LuxR transcription regulator and thus regulates AHL-mediated transcription of various virulent genes. In the present work, we employed a computational biology approach to identify novel inhibitors of SdiA by screening natural compounds. First, we modelled the structure of SdiA using Modeller using 4LFU as template (64.6% sequence identity and 100% coverage). ProCheck, Verify3D, Ramachandran plot scores (>99.5% residues in the favourable and allowed regions) and ProSA-Web (Z-score=-8.45) have been employed to ascertain the good quality of the generated SdiA model. RMSD between SdiA model and 4LFU template was estimated to be 0.21 Å. The secondary structural contents of SdiA model was predicted using PDBsum. Further, CASTp identified only one pocket as the most probable binding site of SdiA (area=523.083 Å2 and volume=351.044 Å3). Autodock Vina was employed to perform high throughput virtual screening (HTVS) on a library of natural compounds available at InterBioScreen Ltd. The potential inhibitors were again docked on SdiA using flexible docking in Autodock. Further, molecular mechanics-general born surface area (MM-GBSA) was used to evaluate the effect of solvent on protein-ligand complex stability. Finally, molecular dynamics simulation was performed on the two best inhibitors to confirm the potential of the identified inhibitors against SdiA

P-355

Unraveling dynamic metabolomes underlying different maturation stages of berries harvested from Panax ginseng

PRESENTING AUTHOR: Mee Youn Lee, Konkuk University, South Korea

CO-AUTHORS: Mee Youn Lee, Han Sol Seo, Digar Singh, Sang Jun Lee, Choong Hwan Lee

Ginseng berries (GBs) show temporal metabolic variations among different maturation stages, determining their organoleptic and functional properties. We analyzed metabolic variations concomitant to five different maturation stages of GBs including immature green (IG), mature green (MG), partially red (PR), fully red (FR), and overmature red (OR) using mass spectrometry (MS)-based metabolomic profiling and multivariate analyses. The PLS-DA score plot based on GC-MS datasets highlighted metabolic disparity between preharvest (IG and MG) and harvest/postharvest (PR, FR, and OR) GB extracts along PLS1 (34.9%) with MG distinctly segregated across PLS2 (18.2%). Forty-three significantly discriminant primary metabolites were identified encompassing five developmental stages (VIP > 1.0, p < 0.05). Among them, most amino acids, organic acids, 5-C sugars, ethanolamines, purines, and palmitic acid were detected in preharvest GB extracts, whereas 6-C sugars, phenolic acid, and oleamide levels were distinctly higher during later maturation stages. Similarly, the partial least squares discriminant analysis based on LC-MS datasets displayed preharvest and harvest/postharvest stages clustered across PLS1 (11.1%); however, MG and PR were separated from IG, FR, and OR along PLS2 (5.6 %). Overall, 24 secondary metabolites were observed significantly discriminant (VIP > 1.0, p < 0.05), with most displaying higher relative abundance during preharvest stages excluding ginsenosides Rg1 and Re. Furthermore, we observed strong positive correlations between total flavonoid and phenolic metabolite contents in GB extracts and antioxidant activity. Comprehending the dynamic metabolic variations associated with GB maturation stages rationalize their optimal harvest time per se the related agroeconomic traits.
**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

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

**P-356**

Development of biomarkers of macronutrient intake by integrated analysis of metabolome data in both human samples and food compositions

PRESENTING AUTHOR: Yoshihito Fujita, Department of Diabetes, Endocrinology and Nutrition, Kyoto University, Japan

CO-AUTHORS: Nozomu Sakurai, Kaori Ikeda, Nozomi Isomura, Fumika Mano, Futoshi Furuya, Daisuke Shibata, Takeshi Ara, Hiroto Minamino, Tsuyoshi Goto, Yasuki Matsumura, Teruo Kawada, Nobuya Inagaki

Environmental factors such as excessive dietary intake have a critical role in the pathogenesis of life style-related diseases including diabetes and obesity; nutrition education contributes to the prevention of these diseases. Novel biomarkers of dietary intake are required to apply nutrition education effectively. To identify biomarkers of macronutrient intake, we have used a metabolomics approach. As a pilot study, we performed metabolomic analysis using human samples after test meal (Terumeal(R)). After intake of the test meal, increases and/or decreases in peaks detected by LC/MS analysis were distinguished in human plasma and urine samples. Correlation analysis of the metabolomic data of the test meal itself and the human samples revealed that many components derived from Terumeal(R) could be detected, and that the values were altered time-dependently after meal in human samples. Based on findings from this study, we established a protocol for the development of biomarkers of macronutrient intake. Next, we performed metabolomic analysis using human samples after white rice or white bread intake as representatives of carbohydrate, a component of macronutrient. Samples in 8 healthy volunteers were taken 7 times over a period of 24 hours. Increases in peaks by LC/MS analysis were detected after bread intake but were not detected after rice intake in human plasma samples. Components derived from bread or its derivatives modified in the body were detected in human plasma samples after bread intake. Thus, integrated analysis of metabolome data both in human samples and various food compositions revealed candidates for biomarkers of macronutrient intake.

**P-357**

Using different untargeted GC-MS trapping techniques reveals the comprehensiveness of volatile compounds in flavouring ingredients

PRESENTING AUTHOR: Carmen Diez Simon, Wageningen UR, Netherlands

CO-AUTHORS: Roland Mumm, Robert D. Hall

Metabolomics has become a valuable data-driven, holistic approach that is able to generate novel hypotheses, identify novel biomarkers and elucidate underlying modes of action. Whereas metabolomics has been applied to diverse plant materials, applications in processed foods is still in its infancy, especially in relation to flavour aspects. Nevertheless, recent studies have shown that metabolomics has great potential to identify sensory-relevant compounds. From this, it became evident that existing technologies have huge potential but need further development to obtain robust analytical data for complex food products. We have developed methods for the untargeted analysis of volatiles from process flavours. The challenge lies in developing optimal headspace trapping techniques that give the most comprehensive picture together with an efficient and robust detection of volatiles. A number of volatile trapping techniques including solid-phase microextraction (SPME), stir-bar sorptive extraction (SBSE), and dynamic headspace (DHS) were compared for their relative values and robustness for the analysis of the volatiles present. Results showed that the techniques differ in their efficiency in trapping of different classes of volatile compounds suggesting a need to tailor the volatile extraction technique depending on the specific composition of the process flavour. Methodologies have been developed which: deliver robust data on sensory-relevant compounds; allow for identifying unknown (Maillard-related) compounds that contribute to the prediction of sensory attributes and; give us deeper insights into the most relevant chemical reactions. Acknowledgements: The authors gratefully acknowledge funding from NWO, Unilever and DSM for the work presented.

**P-358**

Urinary Metabolomic Profiling to Identify Biomarkers of Carrot Intake

PRESENTING AUTHOR: Siva Charan Sri Harsha Pedapati, University College Dublin, Ireland

CO-AUTHORS: Roshaida Abdul Wahab, Lorraine Brennan

Metabolomics has emerged as a key technology to identify novel food-intake biomarkers in a range of foods. The objective of this research was to identify urinary biomarkers of carrot intake using an untargeted metabolomics approach. In this context, a randomized cross-over acute intervention study was conducted on eleven participants who consumed carrots (Test food) and turnips (control food) in random order. Urine samples were collected postprandially and analysed by UPLC-QTOF-MS. The data obtained was reduced to 3148 features in positive mode and 3015 features in negative mode applying filtering procedures such as data alignment and sample occurrence frequency. Robust PLS-DA models were obtained when comparing fasting samples with 12h (R2X=0.725, Q2=0.331) time point post-consumption of carrots. The variable importance for the projection (VIP) list was created from PLS-DA plots with scores ≥1.5 considered as discriminant features between the two time points. Time series plots revealed 29 features (15 in positive mode and 14 in -ve mode) with a time response following carrots consumption with 11 features demonstrating a differential time course compared to the control food. Ferulic acid 4-sulphate (m/z 273.0074), Pyrocatechol sulphate (m/z 188.9864), N-Feruloylglycine (m/z 251.0721), L-Tyrosine (m/z 182.0802) were identified as metabolites reflective of carrot intake using databases such as METLIN, HMDB and confirmed by LC-MS/MS. Future work will focus on confirmation of the remaining discriminating features and examine dose response relationship to ascertain biomarker validity. Overall, the untargeted metabolomics approach developed proved to be successful in the identification of urinary markers of carrot intake.
**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

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-359 Australian native plants and their application in food

PRESENTING AUTHOR: Sara Ghorbani Gorji, School of Agriculture and Food Sciences, The University of Queensland, Australia

CO-AUTHORS: Sara Ghorbani Gorji, Siyuan Chen, Zhenzhen Li, Melissa Fitzgerald

Australia has a diverse native flora, that contains plants that have been used as food or medicine by Indigenous people for many thousands of years. However, to date Western knowledge about the properties of these plants is limited, and where information is available, it is clear that these plants offer unique properties in terms of flavour and nutritional value. Recently, there have been many reports about the nutritional value of Indian turmeric (Cucurma longa). In Northern Queensland, a native Australian turmeric (Cucurma australasica) is found, and it is reported that Indigenous people roast and consume the rhizomes. In this study we have explored the rhizomes of Australian native turmeric for the presence of phytochemicals important to human health, unique flavours, and nutritional properties, and compared these with the traditionally consumed Indian turmeric rhizomes. With the aid of metabolomics, the phytochemical, volatile compounds and minerals of Australian and Indian turmeric were studied. Various antioxidant assays were used to compare the antioxidant activity. The potential of using Australian turmeric in niche products such as turmeric latte or confectionary was also studied. Sensory analysis provided new insights about the application of Australian turmeric in the market. Our results showed that the Australian turmeric has comparable nutritional value to Indian Turmeric and furthermore, consumers preferred Australian turmeric over Indian turmeric in a formulation for turmeric latte. This research helps to understand potential applications of Australian native turmeric, ultimately increasing its economic value, and offering a native competitor to the now highly sought Indian turmeric.

P-360 UPLC-Orbitrap-MS and GC-TOF-MS based Metabolomics Analysis on the differentiation of Red Ginseng of Two Different Geographical Origins

PRESENTING AUTHOR: Ailsa Chui-Ying YUEN, The Hong Kong Polytechnic University, Hong Kong

CO-AUTHORS: Siu-Wai WAN, Tung-Ting SHAM, Chi-On CHAN, Daniel Kam-Wah MOK

Red ginseng (also called Hongshen in Chinese) is the steamed dried root and rhizome of Panax ginseng C.A. Mey and is a famous Chinese medicine for tonifying qi. The northern part of China and South Korea are two main cultivation areas of red ginseng, and ginseng produced in these locations differs in qualities and thus in their market prices too. Previous studies have used hydrogen, carbon and nitrogen isotopic ratios to differentiate ginseng collected from these two locations. However, this technique requires specialized instrument and does not provide information about the quality of red ginseng. Meanwhile, the analysis of ginseng mainly focuses on its ginsenoside content, however, it could be affected by various factors such as the steaming process, age and the part of the ginseng (main or lateral root). In this study, red ginseng samples collected from China and Korea were extracted using the modified Bligh and Dyer's liquid-liquid extraction method with subsequence analysis using GC-TOF-MS (aqueous layer) and UPLC-Orbitrap-MS (organic layer). Multivariate analysis was carried out to identify the key metabolites for differentiating between red ginseng from these two locations. Our results showed that carbodiimide, propionic acid, disaccharide (maltose, fructose, sucrose), ginsenoside Re, Rf, Notoginsenoside R2 could be tentatively identified as the metabolites to differentiate between ginseng from these two origins. The results suggested that the metabolomic profile of red ginseng could provide more comprehensive information to differentiate between Chinese and Korean red ginseng.

P-361 Metabolomics-based Approach for the Evaluation of Post-Harvest Quality of White Leg Shrimp (Litopenaeus vannamei) Cultured in Different Salinity

PRESENTING AUTHOR: Eiichiro Fukusaki, Osaka University, Japan

CO-AUTHORS: Sri Hayati, Julie Ekasari, Sastia P. Putri

Shrimp is one of the most important commodities in aquaculture industry and has important significance in global food supply. Salinity plays an essential role in shrimp aquaculture and is considered to affect post-harvest quality (physical and sensory attributes). However, there is no study on the correlation between salinity and metabolite profiles. The objective of this study is to investigate the effect of different salinity on physical and sensory attributes, and metabolome of white leg shrimp (Litopenaeus vannamei). As an initial analysis, shrimp was cultured in different salinity (10, 20, 30 ppt) for ten weeks under controlled condition and were subsequently analyzed for physical attributes (texture and color analysis), sensory analysis, and untargeted GC/MS analysis. The result for texture profile analysis showed that there is an increase trend in springiness and cohesiveness value along with increase of salinity level. Moreover, color measurement showed that ΔL*(lightness-darkness) and Δb*(yellow-blue) score showed a positive trend along with increase of salinity, and thus both of texture and color could be used as a reference for selected sensory attributes. GC/MS based metabolome data were subjected to Principle Component Analysis to clarify the important metabolites varying depending on salinity change to be several amino acids. The findings from this study provide an insight on the effect of different salinity towards metabolite composition of shrimp and can act as a useful feedback for shrimp aquaculture industry.
*AWARD WINNERS

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-362 Discovering the flavor of the fine flavor cocoa of Piura/Peru

PRESENTING AUTHOR: Madina Mansurova, Pontificia Universidad Catolica del Peru, Peru

CO-AUTHORS: Stephanie Michel, Alfredo J. Ibañez

Flavor and aroma are the most important characteristics of chocolate and is key to determining the final price that the customer is willing to pay for the product. Peru is one of the countries that produces and exports the finest cocoa beans (Fine Flavor Cocoa, or FCC), and the chocolate as a final product. In this project we have developed an analytical method based on mass spectrometry to identify the volatile compounds profile responsible for the fine flavor of the Peruvian chocolate from Piura. Furthermore, we have observed the development of those taste markers during the manufacturing of the chocolate from cocoa beans. Finally, we cross-correlated the results of our measurements with the quality control team of the FCC-based chocolate company to corroborate with a panel the taste markers that we have detected molecularly. In summary, our methodology allowed us to map chemical compound patterns that represent the flavor and aroma of the final product (chocolate), making it possible to differentiate it from other ordinary cocoa chocolates, as well as from other fine flavor chocolate producers with different cacao origin or different manufacturing. Hence, this approach can be applied to other chocolate producers, as well as other food products to define, analyze and quantify flavor and aroma compounds. This information would be of great help for those companies whose sale price depends on their organoleptic characteristics.

P-363 Dynamics of metabolic profiling in thirteen pepper cultivars during fruit ripening

PRESENTING AUTHOR: Jae Kwang Kim, Incheon National University, South Korea

CO-AUTHORS: Tae Jin Kim, Hyejin Hyeon, Nam Il Park, Tae Gyu Yi, Sang Un Park, Yongsoo Choi, Jae Kwang Kim

To the best of our knowledge, none of the studies have conducted a comprehensive profiling of the primary and secondary metabolites in various phenotypes of pepper in relation to the stage of fruit maturity, nor did they have examined the metabolic networks by linking the large-scale metabolite data with metabolic pathways. We performed metabolic profiling of phytochemicals, namely amino acids, organic acids, sugars, sugar alcohols, free phenolic acids, capsaicinoids, carotenoids, phytosterols, policosanols, tocopherols, volatiles, and fatty acids, in 13 pepper cultivars at different fruit maturity stages. Our results comprised 10,218 data points resulting from three repetitions of 131 metabolites obtained from the fruit samples. Next, the multivariate analysis (principal component analysis, PCA; Pearson's correlation analysis; hierarchical clustering analysis, HCA) and PathVisio 3.3.0 were used with these data to obtain a more comprehensive understanding of the relationships among the metabolites and pepper samples. In the mevalonic acid pathway, the content of phytosterols such as campesterol, beta-sitosterol, cholesterol, and stigmasterol were reduced in the fruit mature stage, thus confirming the negative correlation between carotenoids and tocopherols revealed by the PCA and HCA. A complex carbon and nitrogen metabolic network was modulated by fruit ripening. Correlation-based network analysis suggested that metabolism is substantially coordinated during the ripening.

P-364 Regression model-based metabolomics approach to propose new quality marker for specialty coffee

PRESENTING AUTHOR: Sastia P. Putri, Osaka University, Japan

CO-AUTHORS: Tomoya Irifune, Yusianto, Eiichiro Fukusaki

Specialty coffee is defined based on cupping scores of 10 attributes (fragrance/aroma, flavor, aftertaste, acidity, body, uniformity, balance, clean cup, sweetness, overall) in the protocol established by the Specialty Coffee Association of America. Despite the increasing demand of specialty coffee, there are very few research on the relationship between component profile and specialty coffee quality. Such study is important for further quality improvement of coffee. Here, a regression model-based metabolomics approach was carried out to propose quality marker of specialty coffee. A total of 97 hydrophilic low molecular weight compounds were detected in 10 specialty coffee samples by GC/MS-based component profiling. Prediction models for 7 attributes (fragrance/aroma, flavor, aftertaste, acidity, body, balance, overall) and final score were constructed by Orthogonal Projection to Latent Structure regression analysis and these prediction models showed high performance (R2>0.9, Q2>0.9). Subsequently, important compounds for each attribute were proposed based on Variable Influence on Projection (VIP). This study proposed galactinol as a new candidate of quality marker of specialty coffee. Galactinol was shown as the compound with the highest VIP in the prediction models of fragrance/aroma, flavor, aftertaste, acidity and overall score. Quantitation of galactinol in samples with lowest and highest score samples in the prediction model showed that there is a significant increase of galactinol concentration in which highest scoring sample (cupping score 86.75) showed 8 times increase of galactinol concentration compared to lowest scoring sample (cupping score 80.25). Identification of galactinol as a new quality marker would be useful for quality improvement of coffee.
*AWARD WINNERS

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-365 Metabolomics, an invaluable phenotyping tool in food science

PRESENTING AUTHOR: Melissa Flitzgerald, University of Queensland, Australia

CO-AUTHORS: Venea Dara Daygon, Sara Ghorbani Gorji, Crystal Concepcion, Lourdes Urban Andalete

The field of metabolomics developed over the years in chemistry laboratories. However, over the past few years, the tools have become more available for routine use for phenotyping. In addition, the stability and reproducibility of the tools enables the phenotyping datasets to be associated with other datasets collected with other omics data, such as lipidomics and genomics. This enables us to complete the picture from the gene, its expression into a protein and the phenotype produced by the action of that protein. As we develop new ideas to improve the flavour, taste and nutritional value of food, metabolomics and other omics platforms have provided invaluable data for transformational solutions to the food industry. Metabolomics has enabled the discovery of different genes to improve the aroma of rice, different ways to provide natural ingredients to prolong the shelf-life of food, knowledge of how lipids impact the sensory properties of rice, and different ways to define rancidity based on volatile compounds. We will report our discoveries for different foods, using mostly metabolomics, lipidomics and genomics platforms, and demonstrate the value of these discoveries to agriculture and food industries in Australia.

P-366 Discovery of plasma biomarkers in human samples that reflect carbohydrate intake

PRESENTING AUTHOR: Nozomi Isomura, Department of Diabetes, Endocrinology and Nutrition, Kyoto University, Japan

CO-AUTHORS: Yoshihito Fujita, Nozomu Sakurai, Kaori Ikeda, Fumika Mano, Futoshi Furuya, Daisuke Shibata, Takeshi Ara, Hiroto Minamino, Tsuyoshi Goto, Yasuki Matsumura, Teruo Kawada, Nobuya Inagaki

Excessive carbohydrate intake is known as a major risk factor of lifestyle-related diseases such as diabetes. Clinical application of biomarkers that reflect dietary intake is required for nutrition education. In this study, we screened for biomarkers reflecting carbohydrate intake using a metabolomics approach. One-week consecutive intake of white bread followed by similar intake of white rice for the same period was applied for 7 healthy volunteers. Plasma samples were collected before and after each intervention, and non-targeted analysis using LC/MS was performed. In partial least squares (PLS) analysis, we discriminated 4 groups, which revealed dynamic changes of metabolites reflecting white bread or rice intake. To further investigate the biomarkers reflecting white bread intake, we compared PLS analyses to compare samples after the periods of bread intake or rice intake. We screened for peaks in mass chromatogram that reflected white bread intake specifically. Next, to ascertain whether these metabolites changed time-dependently, we performed analysis using samples after single intake of white bread or white rice. Plasma samples were collected 7 times over a period of 24 hours, and we found metabolites identical to those after 1-week intake. It is likely that these include external metabolites derived from the food composition itself. Thus, we found candidates for biomarkers that reflect carbohydrate intake such as white bread.

