Nontargeted and Targeted Metabolomics Identifies Dietary Exposure Biomarkers for Navy Bean and Rice Bran Consumption in Children and Adults.

Dietary exposure biomarkers are needed for advancing knowledge on healthy foods. This study examined biomarkers for navy beans and rice bran in children and adults. Plasma, urine, stool, and study foods from dietary intervention studies were analyzed by metabolomics. A total of 38 children and 49 adults were assessed after consuming navy beans and/or rice bran for 2-, 4-, 6-, or 12 weeks. From the 138-175 metabolites modulated by diet, 11 were targeted for quantification. Trigonelline and pipecolate concentrations increased in children and adult plasma after 4 weeks compared to baseline. Increased xanthurenate (46%) was observed in children plasma after rice bran intake for 4 weeks. Study foods with navy beans had higher S-methylcysteine compared to control and supported the increased urine S-methylcysteine sulfoxide. Nontargeted metabolomics was moderately effective to identify target molecules as candidate biomarkers. Study limitations include interindividual metabolite variations before diet intervention. Validation is warranted using cross-over designs and larger sample sizes.

Tissue-wide metabolomics reveals wide impact of gut microbiota on mice metabolite composition.

The essential role of gut microbiota in health and disease is well recognized, but the biochemical details that underlie the beneficial impact remain largely undefined. To maintain its stability, microbiota participates in an interactive host-microbiota metabolic signaling, impacting metabolic phenotypes of the host. Dysbiosis of microbiota results in alteration of certain microbial and host metabolites. Identifying these markers could enhance early detection of certain diseases. We report LC-MS based non-targeted metabolic profiling that demonstrates a large effect of gut microbiota on mammalian tissue metabolites. It was hypothesized that gut microbiota influences the overall biochemistry of host metabolome and this effect is tissue-specific. Thirteen different tissues from germ-free (GF) and conventionally-raised (MPF) C57BL/6NTac mice were selected and their metabolic differences were analyzed. Our study demonstrated a large effect of microbiota on mammalian biochemistry at different tissues and resulted in statistically-significant modulation of metabolites from multiple metabolic pathways (p ≤ 0.05). Hundreds of molecular features were detected exclusively in one mouse group, with the majority of these being unique to specific tissue. A vast metabolic response of host to metabolites generated by the microbiota was observed, suggesting gut microbiota has a direct impact on host metabolism.

LongITools: Dynamic longitudinal exposome trajectories in cardiovascular and metabolic noncommunicable diseases.

The current epidemics of cardiovascular and metabolic noncommunicable diseases have emerged alongside dramatic modifications in lifestyle and living environments. These correspond to changes in our "modern" postwar societies globally characterized by rural-to-urban migration, modernization of agricultural practices, and transportation, climate change, and aging. Evidence suggests that these changes are related to each other, although the social and biological mechanisms as well as their interactions have yet to be uncovered. LongITools, as one of the 9 projects included in the European Human Exposome Network, will tackle this environmental health equation linking multidimensional environmental exposures to the occurrence of cardiovascular and metabolic noncommunicable diseases.

Identification and Distribution of Sterols, Bile Acids, and Acylcarnitines by LC-MS/MS in Humans, Mice, and Pigs-A Qualitative Analysis.

Sterols, bile acids, and acylcarnitines are key players in human metabolism. Precise annotations of these metabolites with mass spectrometry analytics are challenging because of the presence of several isomers and stereoisomers, variability in ionization, and their relatively low concentrations in biological samples. Herein, we present a sensitive and simple qualitative LC-MS/MS (liquid chromatography with tandem mass spectrometry) method by utilizing a set of pure chemical standards to facilitate the identification and distribution of sterols, bile acids, and acylcarnitines in biological samples including human stool and plasma; mouse ileum, cecum, jejunum content, duodenum content, and liver; and pig bile, proximal colon, cecum, heart, stool, and liver. With this method, we detected 24 sterol, 32 bile acid, and 27 acylcarnitine standards in one analysis that were separated within 13 min by reversed-phase chromatography. Further, we observed different sterol, bile acid, and acylcarnitine profiles for the different biological samples across the different species. The simultaneous detection and annotation of sterols, bile acids, and acylcarnitines from reference standards and biological samples with high precision represents a valuable tool for screening these metabolites in routine scientific research.

