Urinary Metabolite Profiles Research Articles

Prenatal exposure to per- and polyfluoroalkyl substances and sex-specific associations with offspring adiposity at 10 years of age: Metabolic perturbation plays a role

BackgroundPer- and polyfluoroalkyl substances (PFAS) have been suspected as potential environmental obesogens, with several mechanisms being proposed, including the role of metabolomics. However, current epidemiological studies have yielded inconclusive findings. ObjectivesWe aimed to estimate the associations of prenatal exposure to PFAS with offspring adiposity measures, and to explore the potential metabolic pathways underlying these associations. MethodsA total of 464 mother–child pairs from the Sheyang Mini Birth Cohort Study (SMBCS) were included in this study. Cord serum concentrations of 12 PFAS and urine metabolite profiles at age 10 were obtained from the SMBCS database. Adiposity-related anthropometric measurements and body composition estimates of children aged 10 were used to assess offspring obesity. Multiple linear regression models and quantile g-computation were conducted to estimate the associations of prenatal exposure to individual and multiple PFAS with obesity at 10 years old. Metabolomics analysis was performed to characterize the biological pathways associated with PFAS exposure or obesity, subsequently identifying the overlapping metabolic pathways underlying the PFAS-obesity relationship. ResultsPrenatal exposure to several PFAS was significantly associated with elevated obesity-related markers in 10-year-old children. After stratification by sex, the effects were more pronounced in girls. Quantile g-computation results indicated that exposure to higher levels of PFAS mixtures during pregnancy was associated with increased odds of obesity in girls, with PFNA emerging as the predominant driving compound. Untargeted metabolomics results showed that several amino acid metabolic pathways were characterized as the overlapping pathways underlying the above associations. ConclusionsTaken together, our findings suggested the potential obesogenic effects of prenatal exposure to PFAS and offered insight into the possible metabolic mechanisms underlying PFAS-related offspring obesity.

Unraveling the role of gut microbiota by fecal microbiota transplantation in rat model of kidney stone disease

Emerging research on the microbiome highlights the significant role of gut health in the development of kidney stones, indicating that an imbalance in gut bacteria or dysbiosis can influence the formation of stones by altering oxalate metabolism and urinary metabolite profiles. In particular, the overabundance of specific bacteria such as Enterococcus and Oxalobacter spp., which are known to affect oxalate absorption, is observed in patients with urolithiasis. This study investigates the effects of gut dysbiosis on urolithiasis through fecal microbiota transplantation (FMT) from patients to rats and its impact on urinary mineral excretion and stone formation. Fecal samples from eight patients with calcium oxalate stones and ten healthy volunteers were collected to assess the gut microbiome. These samples were then transplanted to antibiotic-pretreated Wistar rats for a duration of four weeks. After transplantation, we evaluated changes in the fecal gut microbiome profile, urinary mineral excretion rates, and expression levels of intestinal zonula occluden-1 (ZO-1), SLC26A6 and renal NF-κB. In humans, patients with urolithiasis exhibited increased urinary calcium and oxalate levels, along with decreased citrate excretion and increased urinary supersaturation index. The fecal microbiota showed a notable abundance of Bacteroidota. In rodents, urolithiasis-FMT rats showed urinary disturbances similar to patients, including elevated pH, oxalate level, and supersaturation index, despite negative renal pathology. In addition, a slight elevation in the expression of renal NF-κB, a significant intestinal SLC26A6, and a reduction in ZO-1 expression were observed. The gut microbiome of urolithiasis-FMT rats showed an increased abundance of Bacteroidota, particularly Muribaculaceae, compared to their healthy FMT counterparts. In conclusion, the consistent overabundance of Bacteroidota in both urolithiasis patients and urolithiasis-FMT rats is related to altered intestinal barrier function, hyperoxaluria, and renal inflammation. These findings suggest that gut dysbiosis, characterized by an overgrowth of Bacteroidota, plays a crucial role in the pathogenesis of calcium oxalate urolithiasis, underscoring the potential of targeting the gut microbiota as a therapeutic strategy.

Longitudinal study and predictive modelling of urinary pesticide metabolite concentrations in residents of Guangzhou, China