P-367 Compositional assessment of genetically modified soybean containing thioredoxin-encoding gene

PRESENTING AUTHOR: Ye Jin Kim, Incheon national university, South Korea

CO-AUTHORS: Young Jin Park, Ju-Seok Seo, Jung-Ho Park, Chang-Gi Kim, Jae Kwang Kim, Sung-Dug Oh

The development and cultivation of genetically modified organism (GMO) has been increased continuously over recent years. As cosmetic materials, genetically modified (GM) soybean event CT-4025 was developed. This soybean contains gene that encode thioredoxin (TRX) gene for improving skin whitening function. In addition, the phosphinothricin-N-acetyltransferase gene was used as a selectable marker gene for glufosinate tolerance. However, commercialization of the GM soybean requires the safety assessment. Therefore, in this study, to assess compositional equivalence between GM soybeans (non-sprayed and sprayed with glufosinate) and non-transgenic soybeans, forty-six key nutrients (proximates, amino acids, fatty acids, isoflavones, vitamins and anti-nutrients) were analyzed. The soybeans were cultivated in 2017 at two representative regions (Ochang and Jeonju) located in the Republic of Korea. Statistical analysis was performed by using the linear model and principal component analysis (PCA). As a result, most of the analyzed components in the GM soybeans had non-significant differences compared with its non-transgenic soybean in the linear model. A result of the PCA model showed that there was a clear discrimination between two regions rather than the soybean genotypes. In a loading plot, the isoflavones such as malonyldaidzin, malonylglycitin, glycitin and genistin mainly contributed to separation between Ochang and Jeonju. Consequently, these comparisons supported that CT-4025 is compositionally equivalent to the non-transgenic soybeans. Furthermore, the results of this study could be a useful tool to investigate compositional similarities between GM and non-GM food.
*AWARD WINNERS

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-368

Differences in the metabolomic profile and microbial composition of soil supporting Burkea africana growth

PRESENTING AUTHOR: Lufuno Ethel Nemadodzi, University of South Africa, South Africa

CO-AUTHORS: Jacques Vervoort, Gerhard Prinsloo

Burkea africana (Wild syringa) is a leguminous tree widely used for medicinal purposes, with a distribution in Namibia, Nigeria, Ethiopia as well as South Africa. The trees are found and grow in clusters, on dry, acidic sandy soils impoverished from most nutrients with a particularly low phosphorus level. The study determined the factors responsible for successful growth in nature of the Burkea africana trees as all efforts for commercial production has been proven unsuccessful. An investigation was carried out to comprehend the metabolic profile, chemical composition and microbial composition of the soils where Burkea africana grows (Burkea soil) versus where it does not grow (non-Burkea soil). 1H-NMR metabolomic analysis showed different metabolites in the respective soils. Trehalose, betaine, choline-like and carnitine-like compound, were found to be in higher concentration in the Burkea soils, whereas, acetate, lactate and formate were found to be more prevalent in the non-Burkea soils. LC-MS analysis revealed the presence of numerous amino acids such as aspartic acid and glutamine to be higher in the Burkea soils. Deoxyribonucleic acid (DNA) was extracted from the soil and subsequently sequenced. A Basic Local Assignment Search Tool (BLAST) was used to analyze the microbial diversity (bacterial and fungal) and composition found in both soils, for a comprehensive understanding of the soil composition. Penicillum species was found to be prevalent in Burkea soils and was the main discriminant between the two soils. The variances in fungal composition suggests that species supremacy play a role in development of Burkea africana trees.

P-369

Profiling Glucosinolates Using LC-MS in Seeds of Camelina sativa L. Crantz

PRESENTING AUTHOR: Rong Zhou, Agriculture and Agri-Food Canada, Canada

CO-AUTHORS: Ning Xu, Kevin C. Falk, Margaret Y.Gruber

Camelina sativa L. Crantz, a member of Brassicaceae family, is heat and drought tolerant. Thus it is thought to require fewer agronomic inputs and is a good alternative crop for marginal lands. Its seed contains three glucosinolates: glucoarabin (9-(methylsulfinyl)nonylglucosinolate or GS9), glucocamelinin (10-(methylsulfinyl)decylglucosinolate or GS10) and 11-(methylsulfinyl)undecylglucosinolate or GS11). The structures of these glucosinolates are similar to that of glucoraphanin (4-(methylsulfinyl)-butylglucosinolate). The degradation product of glucoraphanin is thought to be an anti-cancer compound. Glucosinolates were extracted and converted to desulfoglucosinolates based on AOCS Official Method Ak 1-92 before analyzed by a Waters Ultra Performance Liquid Chromatography-Photodiode Array Detector-Tandem Mass Spectrometry (UPLC-PDA-TQD) equipped with a BEH Shield RP18 column (2.1 x 50 mm; 1.7 µm) and held the initial conditions (100% water at 0.8 mL/min) for 0.3 minutes and then a linear solvent gradient of 0% to 25% acetonitrile (v/v) over the next 6.7 min. The desulfoglucosinolates were quantified at 229 nm and identified by monitoring the characteristic loss of 162.2 mass units using MS/MS constant neutral loss scans. UPLC reduced the typical LC run time of 25-50 minutes for camelina desulfoglucosinolates to 10 minutes and consumed 60% less acetonitrile. The camelina accessions surveyed contained 21 - 31 µmol glucosinolates per gram of seed: GS9 (4.6-7.0 µmol/g seeds), GS10 (13.7-20.2 µmol/g seeds) and GS11 (2.5-3.6 µmol/g seeds). Additionally, we identified several putative minor glucosinolates including 8-(methylsulfinyl)octyl glucosinolate (GS8) based on its mass spectrum. Further work is needed to confirm the identity of the minor glucosinolates.

P-370

Bitter Melon Triterpenoids Regulate Human Gut Microbiome and Potentially Benefit Metabolism Condition

PRESENTING AUTHOR: Jia Liu, Shanghai Institute of Materia Medica, China

Bitter melon, a widely cultivated vegetable and medicinal herb from Cucurbitaceae family, has been used as a botanical dietary supplement in Asia for treatment of dyslipidemic conditions, diabetes, and obesity. The impact of bitter melon major components triterpenoids to gut microbiome was investigated. The results showed that bitter melon triterpenoids (BMTs), including tetracyclic triterpenes and pentacyclic triterpenes, could influence gut microbiota, but the effects varied for different volunteers. According to influence of microbiota, BMTs could be clustered. While some of them showed much stronger effect. Proportions of some beneficial bacteria were significantly promoted by BMTs; meanwhile, proportions of opportunistic pathogens were considerably decreased. Moreover, BMTs influenced the bile acids metabolism. After treatment, concentration of the TGR5 agonists and Farnesoid X Receptor antagonist bile acids increased, which indicated an increase of glucagon-like peptide-1 secretion. After component analysis of bitter melon from different regions, we found that proportions of pentacyclic triterpenes in bitter melons from Guangdong were significantly higher than that from Shandong and Hunan. Since different BMTs influenced microbiota condition and bile acid metabolism diversely, the influences of bitter melons with different chemotypes could vary. In conclusion, bitter melons could benefit to treatment of metabolic disorders through the influence to microbiota.
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

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-371 **Metabolomic mapping of chemodiversity in microbial strains using NMR and multivariate statistical process control (MSPC) to find potential biomarkers**

PRESENTING AUTHOR: John Gauvin, DSM, Netherlands

CO-AUTHORS: Abel Folch Fortuny, Margriet Hendriks, Adriana Carvalho de Souza, Peter van de Vondervoort

NMR has become an established technique for use in metabolic profiling, delivering information on a broad range of compound classes at various concentrations. In proton NMR many compounds give multiple signals within the proton spectrum, which can be used in statistical analysis to detect patterns in multivariate approaches. In a novel approach, we used the metabolic profiles of microbial strains to build a MSPC-based model to find possible biomarkers from the chemodiversity portrayed in spectra. MSPC has been used within industrial processes with many complex parameters and strongly multivariate data, to detect and diagnose deviations in incoming production batches. MSPC can be used to build a model from the "non-hits", considering them as the normal operating conditions results. Then, "hits" can be projected in the model and analyzed as outlier observations (since they were hits). Finally, in this work a MSPC-based model was created to map and distinguish biomarkers from general metabolites.

P-372 **Metabolic engineering of Ustilago trichophora TZ1 for the improvement of malic acid production from glycerol using metabolomics approaches**

PRESENTING AUTHOR: An Phan, RWTH Aachen, Germany

CO-AUTHORS: An N.T. Phan, Lisa Prigolovkin, Lars M. Blank

U. trichophora has attracted increasing attention due to their capability in using glycerol, the main waste streams of biodiesel production process, to produce malic acid. Unlike other commercial filamentous fungi that tend to form pellets during cultivation, Ustilaginaceae can stay haploid and non-filamentous, which are the vital features for industrial applications. Previous optimizations could achieve the strain U. trichophora TZ1, which has the highest reported malic acid titre for microbial production from glycerol. Though, the production yield was quite low. The main obstacle for further strain improvement is the lack of knowledge on U. trichophora metabolic network. In this study, we aim to decipher and reconstruct the metabolic network in U. trichophora TZ1 to improve malic acid production using GCMS-based metabolomics. Metabolites in central carbon metabolic pathways were identified and quantified during malic acid production phase. As a results, the main by-product formation pathways in malic acid production from glycerol were revealed. By using metabolic engineering and medium optimization, we could decrease the level of by-products and successfully improve malic acid production in U. trichophora TZ1.

P-373 **LC/MS-based metabolomics analysis for the improvement of 1-butanol production in engineered cyanobacterial Synechococcus elongatus**

PRESENTING AUTHOR: Artnice Fathima, Osaka University, Japan

CO-AUTHORS: Artnice Mega, Derrick Chuang, Walter Laviña, James Liao, Sastia Putri, Eiichiro Fukusaki

Sustainable 1-butanol production using photosynthetic organisms, such as cyanobacteria, has garnered interest among researchers. 1-Butanol is natively produced via CoA-dependent pathway in Clostridia species. Therefore, a cyanobacterial, Synechococcus elongatus strain capable of producing 1-butanol, named BUOHSE, was developed by introducing a modified CoA-dependent pathway. However, 1-butanol production in this strain is low compared to other model microorganisms. Metabolomics approach for strain improvement has been widely employed in various microorganisms, which enables rapid detection of important metabolites and rate-limiting steps in the production pathway. Thus, comparative metabolome analysis of cyanobacteria strains producing 1-butanol was performed to further improve 1-butanol titer. By using ion-pair reversed-phase LC/MS, the reaction from butanoyl-CoA to butanal, catalyzed by PduP enzyme, was considered as a rate-limiting step. Accordingly, a new strain DC7 with an improved activity of PduP was developed. This strain showed a 33% increase in 1-butanol titer and a decreased level of butanoyl-CoA, indicating that butanoyl-CoA to butanal reaction was improved. To understand the effect of PduP optimization, metabolome analysis of DC7 and BUOHSE was carried out. Results showed acetyl-CoA was highly accumulated in DC7 compared to BUOHSE. To utilize this enhanced level of acetyl-CoA for 1-butanol formation in DC7, acetyl-CoA carboxylase was overexpressed. The resulting strain DC11 was able to reach a production titer of 418.7 mg/L in 6 days, cutting production time in half. In conclusion, metabolomics-assisted strain improvement employed in this study demonstrated the utility of metabolomics to effectively find a rate-limiting step and target for improvement in a biosynthesis pathway.
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PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-374 GC-MS metabolic profiling of volatile compounds produced by Actinobacteria

PRESENTING AUTHOR: Lada Brazdova, Institute of Microbiology, Czech Republic

CO-AUTHORS: Bara Seidlova, Zdenek Kamenik

Actinobacteria, particularly the genera Streptomyces, Amycolatopsis, Rhodococcus, and others, are important producers of bioactive metabolites including antimicrobials, antitumor agents, immunosuppressants, or signaling molecules. However, the mainstream research tends to focus on compounds, which are suitable for liquid chromatography, while volatile compounds may be missed unless they are analyzed by gas chromatography. We cultured various Actinobacteria strains with novel gene clusters including those encoding biosynthesis of new compounds with 4-alkyl-L-proline moieties. We determined the profile of volatile compounds by gas chromatography with mass spectrometry and processed the data using various approaches including GNPS molecular networking for GC-MS data.

P-375 LC-MS/MS and GC-MS metabolomics identifies feedstock components inhibitory to growth of engineered microbial strains

PRESENTING AUTHOR: Mona Elbadawi-Sidhu, Amyris, United States

CO-AUTHORS: Chia-Wei Lu, Perry Kumagai, Quinn Mitrovich, Judith Denery

Amyris strives to deliver high-quality, renewable products from sustainable resources, including cellulosic sugars. To achieve this, advanced microbial engineering coupled with state-of-the-art analytics are innovatively implemented to propel industrial synthetic biology and manufacturing of eight new renewable products in the past ten years. Targeted and untargeted mass spectrometry-based metabolomics promotes data-driven strain and process design that enables manufacturing of our products. One application of these approaches focused on development of strains capable of thriving on wood-derived cellulosic feedstocks. The conversion of wood into microbially-consumable monosaccharides produces a variety of side products that inhibit growth of highly engineered yeast strains. To better characterize the compounds affecting engineered strains, metabolomics was used to elucidate the complex composition of the feedstock. Polar and non-polar chromatography methods coupled to both LC-MS/MS and GC-MS were used to broaden the scope of the metabolic profiles. A multi-pronged approach was applied to determine the identities of the inhibitors: bioassay-guided fractionation of hydrolysate and spent hydrolysate informed the untargeted analysis, which in turn informed the targeted analysis, along with literature search of known feedstock components. Using untargeted metabolomics, we identified six novel components of cellulosic hydrolysate. In total, 70 compounds were tested, and we found that a combination of 26 metabolites could account for the total observed growth inhibition of the Amyris yeast. A summary of the analytical workflow and biological impact is presented.

P-377 Intracellular metabolite profiling and the optimization of metabolome extraction methods for Clostridium carboxidivorans fermenting syngas

PRESENTING AUTHOR: Yu Eun Cheong, Korea University, South Korea

CO-AUTHORS: Jungyeon Kim, Kyoung Heon Kim

Clostridium carboxidivorans, which ferments CO to biofuels via the Wood-Ljungdahl pathway, has received increased attention. Since CO is a unique substrate and is known to be metabolized via metabolisms different from other common carbon sources such as glucose, it is necessary to analyze intracellular metabolite profiles for CO fermentation by C. carboxidivorans to suggest clues for metabolic engineering. Moreover, when fermenting syngas, the metabolite profiles of C. carboxidivorans can be quite different from those under the normal conditions. Therefore, it is essential to optimize the metabolite extraction solvent for syngas-fermenting C. carboxidivorans. In comparison with glucose media, CO-containing syngas media allowed higher levels of intracellular fatty acid synthesis and changes of fatty acid metabolism possibly due to cofactor imbalance. The high levels of intracellular fatty acids could be used to produce high-value chemicals by further microbial engineering. Also, the evaluation of extraction solvents revealed that the mixture of water-isopropanol-methanol (2:2:5, v/v/v) is the best extraction solvent, which showed a higher extraction capability and reproducibility than pure methanol, the conventional extraction solvent for clostridia. This is the first metabolomic study to demonstrate the unique metabolite profiles of CO fermentation, and the mixture of water-isopropanol-methanol as the optimal metabolite extraction solvent for C. carboxidivorans fermenting CO.
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PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-379

Development of LC-MS/MS based acyl-CoAs analysis method and its application to Streptomyces coelicolor

PRESENTING AUTHOR: Katsuaki Nitta, Osaka University, Japan

CO-AUTHORS: Masahiro Furuno, Sastia Prama Putri, Eriko Takano, Eiichiro Fukusaki

Acyl-CoAs are important intermediates in central carbon metabolism, fatty acids metabolism and other important pathways. In addition, secondary metabolism such as polyketide antibiotics synthesis also utilizes acyl-CoAs for intermediate elongation. Hence, a comprehensive analysis platform of acyl-CoAs is necessary to facilitate above pathway intermediate analysis. One of challenges of acyl-CoAs analysis is strong interaction of acyl-CoAs with metal part of LC-MS system and this results in strong peak tailing in liquid chromatography. Consequently this makes it impossible to detect low concentration of acyl-CoAs. In this study, the following five points were investigated to avoid the peak tailing: 1) metal free ODS column selection, 2) mobile phase pH and elution power selection, 3) chelating agent selection, 4) to inert ESI capillary, 5) to inert LC pomp and injector. In summary, non-metal column, which is resistant to highly basic mobile phase, was chosen for sustainable analysis while ammonium bicarbonate and acetonitrile was chosen as mobile phase to deprotonate the phosphate group of acyl-CoAs and avoid metal adsorption. To reduce metal part of LC-MS, fused silica capillary was employed as ESI capillary. In addition, bioinert pump and injector was employed. The peak shapes of acyl-CoAs were improved significantly in the optimized condition. In addition, different extraction solvents were investigated to achieve better coverage and mixture of acetonitrile, isopropanol & phosphate buffer showed good coverage. It was applied to Streptomyces coelicolor. A total twelve acyl-CoAs including highly hydrophobic(C16) acyl-CoA were detected and the LC-MS/MS analysis and extraction method showed good applicability to biological samples.

P-380

Evidence for Streptomyces venezuelae strains harboring potential anthelvencin biosynthetic gene cluster

PRESENTING AUTHOR: Mira Choi, Korea Basic Science Institute, South Korea

CO-AUTHORS: Namil Lee, Woori Kim, Byung-Kwan Cho, Kyoung-Soon Jang

Secondary metabolite biosynthetic gene clusters (BGCs), which encode bioactive compounds such as antibiotics and anti-cancer drugs, in Streptomyces species are often considered as a clue understanding the physiology and evolutionary aspect of those bacteria. In this study, we present highly resolved classification of 25 Streptomyces strains, representing ten S. venezuelae strains and other 15 species, using a matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometer and whole-genome mining. The close relationship of S. venezuelae ATCC 14583, 14584 and 14585 strains were revealed by comparison of the bacterial protein fingerprints by MALDI-TOF MS analysis, meanwhile those strains were not clustered by conventional 16S rRNA sequence alignment. Following whole-genome sequence alignment using 9 publicly available genome sequences of Streptomyces species and three S. venezuelae strains was used to evaluate their phylogenetic relationships, and the results correlate well with the MALDI-MS results. In addition, secondary metabolite BGC analysis of the genomes revealed that S. venezuelae ATCC 14583, 14584 and 14585 distinctly shared the congocidine BGC. Our findings in this study illustrate the importance of accurate bacterial classification and discovery of novel secondary BGCs for better utilization of metabolically relevant bacterial species.

P-381

The metabolisms in oxalate rich-leaf beetle via gut bacteria

PRESENTING AUTHOR: Atsuko Miyagi, Grad. Sch. of Sci. & Eng., Saitama Univ., Japan

CO-AUTHORS: Noriyuki Ojima, Wakako Ikeda-Ohtsubo, Maki Kawai-Yamada

Rumex obtusifolius is an invasive perennial weed, which is one of the most soluble oxalate-accumulated plant among Rumex species. Their oxalate accumulation is maintained at various constraints (high CO2 concentration, nutrition, dark, low temperature, and Al ion stress). Excess intake of soluble oxalate causes mineral (such as calcium and iron ions) deficiency or renal calculus for human and vertebrate. However, an oxalate rich-leaf beetle Gastrophysa atrocyanea feed R. obtusifolius as a host plant. To investigate the mechanism of oxalate metabolisms in the oxalate rich-leaf beetle, we performed metabolomic analysis of G. atrocyanea larva and adults using CE-MS. The results showed that oxalate was not accumulated in both larva and adults. Moreover, other organic acids such as lactate or citrate was highly accumulated. To clarify the effects of gut bacteria on oxalate degradation in leaf beetle, we measured the oxalate and other metabolites in the larva fed with an antibacterial agent tetracycline-treated R. obtusifolius leaves. The results showed that oxalate contents in the larva fed with tetracycline-leaves were highly accumulated than those in the larva fed with control leaves. Moreover, the tetracycline experiments revealed that the leaf beetle would obtain the gut bacteria from not their parents but their feeding leaves.
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.