Daily Rice Bran Consumption for 6 Months Influences Serum Glucagon-Like Peptide 2 and Metabolite Profiles without Differences in Trace Elements and Heavy Metals in Weaning Nicaraguan Infants at 12 Months of Age.

BACKGROUND: Environmental enteric dysfunction (EED) is associated with chronic gut inflammation affecting nutrient absorption and development of children, primarily in low- and middle-income countries. Several studies have shown that rice bran (RB) supplementation provides nutrients and modulates gut inflammation, which may reduce risk for undernutrition. OBJECTIVE: The aim was to evaluate the effect of daily RB dietary supplementation for 6 mo on serum biomarkers in weaning infants and associated changes in serum and stool metabolites. METHODS: A 6-mo randomized-controlled dietary intervention was conducted in a cohort of weaning 6-mo-old infants in León, Nicaragua. Anthropometric indices were obtained at 6, 8, and 12 mo. Serum and stool ionomics and metabolomics were completed at the end of the 6-mo intervention using inductively coupled plasma MS and ultra-high performance LC-tandem MS. The ɑ1-acid glycoprotein, C-reactive protein, and glucagon-like peptide 2 (GLP-2) serum EED biomarkers were measured by ELISA. RESULTS: Twenty-four infants in the control group and 23 in the RB group successfully completed the 6-mo dietary intervention with 90% dietary compliance. RB participants had higher concentrations of GLP-2 as compared with control participants at 12 mo [median (IQR): 743.53 (380.54) pg/mL vs. 592.50 (223.59) pg/mL; P = 0.04]. Metabolite profiles showed significant fold differences of 39 serum metabolites and 44 stool metabolites from infants consuming RB compared with control, and with significant metabolic pathway enrichment scores of 4.7 for the tryptophan metabolic pathway, 5.7 for polyamine metabolism, and 5.7 for the fatty acid/acylcholine metabolic pathway in the RB group. No differences were detected in serum and stool trace elements or heavy metals following daily RB intake for 6 mo. CONCLUSIONS: RB consumption influences a suite of metabolites associated with growth promotion and development, while also supporting nutrient absorption as measured by changes in serum GLP-2 in Nicaraguan infants. This clinical trial was registered at https://clinicaltrials.gov as NCT02615886.

Plasma and Urine Metabolite Profiles Impacted by Increased Dietary Navy Bean Intake in Colorectal Cancer Survivors: A Randomized-Controlled Trial.

Navy beans contain bioactive phytochemicals with colon cancer prevention properties as demonstrated in carcinogen-induced animal models. Human studies support that dietary navy bean intake modulates metabolism by the gut microbiome. This study investigated the effect of navy bean ingestion on plasma and urine metabolite profiles of overweight and obese colorectal cancer survivors. Twenty participants completed a single-blinded, randomized-controlled dietary intervention with precooked navy beans (35 g bean powder/day) or control (0 g/day) for 4 weeks. Plasma and urine were collected at baseline, 2 weeks, and 4 weeks following consumption. Nontargeted metabolomics was applied to study meals and snacks, navy beans, plasma, and urine. Increased navy bean consumption was hypothesized to (i) delineate dietary biomarkers and (ii) promote metabolic shifts relevant for cancer protection in the plasma and urine metabolome. At 4 weeks, 16 plasma and 16 urine metabolites were significantly different in the navy bean intervention group compared with placebo control (P < 0.05). Increased plasma 2,3-dihydroxy-2-methylbutyrate (1.34-fold), S-methylcysteine (1.92-fold), and pipecolate (3.89-fold), and urine S-adenosylhomocysteine (2.09-fold) and cysteine (1.60-fold) represent metabolites with cancer-protective actions following navy bean consumption. Diet-derived metabolites were detected in plasma or urine and confirmed for presence in the navy bean intervention meals and snacks. These included 3-(4-hydroxyphenyl)propionate, betaine, pipecolate, S-methylcysteine, choline, eicosapentaenoate (20:5n3), benzoate, S-adenosylhomocysteine, N-delta-acetylornithine, cysteine, 3-(4-hydroxyphenyl)lactate, gentisate, hippurate, 4-hydroxyhippurate, and salicylate. The navy bean dietary intervention for 4 weeks showed changes to pathways of metabolic importance to colorectal cancer prevention and merit continued attention for dietary modulation in future high-risk cohort investigations. PREVENTION RELEVANCE: This clinical study suggests that increased consumption of navy beans would deliver bioactive metabolites to individuals at high risk for colorectal cancer recurrence and produce metabolic shifts in plasma and urine profiles.