Continuous human biomonitoring and predictive modelling of urinary pesticide metabolites are critical for evaluating pesticide exposure trends and associated health risks. We conducted repeat cross-sectional surveys to determine the urinary concentrations of eight pesticide metabolites in the residents of Guangzhou, China, from 2018 to 2022. We longitudinally analyzed the changes in these metabolite concentrations over the years and assessed the potential non-carcinogenic risks by calculating the hazard quotient and hazard index. No significant differences were observed in the total urinary pesticide metabolite concentrations over the 5 years (9.16–12.99 μg/L). The urinary concentrations of 3,5,6-trichloro-2-pyridinol and 2,4-dichlorophenoxyacetic acid reached their lowest levels in 2020 (1.47 and 0.11 μg/L). Conversely, urinary para-nitrophenol concentrations exhibited an inverse trend, peaking in 2020 (6.16 μg/L). The composition profiles of urinary pesticide metabolites showed that para-nitrophenol consistently constituted the largest proportion each year. Males consistently showed higher median concentrations of total urinary pesticide metabolites and individual metabolites of 3,5,6-trichloro-2-pyridinol, trans-3-(2,2-dichlorovinyl)-2,2-dimethylcyclopropane-1-carboxylic acid, and para-nitrophenol than females. The concentrations of cis-3-(2,2-dichlorovinyl)-2,2-dimethylcyclopropane-1-carboxylic acid in adults' urine were significantly higher than those in minors' urine each year. The total pesticide metabolite concentrations in adults' urine were significantly higher than those in minors' urine in 2018 and 2020, whereas no significant differences were observed in other years. No significant differences in urinary pesticide metabolite concentrations were observed among different BMI groups. Results showed that 14.17% of the population had hazard index values above 1, indicating a higher risk of health hazards. Three predictive models were employed to predict urinary pesticide metabolite concentrations for 2023–2024, revealing an increasing trend in 3,5,6-trichloro-2-pyridinol concentrations while other metabolites are expected to decrease. The study showed the concentration of para-nitrophenol peaked in 2020 while 3,5,6-trichloro-2-pyridinol and 2,4-dichlorophenoxyacetic acid reached their lowest levels, suggests that the COVID-19 pandemic may have influenced pesticide exposure patterns.

A gut reaction? The role of the microbiome in aggression

Recent research has unveiled conflicting evidence regarding the link between aggression and the gut microbiome. Here, we compared behavior profiles of control, germ-free (GF), and antibiotic-treated mice, as well as re-colonized GF mice to understand the impact of the gut microbiome on aggression using the resident-intruder paradigm. Our findings revealed a link between gut microbiome depletion and higher aggression, accompanied by notable changes in urine metabolite profiles and brain gene expression. This study extends beyond classical murine models to humanized mice to reveal the clinical relevance of early-life antibiotic use on aggression. Fecal microbiome transplant from infants exposed to antibiotics in early life (and sampled one month later) into mice led to increased aggression compared to mice receiving transplants from unexposed infants. This study sheds light on the role of the gut microbiome in modulating aggression and highlights its potential avenues of action, offering insights for development of therapeutic strategies for aggression-related disorders.

Changes in the urinary metabolome accompanied alterations in body mass and composition in women with overweight – impact of high versus low protein breakfast

IntroductionUnderstanding why subjects with overweight and with obesity vary in their response to dietary interventions is of major interest for developing personalized strategies for body mass regulation.ObjectivesThe aim of this study was to investigate the relationship between changes in the urine metabolome and body mass during a breakfast meal intervention. Furthermore, we aimed to elucidate if the baseline urine metabolome could predict the response to the two types of breakfast meals (high versus low protein) during the intervention.MethodsA total of 75 young, women with overweight were randomly allocated to one of two intervention groups: (1) High-protein (HP) or (2) low-protein (LP) breakfast as part of their habitual diet during a 12-week intervention. Beside the breakfast meal, participants were instructed to eat their habitual diet and maintain their habitual physical activity level. Nuclear magnetic resonance-based metabolomics was conducted on urine samples collected at baseline (wk 0), mid-intervention (wk 6), and at endpoint (wk 12). At baseline and endpoint, body mass was measured and DXA was used to measure lean body mass and fat mass.ResultsThe baseline urine metabolite profile showed a slightly higher correlation (R2 = 0.56) to body mass in comparison with lean body mass (R2 = 0.51) and fat mass (R2 = 0.53). Baseline 24-h urinary excretion of trigonelline (p = 0.04), N, N-dimethylglycine (p = 0.02), and trimethylamine (p = 0.03) were significantly higher in individuals who responded with a reduction in body mass to the HP breakfast.ConclusionsDifferences in the urine metabolome were seen for women that obtained a body weight loss in the response to the HP breakfast intervention and women who did not obtain a body weight loss, indicating that the urine metabolome contains information about the metabolic phenotype that influences the responsiveness to dietary interventions.

Protective signature of xanthohumol on cognitive function of APP/PS1 mice: a urine metabolomics approach by age.