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PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-382

Contribution of gut microbiota to metabolism of dietary glycine betaine

PRESENTING AUTHOR: Ville Koistinen, University of Eastern Finland, Finland

CO-AUTHORS: Olli Kärkkäinen, Klaudyna Borewicz, Iman Zarei, Jenna Jokkala, Valérie Micard, Natalia Rosa-Sibakov, Seppo Auriola, Anna-Marja Aura, Hauke Smidt, Kati Hanhineva

The protective effect of whole grains against several chronic diseases is being supported by mounting evidence. At the same time, our knowledge is increasing on how gut microbiota influences our health and how diet can modify its composition. Herein, we studied C57BL/6J mice fed with bran-enriched diets, conventional and germ-free C57BL/6NTac mice, and the colonic fermentation of rye bran in an in vitro model of the human gastrointestinal system. We performed 16S rRNA gene sequencing and metabolomics to determine the effect of bran-enriched diets on the gut microbial composition and the potential contribution of microbiota to the metabolism of a novel group of betainized compounds. The bran-enriched diets elevated the levels of betainized compounds in the colon contents and changed the composition of the microbiota by increasing the relative abundance of several bacterial taxa, including Akkermansia, Bifidobacterium, Coriobacteriaceae, Lactobacillus, Parasutterella, and Ruminococcus. The levels of betainized compounds in the germ-free mice were significantly lower compared to conventional mice. In the in vitro model, the production of betainized compounds was observed throughout the incubation, while the levels of glycine betaine decreased. Only low levels or trace amounts of other betaines than glycine betaine were observed in whole-grain cereals. In conclusion, our findings provide evidence that the bacterial taxa increased in relative abundance by the bran-based diet are also involved in the metabolism of glycine betaine into other betainized compounds, introducing another compound group as potential mediators of the synergistic metabolic effect of diet and colonic microbiota.

P-383

Application of high-throughput MS screening to disentangle the bacterial metabolome

PRESENTING AUTHOR: Riya C Menezes, Max Planck Institute for Chemical Ecology, Germany

CO-AUTHORS: Paolina Garbeva, Purva Kulkarni, Aleš Svatoš

Bacteria produce a wealth of structurally diverse specialized metabolites with a remarkable range of activity. Recently augmented knowledge about bacterial genomes has revealed that many bacteria have far greater potential to produce secondary metabolites. This potential, however, has been hampered by difficulties in metabolite extraction or non-expression of genes. Many microbial gene clusters may be silent under standard laboratory growth conditions. One promising approach to trigger the activation of cryptic biosynthetic pathways is the co-cultivation of two or more microorganisms in the same confined environment, leading to the production of new metabolites. Because of the complexity of microbial extracts, advanced analytical methods are key for the successful detection and identification of co-culture-induced metabolites. In the present study, the secondary metabolites produced by 12 phylogenetically different bacteria isolated from soil and rhizosphere were analysed. Liquid Extraction Surface Analysis (LESA) combined with a Thermo Scientific LTQ Orbitrap XL mass spectrometer was applied using various solvents. The bacteria were also selectively cocultured and grown under three different nutrient conditions namely, nutrient-rich 1/10 TSBA; nutrient-poor Water Agar and Water Agar supplemented with Artificial Root Exudates. Our results revealed that LESA with high-resolution MS is a powerful tool for high-throughput extraction and detection of secondary metabolites produced by different bacteria under different culture conditions. Of all the solvents tested, ethyl acetate/acetone (65:35, v/v) containing 0.1% formic acid revealed the most effective extraction. Besides known secondary metabolites, several unknown metabolites were detected in bacterial cultures but not in any of the controls consisting of only media.

P-384

Mapping the chemical diversity between mammalian gut microbiomes – a molecular walk in the zoo

PRESENTING AUTHOR: Rachel Gregor, Ben-Gurion University of the Negev, Israel

CO-AUTHORS: Rachel Gregor, Maraike Probst, Goor Sasson, Stav Eyal, Pieter C. Dorrestein, Michael M. Meijler, Itzhak Mizrahi

In the past decade, studies on the mammalian microbiome have revealed that different animal species show wide variations in their microbial compositions. This diversity is strongly connected to a variety of factors, especially diet, digestive strategy, and animal host phylogeny. However, a key metagenomics study indicated functional redundancy, meaning that despite this taxonomic diversity, the functional capabilities of these microbial communities are similar, although certain functions were found to be enriched based on diet. Here, we propose to characterize the metabolic content of mammalian microbiomes as a direct window into ecosystem function. We have performed comprehensive metabolomics analyses on 101 mammalian fecal samples, as well as 34 dietary samples. Our platform includes LC-MS/MS for semi-polar metabolites and lipids, GC-MS for primary metabolites, and GC-FID for short chain fatty acids. Metabolites were identified using an in-house library of standards, and further explored using the Global Natural Product Social Molecular Networking platform (GNPS; gnps.ucsd.edu) to assign putative structures and group compounds into chemical families based on fragmentation patterns. Additionally, the microbiome of the fecal samples was analyzed by 16S rRNA gene sequencing. We found a strong correspondence between the animals' microbiomes and metabolomes (Mantel test: 0.703; significance: 0.001), as well the association of certain chemical families with different animal groups. We are currently examining how this effect differs for different classes of metabolites, as well as correlating between specific metabolites and microbial species, in order to better elucidate the metabolic capabilities of these communities and the ramifications on their ecological functions.
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PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-385

Mass spectrometry based quantification of intracellular metabolites

PRESENTING AUTHOR: Reza Maleki Seifar, DSM, Netherlands

CO-AUTHORS: Reza Maleki Seifar, Lucien Duchateau, Maurien Olsthoorn

Accurate quantitative data on intracellular metabolite from industrial cell cultures provides key information in metabolic engineering studies which is required for producing useful chemicals. In quantitative analysis of intracellular metabolites a number of important considerations regarding sampling and sample processing are need to be taken into the account: 1) short turn over time of some of the metabolites, 2) continuation of the metabolism of microorganisms at the time of sampling, 3) leakage of intracellular metabolites, 4) interferences from extracellular environments, 5) extraction efficiencies. Nowadays LC-ESI-MS(MS) is one of the most widely used analytical techniques in metabolomics studies. However, for absolute quantification of metabolites it is necessary to have access to standard chemicals and stable isotope labelled internal standards. Internal standards are required for correction of losses during extractions and also matrix effects in MS- based quantitative analysis of metabolites.
The fully 13C-labelled internal standards could be considered as ideal internal standards. Here, we present the application of fully 13C-labelled internal standards which have been produced by microorganisms cultured on 13C-fully labelled glucose as exclusive carbon source. The sampling, quenching, extraction analytical method based on LC-MS/MS are presented for quantitative analysis of metabolites from two important mevalonate and isoprenoid pathways.

P-386

Evolution of gene knockout strains of E. coli reveal regulatory architectures governed by metabolism

PRESENTING AUTHOR: Douglas McCloskey, Biosustain, DTU, Denmark

CO-AUTHORS: Sibei Xu, Troy E. Sandberg, Elizabeth Brunk, Ying Hefner, Richard Szubin, Adam M. Feist, Bernhard O. Palsson

Biological regulatory network architectures are multi-scale in their function and can adaptively acquire new functions. Gene knockout (KO) experiments provide an established experimental approach not just for studying gene function, but also for unraveling regulatory networks in which a gene and its gene product are involved. Here we study the regulatory architecture of Escherichia coli K-12 MG1655 by applying adaptive laboratory evolution (ALE) to metabolic gene KO strains. Multi-omic analysis reveal a common overall schema describing the process of adaptation whereby perturbations in metabolite concentrations lead regulatory networks to produce suboptimal states, whose function is subsequently altered and re-optimized through acquisition of mutations during ALE. These results indicate that metabolite levels, through metabolite-transcription factor interactions, have a dominant role in determining the function of a multi-scale regulatory architecture that has been molded by evolution.

P-387

Metabolic profiling of Candida albicans

PRESENTING AUTHOR: Alberto Spisni, University of Parma, Italy

CO-AUTHORS: Tecla Ciociola, Thelma A. Pertinhez, Laura Giovati, Walter Magliani, Alberto Spisni, Luciano Polonelli, Stefania Conti

Microbial metabolomic analysis focused on extracellular and intracellular metabolites can be used for a number of purposes, including species identification. Currently, Saccharomyces cerevisiae Yeast Metabolome Database is the only one available of fungal metabolites. Here we outline the metabolic profile of Candida albicans SC5314 in different growth conditions. The metabolites of budding and germinating yeast cells were analysed by Proton Nuclear Magnetic Resonance (1H-NMR) spectroscopy. Yeast cells from semi-synchronized cultures suspended in fresh YPD and 199 medium were incubated at 30 and 37°C for 1 and 6 h. Extracellular metabolites, separated by centrifugation and ultrafiltration, and intracellular metabolites, obtained by mechanical cells disruption, were profiled by 1H-NMR on a JEOL 600 MHz ECZ600R and identified and quantified using Chenomx NMR suite 7.6. Distinct metabolic profiles are observed for cells grown in selected conditions. Increase in extracellular metabolites is observed over time (1 to 6 h). 50 intracellular metabolites were annotated. Among the major intracellular metabolites, ethanol, acetate, alanine, betaine, and pyruvate, metabolites involved in energy production pathways, were present in different concentrations in budding and germinating cells. Genes coding for enzymes involved in those metabolic pathways have been confirmed: Candida Genome Database (http://www.candidagenome.org). The results obtained, though preliminary, contribute to increase the knowledge of C. albicans metabolic profile and to create its metabolites database. This strategy will allow to unravel the pathways involved in the transition from budding to germinating phenotype. Characterization of C. albicans main metabolic pathways favour the identification of hot spots for new antifungal agents.
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PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-388 Exploring the Metabolic Fate of Dietary Polyphenols

PRESENTING AUTHOR: Nicola Gray, University of Reading, United Kingdom

CO-AUTHORS: Cintia Rabelo e Paiva Caria, Christine Hinz, Chris Titman, Wagner Vilegas, Glenn Gibson, Gunter Kuhnle

Epidemiological and dietary intervention studies suggest that consumption of polyphenol-rich foods may reduce the risk of cardiovascular disease. Increased intake of polyphenols, found at high levels in tea, fruit and vegetables, has been associated with health-promoting benefits including vascular and platelet function, blood pressure and improved plasma lipid profile. However, the mechanisms behind these benefits remain unclear, and the bioavailability of dietary polyphenols is highly variable between individuals. A significant fraction of dietary polyphenols can persist in the colon, where they are exposed to the gut microbiota. Since the study of the human gut microbiota is hindered by the complexity of this ecosystem and accessibility, in vitro gut models provide a powerful tool to build mechanistic knowledge around microbial polyphenol bioconversion. Combined with metabolomics, this approach enables investigation of broad metabolite perturbations and variations in microbial diversity to enhance mechanistic understanding of polyphenol bioactivity and nutritional influences to improve health and reduce risk of CVD through diet. Here, untargeted and targeted LC-MS metabolite profiling was applied to in vitro gut models from human stool donors in the presence of different flavanol-containing substrates to explore polyphenol metabolism and the influence on gut microbial activity. Distinct metabolic differences were observed in the in vitro culture profiles with the addition of polyphenol substrates in a time dependent manner. Targeted phenolic, bile acid and short chain fatty acid profiles allowed for a more detailed exploration of gut microbial metabolism, providing an in-depth workflow to determine the bioconversion of dietary polyphenol compounds.

P-389 Semi-targeted analysis of folate metabolites in yeast using ultra-high resolution mass-spectrometry

PRESENTING AUTHOR: Lena Gmelch, Technical University of Munich, Germany

CO-AUTHORS: B. Kanawati, P. Schmitt-Kopplin, M. Rychlik

Folates are a group of vitamins consisting of more than 50 different vitamers. These vitamers play an important role in many metabolic processes such as methylation reactions, nucleotide synthesis, or oxidation and reduction processes of C1 units. A lack in folates is closely associated with the incidence of neural tube defects and is assumed to be a major risk factor for several chronic diseases such as cardio vascular diseases or dementia. Until now, folates are mainly analyzed in the respective monoglutamate forms. Little is known so far about the distribution of polyglutamates, which are the major vitamer forms in food. Thus, it is important to gain deeper insights into the metabolism of folates including its polyglutamylated forms. Therefore, a semi-targeted FT-ICR-MS approach was conducted to unravel the complete vitamer distribution including metabolic precursors. Baker's yeast served as model organism since it is known to be a rich source of folates. For obtaining higher sensitivity, sample clean-up via SPE had to be performed prior to measurements. Several signals could be unraveled as tentative folate vitamers. For verification of those compounds, targeted analysis by LC-MS/MS using a low resolution triple quadrupole detector was performed. The results were in good agreement with FT measurements and thus, the assignment could be confirmed. To further investigate the folate metabolism, yeast grown in different chemically defined culture media was analyzed as described above. Differences in the vitamer distribution as well as higher folate contents in media containing methionine could be observed as described previously.

P-390 Development of a Caenorhabditis elegans Reference Material for Long-Term LCMS Metabolomics Quality Control and Unknown Compound Identification

PRESENTING AUTHOR: GONCALO GOUVEIA, University of Georgia, United States

CO-AUTHORS: David L. Blum, Alison Morse, Arthur S. Edison, Lauren M. McIntyre

Robust quality assurance/control measures are an essential component of metabolomics studies. Creating a biological reference material (BRM) that is compliant with six-sigma best practices improves our ability to make inferences from metabolomics experiments. However, this is a challenging task. There are several criteria for a BRM: it should contain features that can be identified on different instruments and platforms and should be able to be produced over-time with negligible variation. This enables corrections for runs, instruments, and time. C. elegans cryo-resistance allows for their genetic identity to be preserved, making it an ideal BRM. Genetically identical batches can be produced from the identical genetic source over-time, a primary source of variation in biological systems. The other source of variation is in the environment during growth. To minimize environmental variance, small batches are produced in bioreactors allowing uniform growing conditions and a homogenous batch. The largest remaining source of variation is the food. We have successfully generated a stable C. elegans food source (E. coli) by ensuring that individual batches are homogenous, combined with an adaptation of methods originally developed by "Student" that include small-batch averaging. QC for individual batches and small batch averaging are common strategies in manufacturing that ensure long-term product consistency. We have divided the process into steps: environmental input control, batch QC, and batch averaging and developed QC metrics for batch acceptance or rejection before averaging. We evaluated our approach by measuring metabolites by NMR and our results indicate that that averaged batches can be 99.99% concordant.
**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.

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P-391 Metabolomes of Sander lucioperca and Rutilus rutilus lacustris gills and lenses

PRESENTING AUTHOR: Lyudmila Yanshole, International Tomography Center SB RAS; Novosibirsk State University, Russian Federation

CO-AUTHORS: Ekaterina Zelentsova, Vadim Yanshole, Yuri Tsentalovich

The metabolomic analysis of fish tissues can be successfully used for the study of impact of external factors on the fish health, including water contamination with pesticides, herbicides, low oxygen level. Changes in the metabolomic compositions of tissues reflect the health status, and metabolomic studies helps to elucidate the molecular basis of pathogenesis or the influence of ecological factors. For example, a problem of extremely high cataract prevalence in farmed salmonids leads to the increase in the metabolomic studies of fish lenses. Earlier we developed a method for quantitative metabolomic analysis of different tissues and biofluids. In this work we applied our approach to the analysis of metabolome of roach and pike-perch lenses and gills. The quantitative metabolomic profiling of samples has been performed with the combined use of high frequency 1H NMR and high-resolution LC-MS methods. The concentrations of 60-80 metabolites in fish lenses and gills have been measured. The most abundant metabolites in the fish lenses are N-acetyl-histidine, N-acetyl-aspartate, serine-phosphoethanolamine and threonine-phosphoethanolamine. These metabolites were attributed to osmolytes. Ovothiol A, one of the most powerful antioxidants existing in nature, was found in fish lenses in millimolar concentrations. The results obtained in the work show high dependence of osmolyte concentration on the season - N-acetyl-Histidine predominates in autumn fish lenses, while myo-inositol - in winter lenses. This difference may be attributed to the low oxygen level in water during the winter season and to the seasonal changes in fish nutrition. Supported by RFBR (Projects N° 19-04-00092, 18-29-13023).

P-392 Plasma metabolomic profiling in chickens under heat stress

PRESENTING AUTHOR: Shozo Tomonaga, Kyoto University, Japan

CO-AUTHORS: Hirofumi Okuyama, Tetsuya Tachibana, Ryosuke Makino

In the poultry industry, exposure to high ambient temperature induces harmful effects, such as growth retardation, increased mortality, and decreased quality and quantity of edible parts (meat and eggs). However, the precise mechanisms underlying physiological and pathophysiological changes induced by heat exposure remain unknown. To understand the influence from a metabolic perspective, we investigated the effects of exposure to high ambient temperature on plasma low-molecular-weight metabolite levels in chickens using gas chromatography/mass spectrometry-based metabolomic analysis. The heat exposure suppressed growth and food intake. Of the 92 metabolites identified, the levels of 29 decreased, whereas the levels of 9 increased. An enrichment analysis using the identified metabolites indicates 35 metabolic processes affected by heat exposure. Among them, the sulfur amino acid metabolic pathway was clearly detected. Changes in the kynurenine pathway in tryptophan metabolism, which could be linked to the immune system and oxidative stress, were also observed. Among the kynurenine pathway metabolites, we reconfirmed increase of quinolinic acid by the quantitative method. These results suggest the possible involvement of various metabolic processes in heat-exposed chickens. Some of them would be important to understand the mechanism of biological responses to high ambient temperature in chickens.

P-393 The search for novel anthelmintic targets: Characterizing alternative metabolic pathways in Caenorhabditis elegans

PRESENTING AUTHOR: Margot Lautens, University of Toronto, Canada

CO-AUTHORS: Samantha Del Borrello, Kathleen Dolan, June Tan, Amy A. Caudy, Andy G. Fraser

Parasitic helminths infect around a quarter of the human population and place a large economic burden on agricultural industries but anthelmintic resistance is a growing problem. Helminths survive long periods of hypoxia in their hosts, but their anaerobic metabolism has yet to be fully characterized. These anaerobic metabolic pathways are a promising target for new drugs. The goal of this project is to find novel, selective drug targets in parasitic helminths by characterizing alternative metabolic pathways in the free-living helminth model, Caenorhabditis elegans. Fumarate reduction, a rewiring of the TCA cycle, has been previously identified as a major metabolic pathway involved in the survival of C.elegans and parasitic helminths during hypoxia. It relies on rhodoquinone (RQ), absent in helminth hosts. RQ-deficient mutants are unable to recover from prolonged exposure to the Complex IV inhibitor cyanide (KCN). To test whether KCN induced fumarate reduction, the TCA cycle phenotype of RQ-deficient mutants and rotenone-induced Complex I disruption was assayed by liquid chromatography-mass spectrometry (LC-MS) and found to be similar. The metabolomic profiles of chemical and genetic disruption of fumarate reduction, knockout of the hypoxia transcription factor, hif-1, and recovery from KCN were interrogated by LC-MS and a computational pipeline developed to allow for metabolite hits to be easily called and visualized. So far, changes in vitamin and starch metabolism have been identified. These pathways will be chemically and/or genetically disrupted then subjected to an in-lab imaging assay to determine their importance to KCN recovery and thus their viability as drug targets.
**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

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-394

Integration of metabolomic approaches in contemporary analysis of bioactive natural products

PRESENTING AUTHOR: Maria Halabalaki, National and Kapodistrian University of Athens, Greece

Natural products (NPs) still retain a predominant position among chemical entities since they are characterized by an unmet and untapped structural diversity and uniqueness. This particular nature is the positive consequence of the evolutionary process, which furnished NPs with competitive advantages, enabling them to interact with biological targets for the benefit of every living organism. The NPs privileged features have led to their broad utilization in medicine, pharmacology, and biology but also in nutrition, agriculture, cosmetics, biotechnology, food chemistry, environmental chemistry, and botany, to name just a few. In the last years, NP's discovery process is experiencing a technological breakthrough. New approaches have been incorporated altering traditional concepts and perceptions. One of these approaches, which was rapidly embraced by the experts in the fiend of NPs, is metabolomics. NP metabolomics, have been rapidly employed for the study of natural entities in a more holistic way, finding multiple applications and uses. Thus, they have been incorporated for chemotaxonomic studies, safety and toxicity assessment, quality control, plant defense, exploration of physiological and biochemical effects in different environmental and stress conditions as well as for the investigation of the physiological development of different organisms and drug discovery. The integration and harmonization of metabolomics with traditional and contemporary methods and approaches effectively facilitates the NPs research. In the context of this talk, different applications of NP metabolomics will be presented using specific examples related to NP drug discovery, chemotaxonomy and quality control of natural entities such as medicinal plants and foodstuffs.