"notame": Workflow for Non-Targeted LC-MS Metabolic Profiling.

Metabolomics analysis generates vast arrays of data, necessitating comprehensive workflows involving expertise in analytics, biochemistry and bioinformatics in order to provide coherent and high-quality data that enable discovery of robust and biologically significant metabolic findings. In this protocol article, we introduce notame, an analytical workflow for non-targeted metabolic profiling approaches, utilizing liquid chromatography-mass spectrometry analysis. We provide an overview of lab protocols and statistical methods that we commonly practice for the analysis of nutritional metabolomics data. The paper is divided into three main sections: the first and second sections introducing the background and the study designs available for metabolomics research and the third section describing in detail the steps of the main methods and protocols used to produce, preprocess and statistically analyze metabolomics data and, finally, to identify and interpret the compounds that have emerged as interesting.

Rice bran supplementation modulates growth, microbiota and metabolome in weaning infants: a clinical trial in Nicaragua and Mali.

Rice bran supplementation provides nutrients, prebiotics and phytochemicals that enhance gut immunity, reduce enteric pathogens and diarrhea, and warrants attention for improvement of environmental enteric dysfunction (EED) in children. EED is a subclinical condition associated with stunting due to impaired nutrient absorption. This study investigated the effects of rice bran supplementation on weight for age and length for age z-scores (WAZ, LAZ), EED stool biomarkers, as well as microbiota and metabolome signatures in weaning infants from 6 to 12 months old that reside in Nicaragua and Mali. Healthy infants were randomized to a control (no intervention) or a rice bran group that received daily supplementation with increasing doses at each month (1-5 g/day). Stool microbiota were characterized using 16S rDNA amplicon sequencing. Stool metabolomes were analyzed using ultra-high-performance liquid-chromatography tandem mass-spectrometry. Statistical comparisons were completed at 6, 8, and 12 months of age. Daily consumption of rice bran was safe and feasible to support changes in LAZ from 6-8 and 8-12 months of age in Nicaragua and Mali infants when compared to control. WAZ was significantly improved only for Mali infants at 8 and 12 months. Mali and Nicaraguan infants showed major differences in the overall gut microbiota and metabolome composition and structure at baseline, and thus each country cohort demonstrated distinct microbial and metabolite profile responses to rice bran supplementation when compared to control. Rice bran is a practical dietary intervention strategy that merits development in rice-growing regions that have a high prevalence of growth stunting due to malnutrition and diarrheal diseases. Rice is grown as a staple food, and the bran is used as animal feed or wasted in many low- and middle-income countries where EED and stunting is prevalent.

Modulation of plasma and urine metabolome in colorectal cancer survivors consuming rice bran.

Rice bran has bioactive phytochemicals with cancer protective actions that involve metabolism by the host and the gut microbiome. Globally, colorectal cancer (CRC) is the third leading cause of cancer-related death and the increased incidence is largely attributed to poor dietary patterns, including low daily fiber intake. A dietary intervention trial was performed to investigate the impact of rice bran consumption on the plasma and urine metabolome of CRC survivors. Nineteen CRC survivors participated in a randomized-controlled trial that included consumption of heat-stabilized rice bran (30 g/day) or a control diet without rice bran for 4 weeks. A fasting plasma and first void of the morning urine sample were analyzed by non-targeted metabolomics using ultrahigh-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). After 4 weeks of either rice bran or control diets, 12 plasma and 16 urine metabolites were significantly different between the groups (p≤0.05). Rice bran intake increased relative abundance of plasma mannose (1.373-fold) and beta-citrylglutamate (BCG) (1.593-fold), as well as increased urine N-formylphenylalanine (2.191-fold) and dehydroisoandrosterone sulfate (DHEA-S) (4.488-fold). Diet affected metabolites, such as benzoate, mannose, eicosapentaenoate (20:5n3) (EPA), and N-formylphenylalanine have been previously reported for cancer protection and were identified from the rice bran food metabolome. Nutritional metabolome changes following increased consumption of whole grains such as rice bran warrants continued investigation for colon cancer control and prevention attributes as dietary biomarkers for positive effects are needed to reduce high risk for colorectal cancer recurrence.