Alzheimer's disease (AD) has an increasing prevalence, complicated pathogenesis and no effective cure. Emerging evidences show that flavonoid compounds such as xanthohumol (Xn) could play an important role as a dietary supplement or traditional Chinese herbal medicine in the management of diseases such as AD. This study aims to analyze the target molecules of Xn in the prevention and treatment of AD, and its potential mechanism from the perspective of metabolites. APP/PS1 mice 2- and 6-months old were treated with Xn for 3months, respectively, the younger animals to test for AD-like brain disease prevention and the older animals to address therapeutic effects on the disease. Memantine (Mem) was selected as positive control. Behavioral tests were performed to assess the course of cognitive function. Urine samples were collected and analyzed by high-performance liquid chromatography (HPLC) with tandem mass spectrometry (MS/MS) coupled with online Compound Discoverer software. Morris Water Maze (MWM) tests showed that Xn, like Mem, had a therapeutic but not a preventive effect on cognitive impairment. The expression levels of urinary metabolites appeared to show an opposite trend at different stages of Xn treatment, downregulated in the prevention phase while upregulated in the therapy phase. In addition, the metabolic mechanisms of Xn during preventive treatment were also different from that during therapeutic treatment. The signaling pathways metabolites nordiazepam and genistein were specifically regulated by Xn but not by Mem in the disease prevention stage. The signaling pathway metabolite ascorbic acid was specifically regulated by Xn in the therapeutic stage. In conclusion, dietary treatment with Xn altered the urinary metabolite profile at different stages of administration in APP/PS1 mice. The identified potential endogenous metabolic biomarkers and signal pathways open new avenues to investigate the pathogenesis and treatment of AD.

Urine metabolomics unravel the effects of short-term dietary interventions on oxidative stress and inflammation: a randomized controlled crossover trial

Dietary biomarkers in urine remain elusive when evaluating diet-induced oxidative stress and inflammation. In our previous study, we conducted a randomized controlled crossover trial to compare the short-term (4-weeks) effects of the balanced Korean diet (BKD) with Western diets, including the 2010 dietary guidelines for Americans (2010 DGA) and typical American diet (TAD), on various metabolic indices in obese Korean adults. Building on this work, the current research focuses on the impact of these dietary interventions on oxidative stress (d-ROMs and BAP) and inflammation (CRP, TNF-α, IL-6, IL-1β, MCP-1) biomarkers in serum, and the concurrent urine metabolomes. Each dietary regimen was in silico and experimentally examined for their antioxidant levels using ABTS, DPPH, and FRAP assays, as well as total flavonoid (TFC) and total phenolic (TPC) contents. We assessed post-intervention variations in oxidative stress and inflammation biomarkers in serum, as well as the urine metabolite profiles for the participants (n = 48, average age: 41 years). Antioxidant contents and associated total antioxidant capacity (TAC) were significantly higher for the recommended diets (BKD and 2010 DGA) compared to TAD (p < 0.05). Butanol extracts from recommended diets (BKD and 2010 DGA) showed significantly higher antioxidant activity compared to TAD in ABTS (p < 0.01), DPPH, and FRAP (p < 0.05) assays. Consistent results were observed in total phenolic and flavonoid contents, mirroring their respective antioxidant activities. Following the intervention period, oxidative stress & inflammation markers in serum varied marginally, however, the urine metabolite profiles were clearly demarcated for the BKD and Western dietary groups (PC1 = 5.41%). For BKD group, the pre- and post-intervention urine metabolite profiles were clearly segregated (PLS2 = 2.93%). Compared to TAD, urine extracts from the recommended dietary group showed higher abundance of benzoic acid & phenolic derivatives (VIP > 0.7, p < 0.05). Metabolites associated with oxidative stress were observed higher in the urine samples from Western dietary groups compared to BKD. Urine metabolomics data delineated the post-intervention effects of three dietary interventions which corroborates the respective findings for their effects on metabolic indices.

Insulin-Resistant Male LEW.1WR1 Rats Do Not Develop β-Cell Mass Expansion in Response to a Moderate Sucrose Diet

Characterizing changes in beta cell function during prolonged hyperinsulinemia and dietary stress is important to study to prevent diseases like metabolic dysfunction-associated steatotic liver disease and insulin resistance. This research investigates how a moderate sucrose (MS) diet affects insulin resistance and β-cell mass in two rat strains: LEW.1WR1 and Wistar Furth (WF). LEW.1WR1 rats seem to be sensitive to beta cell disruptions as weanlings. Twenty-one male LEW.1WR1 rats and sixteen male WF rats were studied over 18 weeks. The rats were divided into groups and given either the control or MS diet. Their body weight was monitored twice a week. Insulin tolerance tests (ITTs) and fasting blood glucose measurements were taken at intervals. Urine samples were analyzed to assess metabolic shifts, and pancreas tissue was examined to evaluate changes in β-cell mass. The LEW.1WR1 rats became overweight and showed higher insulin resistance than the WF rats. Both strains of rats on the MS diet displayed changes in urine metabolite profiles in terms of levels of lactic acid and alanine. This study highlights the impact or lack thereof of a moderate sucrose diet on body mass, insulin resistance, and β-cell mass, with notable effects observed specifically in LEW.1WR1 rats. These findings contribute to our understanding of how dietary sugar intake can affect metabolism when observed in models sensitive to metabolic defects.