P-395

Metabolic implications of gut health disorders in Norwegian farmed salmons as studied by high resolution magic angle spinning (HR MAS) 1H-NMR spectroscopy of fecal samples

PRESENTING AUTHOR: Violetta Aru, Department of Science, University of Copenhagen, Denmark

CO-AUTHORS: Bekzod Khakimov, Elvis Chikwati, Alexander Jaramillo Torres, Aleksei Krasnov, Trond Kortner, Paul Johan Midtlyng, Åshild Krogdahl, Søren Balling Engelsen

The "GutMatters" project, funded by the Norwegian Seafood Research Fund (FHF), aims at defining the prevalence of gut health disorders and their incidence in sea farmed Atlantic salmon. The project plan included the sampling of a total of 360 salmons from 6 farm sites along the Norwegian coast and over one year period (3 time points). Histopathological analysis, performed on gut samples, revealed inflammatory changes predominantly in the distal intestine but also in the whole intestinal tract in fish with chronic cestode parasitosis. Additionally, enterocyte steatosis was observed in the pyloric caeca of individuals collected in most of the farm sites. HR-MAS (high resolution magic angle spinning) 1H-NMR (proton nuclear magnetic resonance) spectroscopy was used for the first time to analyze intact fecal samples collected from the pyloric caeca and distal intestine of Atlantic salmons. The samples were measured on a 400 MHz spectrometer. The results of the metabolomics analysis evidenced a similar metabolic composition for the samples from the two intestinal regions. Amino acids (i.e. alanine, methionine, and phenylalanine), organic acids (i.e. lactic acid) and sugars were found to be the main components of the fecal metabolome. Significant alterations in the lipid composition of the fecal samples were observed as a result of both dyslipidemia and gut inflammation.

P-396

Exploring the adaptation of rainbow trout metabolome to novel aquafeeds by 1H-NMR metabolomics

PRESENTING AUTHOR: Simon Roques, INRA, Univ. Pau & Pays Adour, E2S UPPA, UMR 1419 Nutrition Métabolisme et Aquaculture, France

CO-AUTHORS: Catherine Deborde, Nadège Richard, Sandrine Skiba-Cassy, Annick Moing, Benoit Fauconneau

The growing industry of aquaculture faces the limited availability of marine resources used to feed farmed fish. The development of plant-based diets alleviates this bottleneck, but carnivorous fish such as rainbow trout still do not tolerate diets based on plant ingredients only. Our objective was to explore fish responses to plant-based diets complemented with alternative raw materials: insect, microalgae, yeast or processed animal proteins. Metabolites were profiled in fish tissues and biofluids using NMR. We also analysed the composition of alternative raw materials to identify compounds susceptible to operate in fish metabolism. The relevance of metabolomics in fish nutrition is illustrated with three results. (1) Histidine, an essential amino acid, was negatively correlated in muscle and plasma with the specific growth rate whatever the diet. In fish, it is considered as a protective buffer agent and prevents the decrease of glycolytic flux during anaerobic process. (2) Betaine, glycine and serine signals were modified in the muscle of fish fed plant-based diet compared to a commercial one. This may correspond to a one-carbon metabolism alteration or to an accumulation from raw material compounds. (3) A quinone compound, specific of microalgae, could be used to assess their incorporation in diet: it was present in raw material and in the liver of fish fed microalgae. Such an approach opens new ways to integrate the interpretation of fish metabolism in diet effect studies. Funding: FUI 2014 (NINAQUA with Le Gouessant, COPALIS, Algae Natural Food, Phileo-Lesaffre Animal Care), ANRT (CIFRE 2016/0775) and MetaboHUB (ANR-11-INBS-0010).
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.

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PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-398

Metabolomics, mixture models and maggots: solutions to agricultural waste problems and food security

PRESENTING AUTHOR: Elizabeth Dickinson, University of York, United Kingdom

CO-AUTHORS: Mark Harrison, Marc Parker, Michael Dickinson, Guy Beavis, James Donarski, Maureen Wakefield, Rosie Nolan, Aida Rafat, Jason Hallet, Adrian Charlton, Julie Wilson

Food security is a global challenge and industrial waste biomass is a major environmental problem. Palm oil waste is currently burned as a means of disposal, and solutions are required to reduce this industry's environmental impact. Palm waste can be recycled into biogas, however, we aimed to optimise conditions for recycling into a nutritious substrate for farmed insects. As a natural component of the diet of farmed animals such as fish, chicken and pigs, fly larvae could meet the increasing demand for protein in animal feed, and can be grown on plant waste. Palm waste is tough and needs to be broken down using microbes (anaerobic digestion), providing a substrate for larvae and producing methane gas, a sustainable energy source. However, digestion processes require adjustments and optimisation, therefore composition of the substrate was investigated. NMR spectroscopy and LC-HRMS were used to analyse palm empty fruit bunches (EFB) after various pre-processing methods, then after anaerobic digestion. Biogas production was also measured. Statistical pattern recognition techniques, such as principal component analysis (PCA), were used to investigate compositional differences before and after digestion, and partial least squares regression (PLS-R) was used to produce models predicting biogas production. Digested EFB were submitted for preliminary insect feeding trials. Subsequently, various domestic waste biomass was submitted for larvae feeding trials and analysed by NMR spectroscopy. Statistical models are being developed to determine the best mixture of these waste streams to produce the most nutritious substrate for larvae, providing an energy-neutral starting point for optimum insect rearing.

P-399

Unravelling the Flavour of White Asparagus using Metabolomics

PRESENTING AUTHOR: Eirini Pegiou, MSc., Netherlands

CO-AUTHORS: Roland Mumm, Robert Hall

Metabolomics has wide application in plant and food analysis. This includes variety differentiation, identification and monitoring of biomarkers for food quality and assessment of improved food products. Food processing often negatively impacts food quality attributes, such as flavour. Asparagus (Asparagus officinalis) is valued for its nutrients and characteristic flavour. Currently more than 30% of the harvested product is discarded as waste due to the harvesting process and strict market requirements. However, this (waste) material could be useful for the production of asparagus powder for use as an ingredient in soups and sauces. With the current drying techniques the characteristic asparagus flavour is not well retained, provoking the need for additional artificial supplements. In this project we aim to evaluate the flavour profile of processed white asparagus materials, in comparison with fresh white asparagus. By analyzing products from different drying processes, we aim to define the optimal conditions which maintain maximum flavour. We use targeted and untargeted metabolomics approaches to evaluate asparagus aroma using GC-MS. Results reveal the complexity of the biochemical composition of fresh and cooked asparagus and how this is modified during processing. Interpreting the effect of different drying strategies on the asparagus aroma is helping us to define an optimal drying process, while contemplating food naturalness. The outcome of this project will enable current crop waste materials to be converted into high value and high quality dried food components.

P-400

The Livestock Metabolome Database: Enabling Livestock Metabolomics Research

PRESENTING AUTHOR: Seyed Ali Goldansaz, University of Alberta, Canada

CO-AUTHORS: Susan Markus, Mark Berjanskii, Rupasri Mandal, Yan Meng, Hamed Pirimoghadam, Debjani Bhattacharyya Chowdhury, Ying Dong, Jiamin Zheng, Paul Luimes, Zhiquan Wang, Graham Plastow, David Wishart

The application of metabolomics to livestock research lags far behind other applications of metabolomics. To further facilitate livestock metabolomics research, we have: 1) created a freely accessible online database called the Livestock Metabolome Database (LMDB; www.lmdb.ca) that contains comprehensive metabolic data for 5 common livestock species (cattle, sheep, goats, horses, and pigs) and 2) performed a detailed experimental characterization of the sheep metabolome. The LMDB is expected to serve as a general hub to support metabolomics studies in livestock research. This database currently holds 1070 metabolites extracted from existing literature and our experimental data collected from studies of cow and sheep metabolomes. Our sheep metabolomic work focuses on profiling the blood metabolome of healthy sheep for marker-assisted prediction of economically important production traits including feed efficiency, carcass merit, pregnancy and litter size. Initial analyses for feed efficiency (n=250) using NMR, ICP-MS, and DI/LC-MS/MS have identified and quantified 191 serum/blood metabolites. A panel of 5 candidate biomarkers (AUC>0.7) distinguishes between lambs with high and low feed efficiency. In a second study, 158 serum metabolites were identified and quantified from pregnant sheep (n=500) with 3 candidate biomarkers (AUC=0.95) that differentiate pregnant from non-pregnant ewes, 35 days into gestation. We have also identified a different pattern in the serum metabolome between pregnant ewes carrying a single fetus versus those carrying multiples. These results are currently being validated using additional serum samples (n=900). These projects will expand the LMDB and help develop a "pen-side" metabolomic test to facilitate farm management practices and animal selection.
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PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-401

Change of volatile metabolites in strawberries (Fragaria ananassa) spoiled by fungi during decay

PRESENTING AUTHOR: SU-YEON NA, EWHA WOMANS UNIVERSITY, South Korea

Strawberry (Fragaria ananassa) is one of the major fruits cultivated all over the world. However, it is highly susceptible to spoilage during shipping and storage, mainly due to food-borne pathogens. In this study, Cladosporium cladosporioides, which is known as a major pathogen of strawberries, was isolated from strawberries infected and inoculated onto disease-free strawberry fruits. After the inoculation of fungi, the decay of strawberries was measured every 12 hours for 3 1/2 days. Then, volatile metabolites produced by fungi grown on strawberry were analyzed and compared during decay periods. Volatile metabolites of the spoiled strawberries were extracted by solid phase micro-extraction (SPME) and analyzed by gas chromatography-mass spectrometry (GC-MS) during decay. Among them, 3-ethylbenzaldehyde, xylene, propan-2-yl butanoate, hex-2-enyl acetate, ethyl octanoate and propan-2-yl acetate were significantly reduced according to the decay periods. On the other hand, 2-methylbutan-1-ol, 3-methylbutan-1-ol, methyl 2-methylbutanoate, methyl (Z)-N-hydroxybenzenecarboximidate and caryophyllene gradually increased over decay periods. In particular, styrene, which is a well-known fungal volatile compound produced from phenylalanine, was produced at relatively high level and significantly increased from 48 hours to 60 hours. Also, data sets were processed by partial least squares-discrimination analysis (PLS-DA) to evaluate the differences in volatile metabolites according to the decay time. The application of PLS-DA revealed that the strawberry samples corrupted by C.cladosporioides could be distinguished according to decay time. Also, major volatile metabolites related to decay were determined. The significant volatile metabolites related to latter state of decay were benzaldehyde, styrene and 2-methylbutyl acetate.

P-403

Non-targeted mass spectrometry-based metabolomics approach for authentication of organic leaf vegetables

PRESENTING AUTHOR: Ka Yi MAN, The Hong Kong Polytechnic University, Hong Kong

Vegetables are an important food source providing essential nutrients, fiber and bioactive phytochemicals. The global trend for consumption of organic vegetable, such as tomatoes, potatoes and leaf vegetables, was increasing rapidly over the last few decades because of the raising demand for healthier and safer diet as well as the raising concerns on environmental sustainability. Despite its profitable prospect and high demand in market, a reliable method to verify whether the plants are cultivated using organic practices is absence. By using non-targeted metabolomics approach, it is possible to acquire comprehensive information on the composition of the metabolite profiles that can be used to identify farming methods. In this research, four types of common leaf vegetables in Hong Kong (n=124), including Amaranthus tricolor L. (red amaranth), Brassica rapa var. parachinensis (choy sum), Ipomoea aquatica Forsk. (water spinach) and Lactuca sativa L. (lettuce), were collected from Hong Kong accredited farms and certificated organic farms. Non-targeted MS-based metabolomics analysis of aqueous and organic layers from liquid-liquid extraction reveals the difference in small molecules profile and lipid profile of leaf vegetables. The result from this study demonstrates that the species and farming methods can be discriminated by a set of primary metabolites and plant lipids.

P-404

Water availability in the soil and its effects on tropane alkaloid metabolism in Datura stramonium

PRESENTING AUTHOR: Fredd Vergara, German Centre for Integrative Biodiversity Research, iDiv, Germany

CO-AUTHORS: Abigail Moreno-Pedraza, Gabriel Jennifer

Datura stramonium is an annual plant that produces tropane alkaloids. Atropine and scopolamine are two of the most studied alkaloids in D. stramonium. Atropine and scopolamine are non-selective muscarinic acetylcholinesterase inhibitors. Atropine and scopolamine affect the central nervous system in animals and act as chemical defenses against herbivores. Molecules of atropine and scopolamine contain nitrogen. There is no evidence that D. stramonium stablishes symbiosis with nitrogen (N2) fixing bacteria. Thus, D. stramonium obtains nitrogen (NO3-) from the soil. The process of nitrogen assimilation requires the dissolving of NO3- in water. It is foreseeable that the availability of water in the soil influences the assimilability of NO3- by the roots of D. stramonium. In turn, the assimilability of NO3- can affect tropane alkaloids biosynthesis. To test this hypothesis we set up an experiment with different irrigations. To estimate the assimilability of NO3- we used tensiometers to quantify the soil water pressure. We analyzed different organs of D. stramonium grown under different irrigations using a non-targeted metabolomics approach (LC-qToF). We also determined the percentages of elemental carbon and nitrogen in different organs. Finally, we performed an absolute quantitation of atropine and scopolamine (LC-QqQ). We identified irrigations correlated with maximum production of tropane alkaloids. These findings are relevant in understanding herbivory patterns in nature.
*AWARD WINNERS

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-405

Effects of the climate conditions around the harvest days on the metabolites in the harvested edamame

PRESENTING AUTHOR: Akira Oikawa, Yamagata University, Japan

CO-AUTHORS: Akira Oikawa, Yamato Horibe, Kei Morita, Katsutaka Takeuchi, Yasutaka Shimizu, Ryosuke Sasaki, Kazuki Saito, Hideki Murayama

Quality of agricultural products is affected by long-term climate changes such as drought and high temperature. Local and short-term climate conditions also have effects on the ingredient in agricultural products. In this study, we focus on the relation between the metabolite changes in edamame (green soybean) and the climate conditions around the harvest days. Edamame was harvested every day of about 10 days for 6 years at the same field. Ionic metabolites and neutral sugars in the harvested edamame were comprehensively analyzed by using capillary electrophoresis mass spectrometer (CE-MS) and liquid chromatography mass spectrometer (LC-MS), respectively. The climate conditions around harvest days were also recorded on the every day. The results of the correlation analysis between the data of the edamame metabolome and the climate conditions revealed that the amounts of some metabolites in edamame were changed depending on the temperature and the day length around the harvest days. Especially, the amounts of lysine in the harvested edamame and the integrated temperature for 5 days before harvest were significantly correlated. Moreover, the amount of sucrose showed weak correlation with the length of day light. Furthermore, although it showed no significant differences, the temperature at the harvest day was inclined to be positively or negatively correlated to the amounts of several metabolites. These results showed the clear relations between the climate around the harvest day and the quality of the harvested edamame, which might lead to add values to agricultural products according to a new viewpoint.

P-406

A Fast LC-QqQ-MS/MS Method for the Quantification of Phytohormones in Stems of Forest Tree Species

PRESENTING AUTHOR: Ana Margarida Rodrigues, ITQB NOVA, Portugal

CO-AUTHORS: Ana Margarida Rodrigues, Swen Langer, Ed Bergström, David Harvey, Tony Larson, Jane Thomas-Oates, Carla António

Phytohormones are key low-abundant metabolites with a critical signaling role to internal and external cues, namely in mediating plant growth and development processes, adaptation to adverse environmental conditions or defence responses. Phytohormones belong to different classes, according to their chemical structure, and are present at very low concentrations in plants, which makes them challenging metabolites to quantify accurately. Thus, continuous improvements of sensitive analytical techniques able to quantify several phytohormones in a single run, and in different plant tissues, are needed. Moreover, most published analytical methods for quantitative analysis of phytohormones were validated for Arabidopsis or crop species, and very few were optimised for complex matrices as forest tree species. The aim of this study was to validate a simple, fast and sensitive analytical method for the quantitative analysis of 14 phytohormones, belonging to six major classes, in the complex matrix of Pinus pinaster Ait. stem tissues, using the triple quadrupole mass spectrometer LC-MS system (LC-QqQ-MS/MS) to take advantage of the high-resolution selected reaction monitoring (SRM) for higher sensitivity and selectivity. This method is fast, reliable, and sensitive, and allows the quantification of 14 phytohormones in a single run (6.6 min). Details on the key analytical validation steps will be presented, namely (i) establishment of calibration curves and assay linearity, (ii) assessment of matrix effects on metabolite quantification, (iii) assessment of the limit of detection and limit of quantification, (iv) assessment of the analytical recoveries during metabolite extraction, and finally (v) determination of the precision of the method.

P-407

Comparative metabolic profiling of soybean seeds from commercial cultivar (Glycine max) and wild soybean (Glycine soja)

PRESENTING AUTHOR: Sung-Dug Oh, Rural Development Administration, National Institute of Agricultural Sciences, South Korea

CO-AUTHORS: Ancheol Chang, Sang Jae Suh, Soo-Yun Park

Metabolomics is a useful tool for determining phenotypic variation in plants. This study aimed to identify the primary metabolites and bioactive secondary metabolites from seeds of wild soybean (Glycine soja) and Korean commercial cultivar (Glycine max, cv. Gwangan) and to explore metabolic differences between their genotypes. We profiled abundant hydrophilic primary metabolite and lipophilic secondary metabolites using gas chromatography-time-of-flight mass spectrometry (GC-TOFMS). Data obtained were subjected to multivariate statistical analyses, principal component analysis, orthogonal partial least squares discriminant analysis, and hierarchical clustering analysis to determine phenotypic variation and relationships between metabolite contents. The identification and profiling of metabolites using GC-TOFMS analysis allows clear discrimination between soybean genotypes. This study determined comprehensive metabolic differences between soybean seeds with different genotypes and provides useful information for genetic manipulation of soybean to influence primary and secondary metabolism.
**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

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-408

A GC-MS platform for assessing the diversity of fruit and leaf volatiles

PRESENTING AUTHOR: JOSÉ L. RAMBLA, Jaume I University. Department of of Agricultural and Environmental Sciences, Spain

CO-AUTHORS: Aurelio Gómez-Cadenas, Antonio Granell

Volatile compounds are important molecules in plants which are involved in several independent processes. Fruit volatiles have a role in the attraction of seed dispersers, and are key compounds in the human perception of fruit flavour. Vegetative tissues also modify their pattern of volatile emission in response to external factors such as the presence of insects and microorganisms, using them as a language for communication with other plant organs and with another plants, and also to communicate with other living organisms such as insects. Here we present some results obtained from our platform for the determination of plant volatiles based on capture by means of Headspace Solid Phase Microextraction (HS-SPME) and separation and detection by means of Gas Chromatography coupled to Mass Spectrometry (GC-MS). This platform was used for the characterization of volatile compounds in different fleshy fruit species, many of them with an impact in fruit flavour, and enabled the identification of genome regions and several genes involved in their biosynthesis. Additionally, a non-targeted approach also allowed the identification of volatile compounds involved in the response of vegetative tissues against both microbial and pest attacks, shedding some light in the mechanisms involved in plant defence.

P-409

Storability and variety-specific metabolite profiles of European onion landraces

PRESENTING AUTHOR: Christoph Weinert, Max Rubner-Institut, Germany

CO-AUTHORS: C.H. Weinert, M. Häußler, M.L. Romo Pérez, B. Egert, M. A. Frechen, B. Trierweiler, C. Zörb, S.E. Kulling

Onions are among the most popular and economically most important vegetables worldwide and contain a range of health-beneficial compounds. Under appropriate conditions, onions can be stored for up to nine months. Modern conventional farming heavily relies on the use of hybrid varieties which may lead to genetic erosion and a loss of biodiversity. The use of open-pollinated landraces and their further development by breeding may be a viable alternative, especially for organic farming. In this trial, we examined the metabolite profiles of nine landraces and a commercial control variety (Sturon) in the fresh state and after cold storage for up to five months at 2-3°C and <60% relative air humidity. Quantitative basic analyses as well as an untargeted GC×GC-MS analysis were performed. More than 200 onion metabolites were relatively quantified. In the fresh state, nine varieties could be separated into three groups, mainly according to differences in the sugar and amino acid profiles. The variety "Jaune des Cévennes" was characterized by remarkably low fructan levels and a higher content of monosaccharides. While this variety suffered from increasing Botrytis and Aspergillus infestations already after two month of cold storage, all other varieties demonstrated minimal water loss and no visual appearance of degradation even after five months. Storage lead, among others, to increased levels of glutamine, glutamate, asparagine, phenylalanine, serine, several sulfur-containing amino acids, fructose, sucrose, xylose, trehalose, raffinose, myo-inositol, phosphate, malate and a substantial decrease in, e.g., citrate, nystose and other di-, tri- and tetrasaccharides.