Contribution of gut microbiota to metabolism of dietary glycine betaine in mice and in vitro colonic fermentation.

BACKGROUND: Accumulating evidence is supporting the protective effect of whole grains against several chronic diseases. Simultaneously, our knowledge is increasing on the impact of gut microbiota on our health and on how diet can modify the composition of our bacterial cohabitants. Herein, we studied C57BL/6 J mice fed with diets enriched with rye bran and wheat aleurone, conventional and germ-free C57BL/6NTac mice on a basal diet, 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 on the study samples 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. RESULTS: The bran-enriched study diets elevated the levels of betainized compounds in the colon contents of C57BL/6 J mice. The composition of microbiota changed, and the bran-enriched diets induced an increase in the relative abundance of several bacterial taxa, including Akkermansia, Bifidobacterium, Coriobacteriaceae, Lactobacillus, Parasutterella, and Ruminococcus, many of which are associated with improved health status or the metabolism of plant-based molecules. The levels of betainized compounds in the gut tissues of germ-free mice were significantly lower compared to conventional mice. In the in vitro model of the human gut, the production of betainized compounds was observed throughout the incubation, while the levels of glycine betaine decreased. In cereal samples, only low levels or trace amounts of other betaines than glycine betaine were observed. CONCLUSIONS: 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, adding another potential compound group acting as a mediator of the synergistic metabolic effect of diet and colonic microbiota.

Method Development of Near-Infrared Spectroscopy Approaches for Nondestructive and Rapid Estimation of Total Protein in Brown Rice Flour.

Rice varietal development and improvement programs are constantly seeking means to shorten the breeding cycle in order to deliver new, consumer-acceptable rice varieties to farmers and to consumers. Advances in molecular biology technologies have enabled breeders to use high-throughput genotyping to screen breeding lines. However, current phenotyping technologies, particularly for rice cooking and eating properties, have yet to match the efficiency of genotyping methodologies. A high-throughput and cost-effective phenotyping suite is essential because without phenotype, the value of genotypic information cannot be maximized. In this book chapter, we explore the application of near-infrared spectroscopy (NIRS), a high-throughput and nondestructive approach in characterizing rice grains, primarily describing method development and validation, instrument calibration, upgrading, and maintenance. We then focus on estimating protein content (PC) in brown rice as a case study because (1) PC is an attribute that contributes to the cooking behavior and the eating properties of cooked rice; and (2) proteins contain chemical bonds that can easily be detected by NIRS.

Comparative Rice Bran Metabolomics across Diverse Cultivars and Functional Rice Gene⁻Bran Metabolite Relationships.

Rice (Oryza sativa L.) processing yields ~60 million metric tons of bran annually. Rice genes producing bran metabolites of nutritional and human health importance were assessed across 17 diverse cultivars from seven countries using non-targeted metabolomics, and resulted in 378⁻430 metabolites. Gambiaka cultivar had the highest number and Njavara had the lowest number of metabolites. The 71 rice bran compounds of significant variation by cultivar included 21 amino acids, seven carbohydrates, two metabolites from cofactors and vitamins, 33 lipids, six nucleotides, and two secondary metabolites. Tryptophan, α-ketoglutarate, γ-tocopherol/ß-tocopherol, and γ-tocotrienol are examples of bran metabolites with extensive cultivar variation and genetic information. Thirty-four rice bran components that varied between cultivars linked to 535 putative biosynthetic genes using to the OryzaCyc 4.0, Plant Metabolic Network database. Rice genes responsible for bran composition with animal and human health importance is available for rice breeding programs to utilize in crop improvement.

Rice Bran Metabolome Contains Amino Acids, Vitamins & Cofactors, and Phytochemicals with Medicinal and Nutritional Properties.