P-410

Increased throughput and coverage for the annotation of Saponins using a structure-based MSn approach on a Tribrid Orbitrap mass spectrometer

PRESENTING AUTHOR: Reiko Kiyonami, Thermo Fisher Scientific, United States

CO-AUTHORS: Caroline Ding, Seema Sharma, Andreas Huhmer

Saponins are major components of Chinese medicines and exert various pharmacological effects, such as cardiovascular protective activity and anticancer activity. Plus, they could reduce the side-effects of radiotherapy and chemotherapy. The comprehensive annotation of Saponins from various Chinese medicines remains challenging because of the limited availability of authentic standards and the structural diversity of this class of compounds. Taking advantages of high resolution MS and MSn capability offered by the tribrid Orbitrap mass spectrometer, we developed a product ion-dependent MSn data acquisition method in which MS2 data is constantly collected and further followed by higher order FTMSn if sugar neutral lose are detected from the MS2 data. The collected MSn tree data were used to identify the Saponin class compounds which contain the triterpenoid or spirostane aglycones. Chemspider and custom databases are further used for final saponin structure annotation. As the proof of concept, 80% methanol extracts from Sanqi, a Chinese medicine was analyzed. A Thermo Scientific™ Orbitrap ID-X™ Tribrid™ mass spectrometer was used for collecting HRAM MS and MSn (up to MS4) data. The MSn data were searched against msCloud spectral library using Thermo Scientific™ Mass Frontier™ 8.0 software for identifying the saponin class compounds. The novel structure ranking tools included in the Thermo Scientific™ Compound Discoverer™ 3.0 software was used for final structure annotation of identified saponin class compounds. More than 60 sapanins were annotated from the Sanqi extract. The MSn data were critical for the identification of triterpenoid or spirostane aglycones.
*AWARD WINNERS

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-411 NMR and Mass based metabolomics applied to the quality control of processed ginseng

PRESENTING AUTHOR: Dae Young Lee, Department of Herbal Crop Research, National Institute of Horticultural and Herbal Science, RDA, South Korea

CO-AUTHORS: Dahye Yoon, Bo-Ram Choi, Jae-Suk Ban, Ryong Gong, Young-Seob Lee, Geum-Soog Kim

The root of Panax ginseng C.A. Meyer has been widely used as ingredients for traditional medicines and functional foods. Furthermore, not only raw ginseng but also several processed ginseng products have been used in the food industry and herbal markets. According to the distinct processing methods, there are four processed P. ginseng products including white ginseng (WG), tae-geuk ginseng (TG), red ginseng (RG), and black ginseng (BG). In this study, HR-MAS NMR and UPLC-QTOF/MS-based metabolomics approaches were applied to assess the metabolic compositions of four processed ginseng. In the results of HR-MAS NMR, primary metabolites were identified such as sugars and amino acids. In the multivariate analyses of NMR spectra, PCA score plot showed that TG and RG were clustered each other, and discriminated from WG and BG. The main metabolites causing these cluster were sucrose and maltose. Maltose and sucrose were not detected in the WG and BG, respectively. In PCA score plot, four groups of processed ginseng were well differentiated. OPLS-DA was also performed to discriminate two selected samples. Several ginsenosides were found as the key components to differentiate four ginseng products. UPLC-QTOF/MS with an in-house library was used to profile ginsenosides from these four ginseng products. The numbers of identified ginsenosides from the four products were as follows: WG (n=26), TG (n=28), RG (n=34), and BG (n=36).

P-412 The effect of storage on the metabolite profile and antibactirial activity of Plectranthus madagascariensis

PRESENTING AUTHOR: Mhlonipheni Msomi, University of South Africa, South Africa

CO-AUTHORS: Gerhard Prinsloo

Plectranthus madagascariensis is a garden plant native to South Africa that is used to treat minor ailments. The study was conducted to investigate metabolite changes of P. madagascariensis using NMR based metabolomics and how it affects antibacterial activity against Staphylococcus aureus. Different storage conditions and the effect of storage conditions on the antibacterial activity of the plant were investigated. In the NMR-based metabolomics analysis, the supervised OPLS-DA separated samples that were stored in -80°C from those that were stored in the fridge and room temperature. Longer storage time of the leaf samples did not make a difference as the samples were scattered with no clustering observed. Drying conditions had no effect on metabolite profile in samples that were stored in different drying conditions. The resazurin-based assay demonstrated samples stored for one month was not active as most of them had a MIC of 1 mg/ml. The antibacterial activity of extracts tested after three months increased. Extracts that were stored in -80°C immediately after drying were unable to inhibit bacterial growth with the MIC values of 1 mg/ml. The best MIC value was 0.015 mg/ml for plants that were grown in the shade, dried in the sun and stored at room temperature. The extracts stored at room temperature showed different MIC values compared to extracts in the fridge, with generally better activity in samples stored at room temperature.

P-413 Terpenoid Diglycosides in the Salicaceae

PRESENTING AUTHOR: Alice Bellisai, Rothamsted Research, United Kingdom

CO-AUTHORS: Gianluca Ruvo, Charlotte Lomax, Michael H. Beale, Jane L. Ward

Willow is well known for the presence of phenolic glycosides, including the famous compound Salicin from which the well-known drug Aspirin was later developed. Terpenes, abundant in many plants are however less well known in members of the Salicaceae such as willow and poplar. Rothamsted Research maintains the National Willow Collection (NWC) as a short-rotation coppice plantation at Rothamsted Research. Comprising 1500 accessions, this germplasm resource is a rich source of novel chemistry which is being systematically screened by metabolomics. A family of novel terpene alcohol diglycosides have been identified which show a variety of carbohydrate components in their diglycoside moiety. Compounds have been isolated from Salix genotypes by HPLC and their structures were elucidated by LC-HRMS-MS and 1D and 2D NMR. We will demonstrate the diversity in both terpenoid and glycoside components, including data describing both pyranose and furanose forms. Finally, we will illustrate how particular molecules are species specific.
PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-414
Response of bush tea's (Athrixia phlycoides DC.) biochemical constituents to Kelpak® application using the LC-MS/MS triple quadrupole and 1H NMR techniques

PRESENTING AUTHOR: Keletso Mohale, University of South Africa, South Africa

CO-AUTHORS: Kafua Lodama, Mudau Fathuwani Nixwell

Biostimulants from seaweed such as Kelpak® (Ecklonia maxima O.) extracts have gained significance in agriculture and improved yields and quality of various crops. However, the effect of Kelpak® (E. maxima) on bush tea's (Athrixia phlycoides DC.) phytoconstituents has not been explored. The objective of this study was therefore to assess the influence of Kelpak® biostimulant on bush tea's chemical profile using LC-MS/MS triple quadrupole mass spectrometer and 1H nuclear magnetic resonance (NMR) spectroscopy. The treatments consisted of 0.2%, 0.1%, 0.05% Kelpak® and control (0.0%) arranged in a completely randomized design (CRD). Kelpak® application stimulated bush tea plant height and increased number of lateral branches produced. The study also revealed that bush tea's primary metabolites including amino acids, organic acids and vitamins were enhanced by application of 0.2% Kelpak®. The partial least squares-discriminant analysis (PLS-DA) demonstrated four distinct groups of bush tea samples treated with different levels of Kelpak®. Based on the 1D NMR spectrum prediction putative annotation of metabolites resulted in detection of 32 metabolites with varying intensities. The application of 0.2% Kelpak® demonstrated high intensities of the detected compounds and could thus be recommended for improved bush tea plant growth and biochemical constituents.

P-416
The Metabolic Characteristics Analysis of Tobacco Leaves from Different Flavor-Style Locations

PRESENTING AUTHOR: Pingping Liu, Zhengzhou Tobacco Research Institute of CNTC, China

CO-AUTHORS: Liu Pingping, Chen Qiansi, Zhang Hui, Zheng Qingxia, Xu Guoyun, Zhai Niu, Wang Chen, Jin Lifeng, Cao Peijian, Zhou Huina

Metabolomics can be used to characterize the metabolites after various internal and external environmental disturbances (gene changes or environment changes)[1-2]. Metabolites are the material basis of formation of tobacco flavor. The flavor of tobacco directly affected by the changes of metabolites concentration. Before the previous research, the flavor of tobacco has a high correlation with the location and the climate environment. In this study, the mature tobacco leaves (middle leaf of fresh tobacco at maturity stage) from 20 sampling sites were collected (including three traditional flavor types) on a national scale, and the metabolic characteristics of these samples were analyzed. The characteristic metabolites which were closely related to flavor style were investigated. The result revealed that, 21 metabolites were related to heavy-flavor type, 16 metabolites were related to fen-flavor type and 10 metabolites were related to neutral-flavor type. The metabolic pathways enrichment results indicated that, the lipid metabolism level of heavy-flavor tobacco leaves was higher, especially the glycolipid metabolism. The triglyceride metabolism level of fen-flavor tobacco was higher and the sugar metabolism level was low. The ascorbic acid salt synthesis and metabolism of neutral-flavor type tobacco leaves was also strong.

P-417
Metabolomic analysis of headspace volatiles of Plectranthus neochilus Schltr. (Lamiaceae)

PRESENTING AUTHOR: Alexandra Sawaya, FCF-UNICAMP, Brazil

CO-AUTHORS: Maria Isabel Galbiatti, Guilherme Perez Pinheiro, Elisa Ribeiro Antunes, Vinícius Veri Hernandes, Marcos Nogueira Eberlin

Plectranthus neochilus Schltr. (Lamiaceae) is popularly known as boldo in Brazil and used for digestive problems, similar to the Chilean plant, Peumus boldus. The essential oil was proven to be schistosomicidal, antimicrobial and reduced whitefly colonization in tomato crops. Despite these promising activities, there were no studies in the literature on the variability of its essential oil throughout the year. Samples were collected from four individuals grown in the field (monthly, morning and afternoon) and inside a greenhouse. All the collected leaves were immediately frozen. Prior to analysis the samples were crushed in liquid nitrogen, a 500 mg aliquot of each was sampled by SPME fiber (PDMS / DVB) and inserted in GC-MS (Agilent) using an automatic injector (splitless), temperature gradient 60 -246 °C at a rate of 3 °C / min. QC samples were analyzed every 16 samples. The chromatograms were aligned and the features extracted by XCMS-Online, chemometrics by Online MetaboAnalyst and GraphPad Prism 6.01 software for ANOVA. One compound, 1-octen-3-ol was a marker for the afternoon samples in the field and the greenhouse samples for all individuals. This compound is present in edible mushrooms, but in plants only one study related it activation of defense genes. Possibly high solar incidence in the afternoon and high temperatures in the greenhouse caused environmental stress for P. neochilus plants. There was, however, no significant monthly variation of the volatiles, indicating that it is possible to harvest leaves throughout the year.
PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-418

Metabolic responses to potassium availability and waterlogging reshape respiration and carbon use efficiency in oil palm

PRESENTING AUTHOR: Jing Cui, Australian National University, Australia

CO-AUTHORS: Marlène Davanture, Michel Zivy, Cathleen Mirande-Ney, Emmanuelle Lamade, Guillaume Tcherkez

Oil palm is a strong potassium (K)-demanding species cultivated in regions where soil K availability is generally low and waterlogging caused by tropical heavy rains can limit further nutrient absorption. However, the metabolic effects of K and waterlogging have never been assessed precisely. This is very surprising because most tropical or wet areas where rice, sunflower, oil palm and other important crops are cultivated combine these two environmental constraints. The aim of this study was to understand the overall impact of waterlogging and limited K availability on metabolism, and to understand how such advert environmental conditions reshape the carbon balance in oil palm. Here, we examined the metabolic response of oil palm saplings in the greenhouse under controlled conditions (nutrient composition with low or high K availability, with or without waterlogging) using gas exchange, metabolomics and proteomics analyses. Our results show that both low K and waterlogging have a detrimental effect on photosynthesis but clearly stimulate leaf respiration, thereby impacting the carbon use efficiency. Omics analyses show differential accumulation of typical metabolic intermediates and enzymes not only of the Krebs cycle but also of alternative catabolic pathways. In addition, we found a strong relationship between metabolic composition and the rate of leaf dark respiration. Overall, advert environmental conditions have an enormous impact on respiration in oil palm. Leaf metabolome and proteome appear to be good predictors not only of K availability but also of CO2 efflux, and this opens avenues for cultivation biomonitoring using functional genomics technologies.

P-419

Assessment of the arsenic exposure in rice (Oryza sativa japonica): Combining untargeted metabolomics and lipidomics

PRESENTING AUTHOR: Miriam Pérez Cova, IDAEA-CSIC, and Universitat de Barcelona, Spain

CO-AUTHORS: Miriam Pérez-Cova, Romà Tauler, Joaquim Jaumot

Arsenic polluted soils have a direct effect on edible plants-base foods such as rice (Oryza sativa japonica). Metabolomics and lipidomics strategies aid to achieve a better understanding of how arsenic is assimilated and translocated, and the main metabolic pathways affected by this pollutant. Two As exposures were applied during the first three weeks of rice growth in an environmental chamber: by irrigation (1 or 1000 µM) and settled in soil before planting (5 or 50 ppm). Aerial parts and roots were collected, extracted (using two extraction protocols, for metabolites and lipids) and analyzed separately. Metabolomics LC-MS analysis was conducted using a TSK Gel Amide-80 HILIC column and a Thermo OrbitrapQExactive mass spectrometer operating in both, Full MS and All Ion Fragmentation mode. Lipidomics LC-MS analysis employed a Kinetex C8 column and a Waters LCT PremierToF. Ionization source for metabolomic and lipidomic analysis was Electrospray in positive and negative mode. Workflow for metabolomics (W4m) platform was used for preprocessing, and multivariate tools (PCA, PLS-DA, ASCA) from Metaboanalyst were employed in the subsequent data analysis. Results obtained after chemometric analysis show that only the highest As concentration levels (both in irrigation water or settled in soil) allowed the differentiation when compared with control samples (without arsenic treatment). No clear differentiation was found between the low concentration samples from both treatments and control samples. Finally, metabolomics and lipidomics results were integrated to have a more comprehensive overview of As exposure effects in rice, by combining the new hypotheses coming from the detected compounds.

P-420

A novel method for identification and quantification of sulfated flavonoids in plants by neutral loss scan mass spectrometry

PRESENTING AUTHOR: Sabine Metzger, University of Cologne, Germany

CO-AUTHORS: Niklas Kleinenkuhnen, Felix Büchel, Silke C. Gerlich, Stanislav Kopriva

Sulfur is present in plants in a large range of essential primary metabolites, as well as in numerous natural products. Many of these secondary metabolites contain sulfur in the oxidized form of organic sulfate. However, except of glucosinolates, very little is known about other classes of such sulfated metabolites, mainly because of lack of specific and quantitative analytical methods. We developed an LC-MS method to analyse sulfated flavonoids, a group of sulfated secondary metabolites prominent, e.g., in plants of the genus Flaveria. The method uses a linear gradient of methanol/formic acid in water on a Restek Raptor C18 Core-Shell column for separation of the compounds. The sulfated flavonoids are detected by mass spectrometry (MS) in a negative mode, using a neutral loss of 80 Da after a collision induced dissociation. With this method we were also able to quantify the sulfated flavonoids. We could detect all (mono)sulfated flavonoids described before in Flaveria plus a number of new ones, such as isorhamnetin-sulfate-glycoside. In addition, we showed that sulfated flavonoids represent a substantial sulfur pool in Flaveria, larger than the thiols glutathione and cysteine. The individual species possess different sulfated flavonoids, but there is no correlation between the qualitative pattern and type of photosynthesis. Similar to other sulfur-containing secondary compounds, the concentration of sulfated flavonoids in leaves is reduced by sulfur starvation. The new LC-MS method will enable qualitative and quantitative detection of these secondary metabolites in plants as a pre-requisite to addressing their functions.
**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

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-421

Construction and new methodology research on untargeted metabolomics database of tabacoo

PRESENTING AUTHOR: Zheng Qingxia, Zhengzhou Tobacco Research Institute of CNTC, China

CO-AUTHORS: Peng Lu, Yongjie Yu, Lifeng Jin, Pingping Liu, Qiansi Chen, Niu Zhai, Guoyun Xu, Hui Zhang, Jingjing Jin, Peijian Cao, Huina Zhou

Metabolomics, a powerful tool to investigate the relationship between genotype and phynotype, attracts an increasing amount of attention. Ultra-high performance liquid chromatography combined with high-resolution mass spectrometry, such as UPLC-QTOF, has become a dominant technique in metabolic studies, particularly for untargeted metabolomics. An obvious advantage of UPLC-QTOF-based untargeted metabolomics is that thousands of components can be simultaneously analyzed, possessing two great challenges for users to data analysis and identify metabolites. Toward these challenges, we constructed an automatic UPLC-HRMS data analysis software, named AntDAS and a freely accessible metabolite database, named TMMLib. AntDAS can be perform data analysis to effective and efficiently extract features of metabolites in samples, whose output involving high-accuracy m/z values, retention time can be directly imported into the TMMLib for compound identification. TMMLib includes an annotated list of known metabolite structural information and their MS-MS spectra from tobacco. TMMLib can realize single and batch searches, as well as MS-MS searches. AntDAS and TMMLib will be regularly updated and to greatly benefit future metabolomics studies in plant. TMMLib is freely available at http://tmmlib.tobaccodb.org and the AntDAS code is available from http://software.tobaccodb.org/software/antdas.

P-422

Leaf metabolite profiling reveals biochemical diversities within wheat genetic resources

PRESENTING AUTHOR: Annick Moing, INRA Bordeaux, France

CO-AUTHORS: Pierre Petriacq, Amelie Flandin, Sylvain Prigent, Cedric Cassan, Stephane Bernillon, Dominique Rolin, Renaud Rincent, Christophe Salon, Christian Jeudy, Jacques Le Gouis, Yves Gibon, A. Moing

BreedWheat (www.breedwheat.fr) and ArchiRac (www.fsov.org) are French national projects aiming at providing new tools and materials for wheat breeding, thereby increasing and facilitating the use of original genetic resources to improve allelic variability in the elite wheat (Triticum aestivum L.) gene pool. A large panel of elite winter-wheat varieties was selected to cover the diversity present in French material and phenotyped in multi-environment trials to identify tolerance traits and QTLs within the BreedWheat project. In addition, two young plants per genotype were cultivated in controlled conditions within specially designed containers (Jeudy et al. 2016, Plant Methods 12: 31) for high-throughput root-phenotyping for three weeks of vegetative growth within the ArchiRac project. Six reference genotypes were used to account for block variation in the experimental design. The largest fully-emerged leaf of each plant was harvested in the morning, rapidly frozen and lyophilized for biochemical studies. Robotized targeted analyses of major metabolites and liquid chromatography coupled to mass spectrometry (UHPLC-Orbitrap-MS) were used to assess the biological variation of primary and specialized metabolites as well as starch and total protein content. Multivariate and univariate statistical analyses were used to mine the profiles of all genotypes. Compositional distances between genotypes were calculated and visualized using clustering in order to compare metabolic and molecular similarities. Metabolomic data will be further combined to other phenotyping data to search for metabolic biomarkers that will link to plant performance. Acknowledgements: MetaboHUB (ANR-11-INBS-0010), PHENOME (ANR-11-INBS-0012), BreedWheat (ANR-10-BTBR-03) and ArchiRac (FSOV 2016K) projects for financing.

P-423

Metabolic flux may active through freeze drying process of tobacco leaves

PRESENTING AUTHOR: Huina Zhou, Zhengzhou Tobacco Research Institute of CNTC, China

CO-AUTHORS: Qiansi Chen, Pingping Liu, Qingxia Zheng, Niu Zhai, Huina Zhou

To fully investigate the effects of vacuum freeze drying on the metabolome of tobacco leaves, the fresh-frozen samples in liquid nitrogen were divided into 3 aliquots, one kept at frozen as fresh samples and the left two freeze-dried under controlled or uncontrolled shelve surface temperature as FD1 and FD2 samples, respectively. Three kinds of samples were balanced in equal dry weight, and analyzed by GC-MS and LC-Q-TOF with six replicates to collect their metabolome data. A total of 79 and 99 metabolites were identified by GC-MS and LC-Q-TOF, respectively. PCA score plots showed that freeze-dried smaples (FD1 and FD2) did different from fresh-frozen samples in metabolomics level, and LC-Q-TOF analysis also revealed a clear difference resulted by controlled shelve surface temperature. Furthermore, based on VIP list and T-test analyses, a tendency was found that different metabolites identified by GC-MS usually decreased upon freeze drying, while different metabolites identified by LC-Q-TOF were mostly increased. Considering the category of different metabolites, sugars, organic acids and amino acids usually down-regulated, while terpenoids and polyphenol/flavonoids usually up-regulated by freeze-drying, the different in metabolome after freeze-drying may result by the active metabolic flux from primary metabolites to secondary metabolites, besides the volatilization of some volatile compounds. And we also found that freeze-drying with controlled shelve surface temperature might slow down the metabolic flux, since the total levels of different metabolites in FD1 were closer to Fresh samples other than FD2.
**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

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-424 A GWAS study of Arabidopsis secondary metabolism

PRESENTING AUTHOR: Marlies Brouckaert, VIB, Belgium

CO-AUTHORS: Rene Hofer, Geert Goeminne, Veronique Storme, Yvan Saeys, Kris Morreel, Wout Boerjan

A combination of high-throughput metabolomics with genome-wide association studies (GWAS) can contribute to the identification of genes underlying metabolic diversity and their relevance to complex traits, such as metabolite levels, in different accessions of the same species. In this research project, the metabolic profiles of 14-day-old seedlings from 225 Arabidopsis thaliana accessions in 5 biological replicates were analyzed by untargeted liquid chromatography-mass spectrometry. Across all chromatograms, 4479 m/z features segregated as binary traits, i.e. being absent or present. Co-segregation analyses with SNPs, obtained from a previously published 250k SNP chip data set (Horton et al., 2012), is currently being performed. In total, 1311 associations (Fisher's exact test P value < 1x10-10) containing 1046 unique genes were found between the SNPs and absence/presence of 194 m/z features. Focusing on the structurally characterized m/z features, 3 glucosinolates were associated to the MAM2 locus, known to be involved in the biosynthesis of this class of metabolites. In addition, an association between three characterized flavonol-glycosides and the BGLU6 gene was found. Moreover, an association between isoleucin and malonyl-isoleucin and DAAR1 could be retrieved. Supported by these previously proven associations, this method will be applied to find new gene-metabolite links. Horton, M.W. et al. Genome-wide patterns of genetic variation in worldwide Arabidopsis thaliana accessions from the RegMap panel. Nature genetics 44, 212-6 (2012).

P-425 Comparative metabolite analysis between Lycoris radiata and Narcissus tazetta

PRESENTING AUTHOR: Chang Ha Park, Department of Crop Science, Chungnam National University, South Korea

CO-AUTHORS: Sang Un Park

Comparative metabolite analysis between Lycoris radiata and Narcissus tazetta. Chang Ha Park and Sang Un Park* - Department of Crop Science, Chungnam National University, 99 Daehak-ro, Yuseong-gu, Daejeon 34134, Korea. This study aimed to comprehensively examine the interface between primary and secondary metabolites in Lycoris radiata (L. radiata) and Narcissus tazetta (N. tazetta), using gas-chromatography coupled with time-of-flight mass spectrometry (GC-TOFMS) and high-performance liquid chromatography (HPLC). There was significant variation of chemical composition in the different parts of L. radiata and N. tazetta. In particular, The level of galantamine was highest in leaves of L. radiata. However, bulbs of N. tazetta contained the highest level of galantamine. Primary and secondary metabolites identified by GC-TOFMS and HPLC were subjected to partial least-squares discriminant, Pearson's correlation, and hierarchical clustering analyses, which indicated significant differences in the primary and secondary metabolisms of L. radiata and N. tazetta. This metabolome study comprehensively describes the relationship between primary and secondary metabolites in L. radiata and N. tazetta and provides the information useful for developing strategies to enhance the biosynthesis of galantamine in Amaryllidaceae plants. Additionally, This work highlights that HPLC and GC-TOFMS-based metabolite profiling is suitable techniques to evaluate morphological variation and determine metabolic differences in L. radiata and N. tazetta. *(Corresponding author) E-mail: supark@cnu.ac.kr, Tel: +82-42-821-6730. (Acknowledgement) This research was supported by the Bio & Medical Technology Development Program of the National Research Foundation (NRF) funded by the Ministry of Science, ICT & Future Planning (2016M3A9A5919548).

P-426 Integrated metabolome and transcriptome analysis of plants identifies biosynthetic pathways for floral volatile organic compounds

PRESENTING AUTHOR: In-Cheol Jang, Temasek Life Sciences Laboratory, Singapore

CO-AUTHORS: Savitha Dhandapani

Plants emit a number of floral volatile organic compounds (VOCs) as secondary metabolites that have ecological functions for their defence against herbivores and pathogens as well as for the pollinator attraction. Plant VOCs are mainly composed of terpenoids, phenylpropanoids/benzenoids, and volatile fatty acid derivatives, which are derived from different biosynthetic routes in plants. The diversity of plant VOCs has stimulated broad systems biology approaches to identify the pathways/genes involved in their biosynthesis. We integrate metabolome and transcriptome analysis of aromatic flowers to unravel biosynthetic pathways for floral VOCs. Our metabolite-guided transcriptomics and molecular and biochemical characterization of genes have identified specific gene members encoding enzymes involved in the biosynthesis of diverse floral scents. Here, we will talk about our recent progress on genes/pathways identification for biosynthesis of floral scents.
**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

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-427 A metabolomics profiling of a traditional Chinese medicine Radix Stemonae

PRESENTING AUTHOR: Chunping Tang, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, China

CO-AUTHORS: Shuai-Zhen Zhou, Chang-Qiang Ke, Yang Ye

Natural products have been a major resource of new drugs due to their highly diverse structures and biological activities. Searching for natural products with the potential to be lead compounds from traditional Chinese medicines (TCMs) have drawn more and more attention. A metabolomics investigation was carried out on a commonly used TCM Radix Stemonae, from which an array of alkaloids reflecting its therapeutic effects were discovered. Radix Stemonae, known as Baibu in Chinese, is a commonly used antitussive agent. The Chinese pharmacopeia documents three species, Stemona tuberosa, S. sessilifolia and S. japonica as its original plants, and spring and autumn as the collecting seasons. Our previous investigations have revealed that the Stemona alkaloids are the characteristic secondary metabolites responsible for the therapeutic effects. In this study, a metabolite profiling method based on the UPLC-QToF technique was established and applied for 218 samples of crude alkaloids prepared from three Stemona species collected in different localities and seasons. PCA and PLS-DA models were used to analyze the data. The results showed that the alkaloidal constituents vary greatly with species and localities while little with harvesting seasons. The PCA model can be successfully applied to identify the species of unknown Stemona samples. Chemical markers of each species were identified by using an PLS-DA model. The results further revealed that the characteristic alkaloid of S. sessilifolia collected in Shandong province is different from that collected in Anhui province. The findings provide scientific evidence to support traditional descriptions of Baibu in medicinal books.

P-428 An evaluation of different ripening conditions and postharvest treatments in mangosteen (Garcinia mangostana) using metabolomics approach

PRESENTING AUTHOR: Anjaritha Aulia Rizky Parijadi, Osaka University, Japan

CO-AUTHORS: Sobir Ridwani, Fenny M. Dwivany, Sastia P. Putri, Eiichiro Fukusaki

Mangosteen is a tropical fruit with a high market value but a relatively short shelf-life. Despite being one of the most important tropical fruits, evaluation of different ripening condition and evaluation the effect after postharvest treatment to prolong shelf-life using metabolomics approach in mangosteen have never been studied. The aims of this study were to evaluate the metabolic changes between different harvesting and ripening condition and to evaluate the effect of postharvest treatment in mangosteen. Mangosteen ripening stage were collected with several different conditions ("on-tree", and "off-tree"). The metabolite changes were investigated for each ripening condition. Additionally, fruit was harvested in stage 2 and was treated with several different treatments (storage at low temperature (LT; 12.3 ± 1.4 °C) and stress inducer treatment (SI; methyl jasmonate and salicylic acid) in comparison with control treatment (normal temperature storage) and the metabolite changes were monitored over the course of 10 days after treatment. Our findings clearly indicate that there is a similar trend of metabolic changes (the accumulation of some aroma precursor metabolites and firmness-related metabolites in the flesh and peel part, respectively) between two ripening conditions although the progression of "off-tree" ripening process observed through color changes occurred faster compared to "on-tree" ripening. Additionally, the metabolome data and color changes observation showed that LT treatment could prolong shelf-life among all treatments in all fruit part. It is the first report of the utilization of metabolomics to support postharvest development strategies in mangosteen.

P-429 Effect of water restriction and rehydration on flavonoid profile in leaves of Eucalyptus urophylla

PRESENTING AUTHOR: Hana Karina Pereira da Silva, University of São Paulo, Brazil

CO-AUTHORS: Thaís Regiani Cataldi, Carlos Alberto Labate

Eucalyptus plantations have a great economic and environmental importance in Brazil. Considering the climate change scenario and the cumulative precipitation deviations observed in these plantation areas in recent years, it is essential to understand how plants respond to stressful conditions. Drought is one of the main factors limiting plant yield and survival. Plants under water deficit can increase their reactive oxygen species content culminating in cell death. Under such conditions, the biosynthesis of flavonoids is induced, suggesting their important role as an auxiliary antioxidant system. This system is very important in plants under severe or multiple stresses once it can help these organisms cope with the given situation. In this work, leaves of two commercial clones (AEC144 and IPB1) of Eucalyptus urophylla under water restriction and rehydration were evaluated in terms of physiological and biochemical parameters. In addition, flavonoids profiles were analyzed combining a flavonoid enrichment extraction method and LC-MS/MS approach. Water limitation provoked alterations in relative water content and in lipidic peroxidation. On the other hand, alterations in these two parameters were not significant when both rehydrated and control plants were compared. When plants under distinct water conditions and genotypes were compared (by volcano plot - MetaboAnalyst), we found differentially abundant metabolites (DAMs). The exact mass of these DAMs corresponding to the target flavonoids (quercetin, luteolin, kaempferol, apigenin and their derivatives). At this moment, we are working on the structural elucidation of these target metabolites by MS/MS.
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

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-430 Large-scale screening of carotenoid and chlorophyll content in tomato cultivars by using the simple, rapid and quantitative method

PRESENTING AUTHOR: Yusuke Aono, University of Tsukuba, Japan

CO- AUTHORS: Yonathan Asikin, Ning Wang, Harry J Klee, Miyako Kusano

Tomato is one of the highest-value crops worldwide. Modern tomato cultivars head to add commercially attractive factors, including flavors. Wild tomato species, e.g., Solanum lycopersicum ver. cerasiforme and S. pimpinellifolium, have a great potential to produce useful volatiles because of their large genomic diversities. Previous study revealed that presence of volatiles in tomato fruits can be improved for taste of sweetness and sourness. Furthermore, it contributes to add favorite flavors for tomato consumers. Of these, apocarotenoid volatiles, which are one of the carotenoid derivatives, are enzymatically cleaved by carotenoid cleavage oxygenases. Additionally, these volatiles have a large impact on the perception of flavor. Therefore, it is important to investigate novel factors in the biosynthetic pathway for production of apocarotenoid volatiles. However, it remains unclear what kinds of key enzymes and/or transcription factors contributing to produce apocarotenoid volatiles that have useful aroma. In this study, we focused on the extent of chemical diversities in tomato cultivars to possess great potential to produce useful volatiles derived from carotenoid pathway. To achieve it, we developed the high-throughput screening method to quantify important pigments, i.e., lycopene and total carotenoid by using microplate reader. We also quantified chlorophylls because it is expected that we could be predict tomato ripening stages. We assayed 362 tomato cultivars including S. lycopersicum, S. lycopersicum ver. cerasiforme, and S. pimpinellifolium by using our developed system. We will present what kinds of tomato cultivars contain specific content of total carotenoids, lycopene and chlorophylls in the assayed tomato fruits.

P-431 Creating the DynLib Mass Spectrometry Database of the Maize Secondary Metabolome using the RDynLib package

PRESENTING AUTHOR: Sandrien Desmet, VIB, Belgium

CO- AUTHORS: Yvan Saeys, Rebecca Dauwe, Hoon Kim, Geert Goeminne, Ruben Vanholme, John Ralph, Kris Morreel, Wout Boerjan

The importance of maize for the production of food and industrially relevant products, e.g. bio-ethanol, as well as the enormous genetic diversity, a vast collection of mutant stocks and the high degree of genomic collinearity with other cereal crops, has led to the construction of several gene/transcript and protein databases, such as gramene and MaizeGDB. However, Mass Spectrometry (MS) spectral databases of especially the secondary metabolome of maize are lagging behind. Here, we present the DynLib MS spectral database for maize that was generated using different tissues (stem, leaf, ear, tassel and cob). The database contains both Quadrupole-Time-of-Flight-based MS/MS and Fourier Transform-Ion Cyclotron Resonance-Ion Trap-based MSn spectra from both the known and unknown Liquid Chromatography (LC)-profiled compounds. The RDynLib package was used to associate MS/MS and MSn spectra belonging to the same compound. Implementing RDynLib into our structural elucidation pipeline led to the characterization of 428 compounds. Based on this set of characterized compounds, we putatively annotated the maize- and tissue-specific conversions that are responsible for the prevailing mass differences that were picked from the LC-MS data.

P-432 Study on anti-depressive constituents and mechanism of Herbal medicine, Chai-hu Shu Gan San

PRESENTING AUTHOR: Zhong-Mei Zou, Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences, China

CO- AUTHORS: Zhong-Mei Zou, Meng Yu, Hong-Mei Jia

It is universally acknowledged that the complexity of TCMs makes the identification of the chemical constituents related to the efficacy and definition of their mechanism of action challenging. Novel approaches are in great demand to provide deeper mechanistic insight into the clinical valuable effects. Metabolomics, as one of the 'omics' technologies of systems biology, is the comparative analysis of metabolites and their dynamic flux associated with the response of living systems to pathophysiological stimuli or genetic modification. By means of advanced analytical tools, including nuclear magnetic resonance (NMR), and mass spectrometry (MS), in conjunction with the multivariate data analysis (MVA), metabolomics approach has been extensively applied in many areas such as diagnosis and treatment of disease, drug toxicity, biomarker discovery, and exploration of pathogenesis. Chai-hu Shu Gan San Chaihu-Shu-Gan-San (CSGS), a traditional Chinese medicine (TCM) formula containing seven herbal medicines, has been used in treatment of gastritis, peptic ulcer, irritable bowel syndrome and depression clinically. This presentation will focus on investigation into its active constituents and mechanism of anti-depressive effect by using LC-MS/MS and HNMR metabolomics.
*AWARD WINNERS

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-433

EcoMetEoR - An enlightning platform for eco-metabolomic research

PRESENTING AUTHOR: Henriette Uthe, Friedrich Schiller University Jena, Germany

CO-AUTHORS: Alexander Weinhold, Fredd Vergara, Nicole M. van Dam

EcoMetEoR - An enlightning platform for eco-metabolomic research. Eco-Metabolomics is a relatively new research discipline and describes the application of metabolomics techniques to ecology with the aim to characterize biochemical interactions of organisms across different spatial and temporal scales 1. Changes in metabolite concentrations can provide mechanistic evidence for biochemical processes that are relevant at ecological scales. These include physiological, phenotypical and morphological responses of plants and communities to environmental changes and interactions with other organisms, too. Although, eco-metabolomics is a powerful tool to combine extrinsic ecological research and intrinsic biochemical research, biodiversity researchers are often limited in applying metabolomics due to lack of facilities and expertise. In addition, ecological research, especially in the field comes along with challenging sample quality and data analysis. To overcome this lack the EcoMetEoR platform uniquely focuses on supporting and teaching ecologists and biodiversity researchers to implement metabolomics analyses in their research programs. This support covers the experimental design, the access to the analytical equipment, sample preparation and data analysis and teaching during workshops and individual research stays.

P-434

Metabolomics as a Discovery Tool for Bioprospecting and Detection of Defense Compounds During Fungal Infection of Spruce Wood

PRESENTING AUTHOR: Marie-Pierre Pavageau, Thermo Fisher Scientific, France

CO-AUTHORS: Anas Kamleh, Marit Almvik, Nina Elisabeth Nagy, Hans Ragnar Norli, Ari Hietala, Sven-Roar Odenmarck, Monica Fongen, Claire Dauly

Every fourth tree in the Scandinavian forest is infected with rot. Root and butt rot fungi attack trees through the root and successively grow into the trunk degrading the core wood. The infected log can no longer be used as construction materials, damaging its value. We have employed tree metabolome analyses to differentiate and identify defense metabolites in Norwegian Spruce for new fungicide discovery as well as discovery of novel pathways that could be targeted by pesticides. Wood cores from healthy and white rot infected (Heterobasidion parviporum) spruce trees were sampled at trunk heights 0, 1.6 and 3.2 meters and from different zones of the core. A total of 73 samples were extracted and analyzed using LC connected to Q-Exactive(TM) and Orbitrap-IDX(TM) Mass spectrometers. Data were processed using Compound Discoverer(TM) software which allows the pre-processing and statistical analysis of data within the same platform as well as including tools for compound identification such as database and library matching. Compound identification was carried out by matching MS2 spectra and retention time to authentic standards, when feasible, or were tentatively annotated based on molecular formula search against compound databases or MSMS matching against mzCloud spectral library. We also looked at compounds that displayed high abundance in the fungal infected core wood as opposed to the reaction zone and found a new compound in spruce with composition C12H8O7, tentatively identified as purpurogallin-4-carboxylic acid, which we believe is a product from the fungal detoxification of wood defense compounds.

P-435

Metabolomic studies on conservation in response of cereal secondary metabolism to fungal infection

PRESENTING AUTHOR: Natalia Witaszak, Institute of Plant Genetics Polish Academy Sciences, Poland

CO-AUTHORS: Anna Piasecka

Several Fusarium spp., are economically devastating pathogens of cereals causing Fusarium head blight (FHB) which leads to reduction in crop yield world-wide. In addition, production of mycotoxins in cereal grain can be harmful to animals and humans. Elaboration of strategies for engineering resistance in cereal crops has to be boost by detailed knowledge about plant response against FHB at the molecular level. Untargeted metabolomics based on LC-MS was applied for studying the conservation of plant immunity response on pathogen infection at metabolomic level among economically important crops barley and wheat as well as taxonomically related model plant Brachypodium distachyon. MZmine2, MetaboAnalyst and MarVis software were used for defining the immunity-related secondary metabolites. Response of metabolomics signals was highly species-specific. Only small percent of differentially accumulated metabolites (DAMs) were common for all studied species. However, conservation of cereals immune system manifested in the similarity of metabolic pathways responding to infection. The most significant changes were observed for compounds annotated to metabolic pathway of tryptophan, especially in Brachypodium. In addition, metabolism of alkaloid gramine and polyamines was re-programmed during infection in all studied plants. Phenylpropanoids, mainly ferulic and p-coumaric acids and its conjugates with polyamines were the most numerous DAMs common among species. LC-MS - based metabolomics proved to be excellent tools for studying the basics of plant immunity reactions to fungal pathogens in closely related species. This study was supported by the National Science Centre grant Sonata 2015/17/D/NZ9/03347
*AWARD WINNERS

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-436

Effect of growing conditions on the tomato root exudate and its associated root microbiome

PRESENTING AUTHOR: Bora Kim, University of Amsterdam, Netherlands

CO-AUTHORS: Bora Kim, Marco Bentlage, Benjamin Thiombiano, Harro Bouwmeester, Anouk Zancarini

The host-microbiome interaction is important for host functions and health. While many endeavors were made to understand how the host shapes its microbiome in the human gut, the effect of plant metabolite exudation on the rhizosphere and root microbiome is still unclear due to the technically challenging root exudate collection and analysis. Indeed, plant root exudates are mostly studied under artificial conditions, such as in hydroponics, to avoid unwanted contaminants from natural soils. However, several questions are often arising with regard to the similarity of root exudate profiles under artificial and natural conditions. To answer these questions, we compared plant root exudate composition under aeroponics and two natural soil conditions using an untargeted metabolomics approach. The results showed that up to 29% of compounds in the root exudate were quantitatively different depending on the growing condition indicating that this has a significant impact on the root exudate composition. In addition, we also analyzed the corresponding root and rhizosphere bacterial communities using 16S rRNA metabarcoding analysis. The advanced statistical analyses used to link the root exudate profile to the microbiome composition to find molecular mechanisms involved in microbiome recruitment by plants will be discussed.