BACKGROUND: Rice bran is a functional food that has shown protection against major chronic diseases (e.g. obesity, diabetes, cardiovascular disease and cancer) in animals and humans, and these health effects have been associated with the presence of bioactive phytochemicals. Food metabolomics uses multiple chromatography and mass spectrometry platforms to detect and identify a diverse range of small molecules with high sensitivity and precision, and has not been completed for rice bran. RESULTS: This study utilized global, non-targeted metabolomics to identify small molecules in rice bran, and conducted a comprehensive search of peer-reviewed literature to determine bioactive compounds. Three U.S. rice varieties (Calrose, Dixiebelle, and Neptune), that have been used for human dietary intervention trials, were assessed herein for bioactive compounds that have disease control and prevention properties. The profiling of rice bran by ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) and gas chromatography-mass spectrometry (GC-MS) identified 453 distinct phytochemicals, 209 of which were classified as amino acids, cofactors & vitamins, and secondary metabolites, and were further assessed for bioactivity. A scientific literature search revealed 65 compounds with health properties, 16 of which had not been previously identified in rice bran. This suite of amino acids, cofactors & vitamins, and secondary metabolites comprised 46% of the identified rice bran metabolome, which substantially enhanced our knowledge of health-promoting rice bran compounds provided during dietary supplementation. CONCLUSION: Rice bran metabolite profiling revealed a suite of biochemical molecules that can be further investigated and exploited for multiple nutritional therapies and medical food applications. These bioactive compounds may also be biomarkers of dietary rice bran intake. The medicinal compounds associated with rice bran can function as a network across metabolic pathways and this metabolite network may occur via additive and synergistic effects between compounds in the food matrix.

An assessment of metal contamination risk in sediments of Hara Biosphere Reserve, southern Iran with a focus on application of pollution indicators.

The aim of this study was to assess the pollution status of metals in sediments of Hara Biosphere Reserve using pollution indicators. For this purpose, sediment samples from nine locations were collected and characterized for metal content (Pb, Cr, Zn, Cu, and Fe) using the total digestion technique. Comparison of metal concentrations with that of sediment quality guidelines (SQGs) demonstrated no association with negative biological effects for Cu and Zn, while the values of Pb and Cr mainly illustrated to have association with negative biological effects. The results of the geo-accumulation index (Igeo) indicated no contamination for Cr, Cu, Zn, and Fe, while the values of Pb demonstrated to have moderate contamination based on I geo values. The analysis of the enrichment factor (EF) showed no enrichment for Cu and Zn and minor enrichment for Pb and Cr. Similar results were also found for quantification of contamination (QoC) analysis, where the values of Cu and Zn demonstrated to have a geogenic source of contamination, while the values of Pb and Cr mainly illustrated to have an anthropogenic source of contamination. According to EF and QoC calculations, the values of Cu and Zn were derived mainly from natural processes and exposure of material from the earth's crust, while the values for Pb and Cr were enriched by anthropogenic activities. The results of the contamination factor (Cf(i)) demonstrated low contamination levels for Fe, Cr, Zn, and Cu and moderate contamination levels for Pb. The pollution load index (PLI), showing the overall contamination of metals, demonstrated moderate pollution status in the study area.

Metal concentrations in tissues of two fish species from Qeshm Island, Iran.

The purpose of this study was to determine the concentrations of metals, cadmium, chromium, lead and nickel in Liza vaigiensis and Johnius carutta, in order to: compare metal concentrations between two species with different gender, and to determine the significance between metal concentrations in the gill, liver and muscle. The highest mean concentrations of cadmium, chromium, lead and nickel in different tissues of these two fish species were found in the liver of L. vaigiensis at 0.68, 0.83, 0.37 and 1.42 µg g(-1), respectively; while the lowest mean concentrations of cadmium, chromium, lead and nickel were observed in the muscle of J. carutta at 0.16, 0.16, 0.03 and 0.29 µg g(-1), respectively. The results showed that the metal concentrations in both species were higher in the females than in the males (except chromium in gill and cadmium in muscle of J. carutta). Also, the results indicated that the metal concentrations were different among fish tissues (one-way ANOVA, p < 0.001), but there was no difference (except lead in gills of J. carutta) between sex (male vs. female).