P-437

Sting nematodes modify metabolomic profiles of host plants

PRESENTING AUTHOR: Denis Willett, Cornell University, United States

CO-AUTHORS: Camila C. Filgueiras

Many nematode endoparasites of plants have sophisticated means of modifying host metabolomes. Root knot and cyst nematodes can create sophisticated feeding cells that co-opt host cellular machinery to upregulate amino acid and sugar production. Given their lifecycle and long history of co-evolution with hosts, it makes some sense that endoparasitic plant nematodes can modify host metabolomes. Here, we show that, based on global metabolomic profiling, nematode ectoparasites of plants can also modify host plant metabolomes. Susceptible, moderately tolerant, and tolerant varieties displayed distinct metabolomic profiles in response to sting nematode feeding. Specifically, sting nematodes suppress amino acids in susceptible varieties. Upregulation of compounds linked to plant defense have negative impacts on sting nematode populations. Pipecolic acid, linked to systemic acquired resistance induction, seems to play a large role in protecting tolerant varieties from sting nematode feeding and could be targeted in breeding programs.

P-438

Metabolomic profiling defines key signals and antibiotics in drought-induced resistance to Cochliobolus heterostrophus

PRESENTING AUTHOR: Shawn Christensen, USDA-ARS, United States

CO-AUTHORS: Casey A. Chamberlain, Charles Hunter, Anna Block

In nature, plants are simultaneously challenged by multiple forms of biotic and abiotic stress that combine for diverse effects on crop production. Here we examined disease resistance under drought stress conditions and observed that the magnitude of fungal-elicit ed maize responses are quantitatively dependent on the duration of drought prior to inoculation (DPI). Comparative analysis of watered and drought stressed Cochliobolus heterostrophus-infected plants using metabolomic fingerprinting resulted in complete multivariate separation of the two global metabolomes with 2,367 significant molecular features. Among these features, phytohormones, amino acids, sugars, and oxylipins were strongly elicited by C. heterostrophus infection in plants undergoing DPI, including the drought responsive signal 12-oxo-phytodienoic acid. The phytohormone abscisic acid predictably increased in response to drought stress but was curiously suppressed in C. heterostrophus-infected tissues vs. damaged controls. Examination of ent-kaurene-, β-macrocarpene- and benzoxazinoid-related antibiotics in fungal-elicit ed tissues displayed drought-dependent increases in production over an eight-day time course. Transcript accumulation of the 1,3-β-glucanase PR6mb, pathogenesis-related 4 (PR4), and chitinase genes also demonstrated positive relationships between pathogen elicitation and the duration of DPI. Collectively, our results indicate that drought stress potentiates maize defense mechanisms, contributing to heightened resistance against the common maize pathogen C. heterostrophus.
**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

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-439 Adaptive metabolomes: defense-related reprogramming in Sorghum bicolor

PRESENTING AUTHOR: Ian Dubery, University of Johannesburg, South Africa

CO-AUTHORS: Charity Mareya, Fidele Tugizimana

Background: Burkholderia andropogonis is the causal agent of bacterial leaf stripe, one of the three major bacterial diseases affecting Sorghum bicolor. However, the biochemical aspects of the pathophysiological host responses are not well understood. An untargeted metabolomics approach was designed to understand molecular mechanisms underlying S. bicolor–B. andropogonis interactions. At the 4-leaf stage two sorghum cultivars (NS 5511 and NS 5655), differing in disease susceptibility/resistance, were infected with B. andropogonis, and the metabolic changes monitored over time. Results: The NS 5511 cultivar displayed delayed signs of wilting and lesion progression compared to the NS 5655 cultivar, indicative of enhanced resistance. The metabolomics results identified statistically significant metabolites as biomarkers associated with the sorghum defence. These include the phytohormones salicylic acid, jasmonic acid and zeatin. Moreover, metabolic reprogramming in an array of chemically diverse metabolites that span a wide range of metabolic pathways was associated with the defense response. Signatory biomarkers included aromatic amino acids, shikimic acid, metabolites from the phenylpropanoid and flavonoid pathways, as well as fatty acids. Enhanced synthesis and accumulation of apigenin and derivatives thereof, was a prominent feature of the altered metabolomes. Conclusions: The analyses revealed an intricate and dynamic network metabolic pathways and metabolites comprising the sorghum defence arsenal towards B. andropogonis in establishing an enhanced defensive capacity in support of resistance and disease suppression.

P-440 Isotope assisted profiling of plant defense metabolites

PRESENTING AUTHOR: Maria Doppler, IFA-Tulln, University of Natural Resources and Life Sciences, Vienna (BOKU), Austria

CO-AUTHORS: Christoph Bueschl, Bernhard Kluger, Rainer Schuhmacher

Application of classical LC-MS based untargeted metabolomics workflows for the comparison of control and stress-treated plants results in the detection of numerous defense related metabolites. Although a large number of metabolites can be detected and classified to be of biological interest, the majority remains unknown or unidentified, which complicates biological interpretation. With our stable isotope labeling workflow we are not only able to effectively and reliably filter biologically relevant metabolites from background compounds, but also to obtain the number of labeling atoms (e.g. C and N) for all detected metabolites. Furthermore, the application of isotopically labeled tracers allows us to detect submetabolomes that only consist of metabolites descending from the applied tracer (i.e. a biological pathway). In the presented study, global-labeling as well as tracer-fate approaches were applied and combined in order to investigate the metabolic response of flowering wheat plants upon treatment with the mycotoxin deoxynivalenol. The detected phenylalanine-derived submetabolome consisted of 172 metabolites and up to 30% of them were classified as defense related. The substance class of hydroxycinnamic acid amides (HCAAs) turned out to be highly involved in the plants defense response as many of these metabolites' abundances were significantly increased under deoxynivalenol stress conditions. Based on these representative metabolites, the potential of SIL approaches for the annotation and identification of unknown metabolites is demonstrated for sum formula generation, database search and further characterization on the fullscan as well as on the MS2 level.

P-441 Comparative host-parasite and within-colony metabolomics: Cerithideopsis californica trematode guild

PRESENTING AUTHOR: Zdenek Kamenik, Institute of Microbiology, Czech Republic

CO-AUTHORS: Daniel Metz, Ryan Hechinger, Martin Kurecka, Petr Marsik

Despite the vast diversity of trematodes and their medical importance, we still lack basic knowledge about the biology of these parasites in their first intermediate host. The fact that these parasites are 'phenotype hijackers' (parasitic castrators), coupled with the recent discovery of social organization in some trematode species, makes the gastropod-trematode host-parasite system an attractive target for studies leveraging the power of comparative metabolomics with the experimental tractability of these organisms. In this study, we used the potamid snail Cerithideopsis californica and selected trematodes from the diverse guild exploiting it to answer three questions using untargeted mass spectrometry-based metabolomics studies. Specifically, we investigated (1) mechanism by which the parasites directly sense changing conditions outside of their host; (2) the differences between the metabolomes of uninfected and infected snails; (3) the possibility that soldier trematodes release an alarm pheromone to recruit other soldiers to the defense of the colony.
*AWARD WINNERS

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-442 Novel sources for Striga resistance: looking toward the rhizosphere

PRESENTING AUTHOR: Benjamin Thiombiano, University of Amsterdam, Netherlands

CO-AUTHORS: Dorota Kawa, Aimee Walmsley, Hanna Vahldick, Siobhan Brady, Harro Bouwmeester

Striga species are roots-parasitic plants widely distributed and able to infest rice, maize, sorghum and different other crops. S. hermonthica is considered the most serious worldwide parasitic weed, with an estimated affected area amounting to many millions of hectares. Aside of intercropping, fertilization, Striga low germinant genotype, soil microbiome has also been explored. Our preliminary data are suggesting that microbiome may lead to a decrease of Striga infestation rate. To better understand the mechanisms of the microbes mediated decrease of Striga infestation rate, evaluating the intercommunication between plant and the microbe through sorghum roots chemical composition and roots exudates chemical profile is needed. Here, we compared root exudates of sorghum under four different growing conditions (i.e. gamma radiated soil in presence and absence of Striga and non-gamma radiated soil in presence and absence of Striga). We analyzed roots and root exudates with a metabolomics approach and assessed phenotypic aspects such as Striga germination and attachment rate. We showed that a Striga attachment was reduced in presence of microbes. Using multivariate analysis, we observed a significant effect of the growing conditions on the root and roots exudate chemical composition. Further advanced statistical analyses will be used to link the roots and root exudate chemical composition to the striga infestation rate in order to find the mechanisms involved in microbiome microbes mediated decrease of Striga infestation rate.

P-443 Metabolomics studies of rhizosphere metabolites from 26 accession of Arabidopsis thaliana involve in plant interaction with rhizosphere microorganism

PRESENTING AUTHOR: Kourosh Hooshmand, Aarhus University, Denmark

CO-AUTHORS: Enoch Narh Kudjordjie, Rumakanta Sapkota, Tong Shen, Oliver Fiehn, Mogens Nicolaisen, Inge S Fomsgaard

Plants may regulate and selectively choose distinct members of a microbial community in the rhizosphere. The belowground plant-microbe association is facilitated by exudation of a wide range of bioactive chemical compounds from the root which may lead to indirect plant resistance against the potential biotic challenges. However, the underlying mechanism enabling the plants to shape its root microbiome structure has been poorly elucidated. In this study, 26 accessions of Arabidopsis thaliana (wild types) were grown in soil, harvested, lyophilized and extracted. The chemical composition of primary metabolites, secondary metabolites and lipids were determined by employing GC-TOFMS, HILIC-TripleTOF MS/MS, and reverse phase-Q Exactive HF mass spectrometer (CSH, lipidomics) respectively. GC-MS data was processed by BinBase and HILIC and CSH data were proceed by MS-DIAL_2.94 followed by post-processing with MS-FLO. The bacterial and fungal composition was determined by Illumina MiSeq sequencing technique. Carbohydrate, amino acids, and fatty acids were the main primary metabolites present in all the accessions. Carboxylic acids, organooxygen, and benzene derivatives were also predominated polar metabolites classes, as well as fatty acyls, glycerophospholipids, and sphingolipids were the core lipids in all the accession. Various statistical methods will be employed to identify the unique chemical substrates from the root of each A. thaliana accessions associated with regulation of distinct members of beneficial rhizosphere microbes which can lead us into a more sustainable plant production.

P-444 Metabolomics to exploit the primed immune system of tomato fruit

PRESENTING AUTHOR: Pierre Pétriacq, INRA & University of Bordeaux, France

CO-AUTHORS: Estrella Luna, Amélie Flandin, Cédric Cassan, Chloé Chevanne, Camélia Feyrouse Kadiri, Yves Gibon, Pierre Pétriacq

Tomato is a major crop suffering substantial yield losses from diseases as fruit decay at a postharvest level can claim up to 50% of the total production worldwide. Due to the environmental risks of fungicides, there is an increasing interest to exploit plant immunity through priming, which is an adaptive strategy that improves plant defensive capacity by stimulating induced mechanisms. Broad-spectrum defence priming can be induced by the compound β-aminobutyric acid (BABA). In tomato plants, BABA induces resistance against various fungal and bacterial pathogens and different methods of application result in durable protection. Here, we examined whether treatment of tomato plants with BABA resulted in a durable induced resistance in tomato fruit against Botrytis cinerea, Phytophthora infestans and Pseudomonas syringae. Targeted and untargeted metabolomics were used to investigate the metabolic regulations that underpin priming of tomato fruit against pathogenic microbes that present different infection strategies. Assessment of infection showed a statistically significant reduction of disease symptoms in tomato fruit that originated from BABA-primed plants, thus confirming the broad effectiveness of BABA priming against various pathogens. Metabolomic analyses revealed major changes after BABA treatment and after infection. Remarkably, primed responses depended entirely on the type of infection, rather than showing a common fingerprint of BABA-induced priming. Altogether, our results demonstrate that metabolomics is particularly insightful towards a better understanding of defence priming in fruit. Further experiments are underway in order to identify key metabolites that mediate broad-spectrum BABA-induced priming in tomato fruit.
*AWARD WINNERS

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-445

Metabolomic responses of tomato fruits and roots according to magnesium excess

PRESENTING AUTHOR: Min Cheol Kwon, Konkuk University, South Korea

CO-AUTHORS: Eun Sung Jung, Yangmin X. Kim, Seulbi Lee, Choong Hwan Lee

Green house cultivation uses excessive amounts of fertilizer due to the characteristics of intensive cultivation, causing nutrient salts that affect plant growth and development. Tomato is an important economic crop that is continually threatened by salt stress. To explore the effect of magnesium excess on metabolites of tomato plants, we performed non-targeted metabolite profiling. Tomato was cultivated under three different conditions, i.e. control, high magnesium stress (MgH), and extremely high magnesium stress (MgEH), and fruits and roots were harvested. In principal component analysis and heat map of fruit and root data, metabolite changing patterns of Mg stress showed different patterns in fruits and roots. Tomato fruit metabolic variations by Mg stress included increases of amino acids, flavonoids and amines and decreases of organic acid, sugar and lysophospholipids. Moreover, the highest anti-oxidant activity of tomato fruits was observed in MgEH group, which was positively correlated to the levels of flavonoids and amines. Also, the decrease of fresh weight per fruit caused by MgEH was positively related to the variations of organic acids and sugars. In tomato roots, MgEH decreased amino acids, organic acids and lysophospholipids. Taken together, our results revealed that tomato fruits and roots responded differently in response to magnesium excess in terms of amino acids, which increased in fruits and decreased in roots.

P-446

Stable isotope labelling and omics approaches to study the spatiotemporal regulation of diterpenoid resin acids biosynthesis in Norway spruce

PRESENTING AUTHOR: Monica Scognamiglio, Max Planck Institute for Chemical Ecology, Department of Biochemistry, Germany

CO-AUTHORS: Felix Feistel, Erica Perreca, Christian Paetz, Jonathan Gershenson, Axel Schmidt

Many conifer species produce a complex mixture of terpenoids, known as resin, acting both as a mechanical and as a chemical barrier against herbivores and pathogens. Although the biosynthetic pathways of the resin components are known, the way some species like Norway spruce (Picea abies) regulate their biosynthesis and accumulation is still not well understood, yet very important in helping elucidating resin's biological and ecological significance. In order to gain new insights into the spatiotemporal regulation of the biosynthesis of diterpenoid resin acids (the main components of resin besides monoterpenes), P. abies saplings were placed in presence of 13CO2 during their growing season in an in-house built labelling chamber under controlled environmental conditions. The 13C enrichment was monitored over time. Thanks to a combination of metabolomics and target chemical analyses, it was possible to obtain information not only on the resin components' biosynthesis during the year, but also on the associated orchestration of plant metabolism. The results suggest that these compounds are synthesized and accumulated very early during the development of new growing tissues, while no turnover was observed in old tissues. The chemical composition of the resin mixture is highly regulated and organ/tissue specific. The identity and relative abundance of the different chemicals are very conserved in the branches, while they vary broadly and change over time in needles. In conclusion, the used experimental approach helped us to shed light on important aspects of resin biosynthesis and accumulation and showed a fine tuning of both phenomena in Norway spruce.

P-447

Elucidation of metabolic reprogramming that characterizes the plant growth promoting rhizobacteria priming of tomato plants

PRESENTING AUTHOR: Msizi Mhlongo, University of Johannesburg, South Africa

CO-AUTHORS: Fidele Tugizimana, Lizelle Piater, Ian Dubery

Plant growth promoting rhizobacteria (PGPR) are beneficial microbes found in the rhizosphere that directly or indirectly stimulate plant growth. Some PGPR can prime plants for enhanced defense against a broad range of pathogens and insect herbivores. In this study, four PGPR strains (Pseudomonas fluorescens N04, P. koreensis N19, Paenibacillus alvei T19 and Lysinibacillus sphaericus T22) were used to induce priming in Solanum lycopersicum (cv Moneymaker) plants. Seedlings were inoculated with an overnight culture of four PGPR strains and the different plant tissues (roots, stems and leaves) were harvested at 24 h and 48 h post-inoculation. Metabolites were extracted with methanol and analysed by UHPLC-MS/MS. Chemometrics methods were applied to mine the data and characterize the differential metabolic profiles induced by these PGPR strains. The results showed that the four PGPR strains induced a defence related metabolic reprogramming in tomato plants, characterized by changes in hydroxycinnamates, benzoates, flavonoids, amino acids and fatty acids. These metabolic alterations point to a preconditioned state that renders the plants primed for an enhanced defense responses. Thus, these results contribute to ongoing efforts in investigating and unravelling the biochemical processes that define the priming phenomenon.
**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

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-448

Metabolomics and Machine Learning Techniques applied to investigate beneficial Plant-Bacteria Interactions

PRESENTING AUTHOR: Lena Fragner, University of Vienna, Austria

CO-AUTHORS: Florian Schindler, Johannes Herpell, Anke Bellaire, Weimin Li, Xiaoliang Sun, Wolfram Weckwerth

Endophytic non-pathogenic colonization of plant tissue by bacteria is a well-known and wide spread phenomenon, even expected to be the case for all angiosperms. Symbiotic plant-bacteria interactions comprise various levels of obligations and ecological benefits to at least one of the partners. The beneficial effects and functions for plants are manifold, including enhanced stress resistance, plant growth promotion or capacity for controlling plant-pathogens. In the present work, we focus on obligatory and constant symbioses occurring in the plant families Rubiaceae, Primulaceae and Dioscoreaceae. Highly specialized bacterial symbionts are mainly host-specific, often not cultivable and their absence can lead to dwarf phenotypes of the host-plants. Endophytic bacteria of leaves can be evenly distributed between the mesophyll cells or accumulated in specific leaf areas or in specialized structures. Most studies focused on genome and proteome analyses suggesting potential alterations of secondary metabolism caused by the presence of beneficial symbionts. However, detailed mechanisms and functions of these highly specialized mutualistic plant-bacteria symbioses are not yet fully understood. In the present study we investigate alterations in the metabolome of colonized leaf tissue. Primary and secondary metabolites were analyzed by GC-MS and LC-MS respectively, complemented with physiological and morphological data, and analyzed with machine learning techniques. Results indicate distinctive mechanisms of the symbiosis in investigated beneficial plant-bacteria interactions and will be discussed in detail.

P-449

Metabolic Profiling of Benzoxazinoids in Weed Suppressive and Early Vigour Wheat Genotypes

PRESENTING AUTHOR: Leslie Ann Weston, Charles Sturt University, Australia

CO-AUTHORS: James M. Mwendwa, Paul A. Weston, Inge Fomsgaard, William B. Brown, Jeffrey D. Weidenhamer, Leslie A. Weston

Wheat cultivar trials were conducted in moderate to low rainfall zones at Wagga Wagga and Condobolin NSW from 2014-2016. Crop and weed growth was monitored at tillering, vegetative, grain filling, harvest and post-harvest stages. Wheat roots, shoots, rhizosphere and bulk soils were collected over time for metabolomic profiling and extracted in methanol using an automated Buchi high pressure extractor while soil samples were extracted for 24h by rotary shaker. Non-targeted analysis by UPLC-ESI MS QTOF (Agilent 6530) was performed and data analysed using Mass Profiler Professional (Agilent). Benzoxazinoids (BXs), including potent allelochemicals, were profiled using targeted analysis in negative ion mode while key bioactive phenoxazinones (microbially-produced APO, AAPO, AAMPO) were profiled in positive ion mode. Detection of up to 20 individual BXs including glycosides, lactones, hydroxamic acids and related microbial metabolites were noted. Both qualitative and quantitative differences in BXs were observed and were cultivar-, growth stage- and location-dependent. Plant part and rhizosphere location (distance from root) also impacted BX concentration. Microbially-produced metabolites with phytotoxic activity were detected at ecologically relevant concentrations in roots and rhizosphere soils. Metabolic profiling provided critical knowledge of seasonal impacts on wheat metabolism, as well as the biosynthesis and release of metabolites associated with weed suppression. Certain wheat cultivars, including heritage and recent commercial accessions, maintained high yield potential and were significantly more weed suppressive, depending on year and location, potentially due to their vigorous early growth habit and canopy architecture as well as the release of BXs and phenoxazinones into the rhizosphere over time.

P-450

The honey bee pollen diet investigated by MS and NMR based metabolomics

PRESENTING AUTHOR: Nanna Hjort Vidkjaer, Aarhus University, Department of Agroecology, Denmark

CO-AUTHORS: Jane Ward, Per Kryger, Inge S. Fomsgaard

Declining honey bee (Apis mellifera, hereafter bees) populations are concerning because bees are important pollinators of food crops. The decline is hypothesized to be driven by multiple factors (1), and bees face many stressors including pathogens, xenobiotics and changes in floral resources affecting their diet. Bees consume pollen containing essential nutrients and multitudes of bioactive plant secondary metabolites (PSMs). Recent findings demonstrate the potential of PSMs to affect bee health e.g. by reducing virus loads (2), but the PSM profile of pollen is sparsely investigated. Different floral resources are available in different environments and throughout the season, but limited knowledge exist on how such variations influences dietary composition. In a Danish field experiment, biweekly pollen samples (hive entrance and fermented pollen stores) were collected from four apiaries located in different environments (agricultural, urban, forest, and meadow). Chemical profiles of the two pollen types were generated by direct infusion ESI-MS, NMR, and HPLC-HRMS. Multivariate data analysis was subsequently used to explore seasonal and landscape variations in diet composition. The results clearly demonstrated differences in the composition of the pollen originating from the apiaries in the four environments. Within each environment, the pollen composition gradually changed throughout the season. Collectively, the results create a knowledge base for future studies of dietary effects on bee health and allows for novel insight into the chemical fate of PSMs upon hive storage and fermentation prior to consumption. 1. Goulson, D. et al. Science 2015, 347:1435. 2. Palmer-Young, E.C. et al. J Econ Entomol 2017, 110:1959.
**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

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-451

From Field to Feature: A global workflow for metabolic fingerprinting in grassland communities

PRESENTING AUTHOR: Susanne Marr, IPB Halle, MLU, iDiv, Germany

CO-AUTHORS: Jos Hageman, Kristian Peters, Nicole van Dam, Helge Bruelheide, Steffen Neumann

Plants, grown in fields, face an undefined number of stresses. How plants respond to these factors, is often studied on the level of visible traits. However, changing communities and seasons will also be reflected in the metabolic fingerprint of a plant. Understanding those dynamics will help to unravel underlying mechanisms of ecosystem functions, such as herbivore resistance. One primary challenge that scientists face when bringing together high-throughput analytical tools with broad ecological questions is the lack of standardised methods that would allow for the automated analyses of metabolomic data within complex experimental designs. We aim to design a global workflow enabling large-scale ecological experiments including many diverse plant metabolomic profiles to be analysed and compared within one dataset. We analysed secondary metabolites – well known to play key-roles in plant defence strategies – in the aboveground tissue of 13 grassland species. In the Jena Experiment, plants were sampled from different communities, consisting of 1 to 8 different species. The plants were collected at four time-points across the growing season to capture seasonal differences. Our workflow enables the automated processing and analysis of raw data, obtained from UPLC-MS spectra, across all species and all treatments simultaneously. We use the number and composition of features – compound-fragments defined by their specific mass and retention time – to describe the metabolic fingerprint. The fingerprints identified each species across the different sampling conditions. On the species level, we found different responses to plant diversity and season, indicating species-specific strategies of adaptation to environmental changes.

P-452

Jasmonate-mediated tomato fruit growth under supplemental LED inter-lighting revealed by metabolomics

PRESENTING AUTHOR: Ivan Paponov, Research Scientist, Norway

CO-AUTHORS: Martina Paponov, Dmitry Kechasov, Michel J. Verheul

Fruit growth and the accumulation of primary compounds, which determine the final fruit quality, are driven by the supply of water, assimilates, and nutrients to the fruits through the phloem and xylem. Consequently, both shoot and root activities contribute to fruit growth. The supplemental LED inter-lighting can modulate both shoot and root activities; therefore, we suppose that this supplemental lighting also can modulate the relative growth rate of tomato fruits during the light and dark periods. How the diurnal rhythm of fruit growth is affected by supplemental LED inter-lighting treatment and the mechanisms of its regulation are unknown and needs further investigation. To address these questions, we investigated the diurnal response in term of fruit relative growth rate and we analyzed the metabolite composition of xylem sap. Diurnal analysis of fruit growth showed that fruits on plants receiving LED light grew quicker during the night than did fruits of control plants. This larger fruit growth during the night was related to an increased root pressure: the main source of night-time water and nutrient transport into the fruits. LED treatment also increased the levels of the phytohormone jasmonate in the xylem. Supplemental LED inter-lighting increased tomato fruit weight by increasing the total assimilates available for fruit growth and by enhancing root activity through increases in root pressure and water supply to support fruit growth during the night.

P-453

Metabolomic responses of Oryza sativa (rice) to high humidity conditions

PRESENTING AUTHOR: Venea Dara Daygon, The University of Queensland, Australia

CO-AUTHORS: Melissa Fitzgerald

Abiotic stresses induce a cascade of metabolic responses in plants to help in adaptation and survival during unfavourable conditions. In some cases, these changes affect the eating quality of food crops. In our previous study, untargeted profiling of volatile compounds in rice has identified indole as a major contributor to unpleasant odour in rice grains and its production is highly affected by environmental factors. To understand the mechanism of indole production, eight japonica rice varieties were grown in different levels of humidity during grain filling stage. Targeted analysis of indole was performed using GC-MS in the matured rice grains and metabolite profiling of auxins and indole derivatives was performed using SPE UHPLC-MS on different stages of rice development. We further compared the expression of candidate genes in varieties with varying levels of indole production. In this study, we demonstrate that high moisture and humidity during the mid to late grain filling stages enhances indole production in susceptible varieties. Indole expression in the matured grains could be a result of an alteration in the tryptophan-dependent biosynthesis of indole-3-acetic acid synthesis during stress conditions.
*AWARD WINNERS

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-454 Metabolomics of Brachypodium – fungal pathogen interaction

PRESENTING AUTHOR: Anna Piasecka, Institute of Plant Genetics, Polish Academy of Sciences, Poland

CO-AUTHORS: Nicolas Jedrzejczak-Rey, Paweł Bednarek

Brachypodium distachyon (Bd) is one of the model systems to study plant-pathogen interaction in grasses. However, the immune function of Bd secondary metabolites is until now unexplored. Therefore, metabolomics of Bd carries great potential to contribute to fundamental knowledge on plant immunity in cereal crops. Untargeted LC-MS approach in combination with MZmine2, MetaboAnalyst and MarVis cheminformatics tools enabled us to study secondary metabolism of Bd during infection with Parastaganospora nodorum, which is economically devastating fungal pathogen of cereals. Our analysis revealed that the number of differentially accumulated metabolites (DAMs) 72 hours post infection (hpi) was three times greater than 24 hpi. Compounds from biosynthetic pathways of indoles, phenylpropanoids, terpenes and polyamines have been identified as DAMs among thousands of detected signals. The most significant changes in metabolite accumulation were observed for serotonin and its derivatives. For instance, dehydrodimer of serotonin was one of the most highly accumulated metabolites 48 hpi. Conjugates of serotonin with p-coumaric and ferulic acids were also identified as DAMs. In addition, tryptamine, which is a precursor of serotonin, also increased its accumulation during infection. These results indicate a tight correlation of tryptophan and hydroxycinnamic acid metabolism with Bd immune response to fungal pathogens. This study was supported by the National Science Centre grants: Sonata 2015/17/D/NZ9/03347 and Sonata Bis 2012/07/E/NZ2/04098.

P-455 Tracking the metabolic fate of 13C-labeled plant-derived allelochemicals in the environment

PRESENTING AUTHOR: Eva Knoch, Gregor Mendel Institute of Molecular Plant Biology, Austria

CO-AUTHORS: Niklas Shandry, Maria Doppler, Christoph Bueschl, Rainer Schuhmacher, Claude Becker

Many plant species release specialized metabolites into the soil that inhibit germination, growth, or development of neighboring plants. These substances are collectively known as allelochemicals, and the related process as allelopathy. A key aspect of allelopathy is the post-release chemical dynamics of these compounds in soil: while some of them are converted to biologically more active forms, others are degraded and lose their toxicity. An important factor in the turnover of allelochemicals in soil is the rhizospheric microbiome, the community of microorganisms that live on and around the plant roots. To understand the metabolic processes that occur after release of the allelochemicals, we take a deconstructed approach that uses plant-derived stable-isotope-labeled allelochemicals and isolated bacterial communities. 13C-labeled allelochemicals are produced by growing plants on 13C-glucose and purified by preparative HPLC. Purified labeled and unlabeled allelochemicals are added to the growth media of defined bacterial communities; we then monitor their metabolic conversion and degradation products by LC-HRMS and will try to identify novel biotransformation products with MS/MS. The product patterns will be correlated with transcriptomes from the bacterial communities to reveal molecular mechanisms of allelochemical turnover by soil bacteria. Here we present our results to date, with focus on the workflow for production of 13C-labeled allelochemicals benzoxazinoids from rye and sorgoleone from sorghum. 13C-enrichment and isotopologue patterns of the target compounds produced in seedlings have been determined by LC-HRMS. We find that the benzoxazinoids are almost fully 13C-labeled, which is a prerequisite for the use of them in our downstream application.

P-456 Metabolomics profiling in patients with non-specific acute low back pain after Thai Herbal Sahatsatara Formula administration

PRESENTING AUTHOR: Manmas Vannabhum, Center of Applied Thai Traditional Medicine, Mahidol University, Thailand

CO-AUTHORS: Pravit Akarasereenont, Manmas Vannabhum, Jantanee Wattanarangsan, Titchaphorn Palo, Kamontip Harnphadungkit, Pinpilai Jutasompakorn, Somruedee Chatsiricharoenkul

Thai Herbal Sahatsatara Formula (STF) is in the Thailand National List of Herbal Medicinal Products for the relief of muscle pain and numbness on hands and feet. STF contains 21 herbal ingredients. The previous study showed that STF reduced pain in patients with low back pain (LBP). However, there are no identifying of related chemical compounds. The aim of this study is to investigate metabolites profiling, using liquid chromatography-time of flight mass spectrometry (LC-TOF MS), in patients with non-specific acute LBP after STF administration. Twenty-eight patients were received STF 1,350 mg, 3 times daily, for 7 days. Plasma samples were collected before STF administration on day 0, and after STF administration on day 1, 4 and 7. The results showed different metabolomics profile at day 1 and 4 when compared to day 0. Interestingly, the pattern of the chemical profile was closely when compared at day 7 to day 0. The most identified compounds were lipids such as lysophosphatidylethanolamine (LPE), palmitoleic acid, linoleate, myristic acid suggesting anti-inflammatory action of STF. Further study on the metabolomics-phenomics relationship will help to understand the mechanism of action of STF.
**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

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-457 **Metabolomics as tool to study the role of semiochemicals in biotic plant interactions**

**PRESENTING AUTHOR:** Roland Mumm, Wageningen Plant Research, Bioscience, Wageningen University and Research, Netherlands

**CO-AUTHORS:** Ric C.H. de Vos, Robert D. Hall

In nature, plants are surrounded by a huge diversity of organisms with which they form a complex network of antagonistic (e.g., pests and pathogens) and mutualistic (e.g., pollinators, beneficial microbes) interactions. These interactions are tuned by an intense chemical communication between the different players mediated by numerous volatile and non-volatile compounds, so called semiochemicals. In this presentation we will give examples of metabolomics approaches as a tool in research towards the role of different types of semiochemicals involved in the chemical communication of plants, microorganisms and insects. We have applied untargeted comprehensive GC-MS and LC-MS profiling for volatile and non-volatile metabolites to elucidate which semiochemicals are important for attracting and repelling of insect pest in contrasting crop species including cabbage, hop, and bean species. In bioassays and field trials we show that compounds that are also valued for their flavour properties likely also play a role in the plant resistance/susceptibility against enemies. In a plant microbial system we show that volatile compounds emitted by certain soil bacteria can have inhibitory effects on the growth of fungal plant pathogens but in turn can also promote the growth of the plant. Once we have identified these key metabolites we can exploit them in follow up research focused upon developing new strategies for improving crop production.

P-458 **Metabolomic Approaches for the Identification of Flavonoids Associated with Weed Suppression in Sele**

**PRESENTING AUTHOR:** Sajid Latif, Charles Sturt University, Australia

**CO-AUTHORS:** Saliya Gurusinghe, Paul A. Weston, Jane C. Quinn, Leslie A Weston

Incorporation of competitive pasture legumes in conservation agricultural systems provides a non-chemical alternative towards integrated and sustainable weed management strategies. While the in-field weed suppressive potential of annual pasture legumes has been previously described, the mechanism of interference with weeds has not been clearly elucidated. We therefore aimed to delineate the role of secondary metabolites present in pasture legumes through a series of studies to: 1) characterize key metabolites present in plant tissues, and the rhizosphere and 2) correlate their presence with weed suppressive properties. In vitro experimentations were conducted to assess the phytotoxic potential of selected annual pasture legumes and targeted and non-targeted metabolic profiling was performed to evaluate the abundance of key metabolites using UHPLC QTOF-MS. Methanolic extracts and dried residues of Biserrula pelecinus L. and Ornithopus compressus L., but not Trifolium vesiculosum Savi., exhibited marked phytotoxicity in a series of laboratory experiments. Metabolic profiling revealed that both foliar tissues and rhizosphere soils of annual pasture legumes possessed a high abundance of various flavonoids and their precursors. Chemometric analyses suggested a clear association of quercetin, isoquercetin, kaempferol, and kaempferol-7-O-glucoside with phytotoxicity and weed suppression under field conditions. Specifically, the abundance of quercetin and kaempferol was significantly higher in soils collected from established stands of biserrula and yellow serradella in contrast to arrowleaf, gland and subterranean clover. Both field and laboratory experimentation provided evidence for the role of annual legume-produced flavonoids in weed suppression in southern Australia and further insight into their localization and release in the soil rhizosphere.

P-459 **Phytochemicals responsible for susceptibility to pest insects in forest species revealed by LC-MS based metabolomics approach**

**PRESENTING AUTHOR:** Jasna Valentina Campos, University of Concepcion, Chile

**CO-AUTHORS:** Sebastián Riquelme, Rosa Alzamora, Claudia Mardones, Rafael Rubilar, Andy J. Pérez.

The forest industry is of great importance for the Chilean economy and the Eucalyptus plantations corresponds to 35% of total planted surface. The two most preferred species by the foresters are E. globulus and E. nitens, but these trees face large biotic stressors. Such is the case of Gonipterus platensis Marelli (Coleoptera: Curculionidae) commonly known as Eucalyptus weevil, which is a defoliator insect native from southeastern Australia that feed on growing Eucalyptus foliage. The most susceptible species to this pest in Chile is E. camaldulensis and E. globulus, whereas E. nitens has been reported for its ability to resist or avoid the attack, fact that has also been confirmed by our research group in the field. In the present work a new approach based on non-targeted reverse phase LC-MS metabolomics analysis was performed to establish a qualitative difference between metabolomes of E. globulus and E. nitens leaves, species with different susceptibility to the insect attack. The multivariate principal component analysis (PCA) and partial lest-squares discriminant analysis (PLS-DA) were performed to investigate the overall variation in the metabolome. The PLS-DA analysis showed the specific metabolites for each species where the most contributor to this separation highly correlated to E. nitens were STILBENOIDS and GALLATE derived compounds that explained the chemical difference with E. globulus, these secondary non-volatile metabolites can satisfactorily explain most of the variation between both species, consider them as potential biomarkers and playing an important role for the host selection by insect.
**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

PLANT, FOOD, ENVIRONMENTAL AND MICROBIAL

P-460

Metabolic profiling approach to understand the immune defense activity in date palm under biotic stress

PRESENTING AUTHOR: Resna Nishad, Qatar University, Qatar

CO-AUTHORS: Talaat Ahmed

The most devastating diseases of date palm are results of necrotrophic fungal pathogens, Ceratocystis radicicola is one among them. C.radicicola is found all over Arabian Peninsula, it is a causative agent of black scorch, root rot, trunk rot and rhizosis. Induced biosynthesis of primary and secondary metabolite is a plant basal immunity mechanism against fungal pathogen, thus in the current study we followed metabolic approaches to study the interaction between date palm and pathogenic fungi. As metabolite production and extraction is more convenient in suspension culture than the whole living plant, we established suitable date palm suspension culture to study the host-pathogen interaction. In order to understand whether same or distinct host defense pathway is operating in both in vitro as well as in planta system we compared the metabolic changes in callus and in planta after exposing plant cell to C.radicicola elicitor and mycelia. Fatty acid profiling was analyzed in response to fungal elicitor and mycelia in callus and leaf respectively by using GC-MS. The fatty acid production was different in callus from the leaf thus the response towards biotic stress was entirely different in both systems. Even though the response were different, the fatty acid synthetic pathway found to be leading SA mediated and JA mediated immune responses. It concludes that date palm suspension system can use for pant-microbe interaction study. This work was made possible by GSRA grant GSRA2-1-0608-14021 from the Qatar national research fund (a member of Qatar foundation).

P-461

In-depth metabolomic profiling reveals genome plasticity in the plant pathogen Fusarium poae

PRESENTING AUTHOR: David Overy, Agriculture and Agri-Food Canada, Canada

CO-AUTHORS: Tom Witte, Amanda Sproule, Anne Hermans, Anne Johnston, Allen Xue, Linda Harris

Fusarium poae (Peck) Wollenweber is cosmopolitan, occurring on a range of hosts and associated with Fusarium Head Blight (FHB) in cereals. European and Asian isolates of F. poae have been reported to produce type A and type B trichothecenes as well as beauvericin, cyclonerodiol, and enniatins, but little is known about the mycotoxigenic potential of Canadian isolates. Historically, mycotoxin production associated with F. poae is quite variable. Several researchers hypothesize that the variation in mycotoxin expression observed in F. poae results from the presence of supernumerary chromosomes and the abundance of transposable elements (TEs) within its genome. Supernumerary chromosomes act as evolutionary cradles for pathogen virulence factors and transposon facilitated translocations into core chromosomes can accelerate genome evolution and therefore present a considerable challenge towards a durable disease management strategy. In depth metabolomic profiling and genome sequencing was carried out on 46 monosporic Canadian F. poae isolates demonstrating a degree of chemotypic diversity. Metabolomic profiling revealed consistent production of multiple "core genome" associated metabolites including the emerging mycotoxins diacetoxyscirpenol and beauvericin. Of particular interest, strain specific production of new mycotoxins were also observed and linked with horizontal gene transfer through supernumerary insertions into the F. poae genome.

P-462

An untargeted lipidomics MS-based approach to identify bioactive compounds of different yeast species associated to Drosophila suzukii

PRESENTING AUTHOR: Flavia Bianchi, Laimburg Research Centre, Italy

CO-AUTHORS: Urban Spitaler, Silvia Schmidt, Peter Robatscher, Daniela Eisenstecken

The spotted wing drosophila, Drosophila suzukii (D. suzukii) is an invasive vinegar fly native to Southern Asia, that has spread across USA and Europe, causing considerable economic damage in agriculture to a wide variety of fruits. Previous studies revealed that yeasts are attractive to D. suzukii, and they can influence the fitness of many Drosophila species being a food source, and favouring larval development, fecundity and oviposition. Lipids play a fundamental role in the diet of Drosophila, since they are the major energy storage molecules in cells, and necessary for egg production. Insect fat body contributes to the development, metamorphosis, and reproduction of the flies. In nature, Drosophila feeds on yeast growing on fruit, and both yeasts and plants constitute a lipid source. Yeast food is enriched in phosphatidylinositols, in contrast, plant food contains mostly triacylglycerols, and a more heterogeneous composition of phospholipid classes. Within the project DROMYTAL the metabolic profile of eight selected yeast species putatively associated to D. suzukii have been explored to develop a lure, based on the association of attractant and phagostimulant yeasts and an insecticide. Intracellular non-polar metabolites have been extracted after inoculation in liquid media (potato dextrose broth) for 30 hours. An untargeted approach in reversed-phase liquid chromatography-quadrupole/time-of-flight mass spectrometry (RPLC-QTOFMS) was used to detect diverse compound classes. Clear differences could be found among yeast species. Overall, about 90 distinct intracellular lipids have been annotated and/or identified, for further investigating which compounds play a bioactive role for the fitness of the insect.