Proton Nuclear Magnetic Resonance (1H NMR) Metabolomics Study in Serum, Urine, and Cystic Fluid for Differentiating Fertility and Staging of Intra-abdominal Hydatid Cyst in Adults.

Cystic echinococcosis (CE) is a parasitic zoonosis caused by the tapeworm Echinococcus granulosus. Over the past few years, a lot of research has been done on liver illnesses using metabolomics techniques to identify biomarkers which could identify the diseases in its early stages. The present study was done to explore biomarkers in serum, urine, and cystic fluid which would help in differentiating, staging, and assessing fertility of intra-abdominal hydatid cyst by using proton nuclear magnetic resonance (1H NMR) metabolomics. In the study, 28 subjects (16 cases and 12 controls) were enrolled. Staging of hydatid cysts was performed using ultrasonography. In patients complying with case and control definition, blood, urine, and cystic fluid were collected for complete blood count, urine culture, Echinococcus IgG enzyme-linked immunosorbent assay (ELISA), and metabolomic analysis. The 17, 15, and 11 metabolites in serum, urine, and cystic fluid samples were quantified, respectively, to differentiate between case and control group. In this study, we observed that there was a significant downregulation of succinate metabolite in urine samples of cases, down-regulation of five metabolites (isoleucine, valine, histidine, tyrosine and formate) and upregulation of alanine in cystic fluid of cases. Current study demonstrates that metabolomics can be used non-invasively for rapid diagnosis of CE. This is one of the very few studies, which used 1H NMR spectroscopy, to analyze the profile of metabolites in serum, urine, and cystic fluid in cases of CE and controls. Raj N, Pandey A, Roy R, et al. Proton Nuclear Magnetic Resonance (1H NMR) Metabolomics Study in Serum, Urine, and Cystic Fluid for Differentiating Fertility and Staging of Intra-abdominal Hydatid Cyst in Adults. Euroasian J Hepato-Gastroenterol 2024;14(1):30-34.

Urinary drug metabolite profiling of tuberculosis treatment failure using proton nuclear magnetic resonance

The underlying cause of tuberculosis (TB) treatment failure is still largely unknown. A 1H NMR approach was applied to identify and quantify a subset of TB drugs and drug metabolites: ethambutol (EMB), acetyl isoniazid (AcINH), isonicotinic acid, pyrazinamide (PZA), pyrazinoic acid and 5-hydroxy-pyrazinoic acid, from the urine of TB patients. Samples were collected before, during (weeks one, two and four) and after standardised TB treatment. The median concentrations of the EMB and PZA metabolites were comparable between the samples from patients with eventually cured and failed treatment outcomes. The INH metabolites showed comparatively elevated concentrations in the treatment failure patients during and after treatment. Variation in INH metabolite concentrations couldn’t be associated with the varying acetylator genotypes, and it is therefore suggested that treatment failure is influenced more so by other conditions, such as environmental factors, or individual variation in other INH metabolic pathways.

Identifying Predictive Biomarkers of Subclinical Mastitis in Dairy Cows through Urinary Metabotyping.

Mastitis is a significant infectious disease in dairy cows, resulting in milk yield loss and culling. Early detection of mastitis-prone cows is crucial for implementing effective preventive measures before disease onset. Current diagnosis of subclinical mastitis (SCM) relies on somatic cell count assessment post-calving, lacking predictive capabilities. This study aimed to identify metabolic changes in pre-SCM cows through targeted metabolomic analysis of urine samples collected 8 wks and 4 wks before calving, using mass spectrometry. A nested case-control design was employed, involving a total of 145 multiparous dairy cows, with disease occurrence monitored pre- and postpartum. Among them, 15 disease-free cows served as healthy controls (CON), while 10 cows exclusively had SCM, excluding those with additional diseases. Urinary metabolite profiling revealed multiple alterations in acylcarnitines, amino acids, and organic acids in pre-SCM cows. Metabotyping identified 27 metabolites that distinguished pre-SCM cows from healthy CON cows at both 8 and 4 wks before parturition. However, only four metabolites per week showed significant alterations (p < 0.005). Notably, a panel of four serum metabolites (asymmetric dimethylarginine, proline, leucine, and homovanillate) at 8 wks prepartum, and another panel (asymmetric dimethylarginine, methylmalonate, citrate, and spermidine) at 4 wks prepartum, demonstrated predictive ability as urinary biomarkers for SCM risk (AUC = 0.88; p = 0.02 and AUC = 0.88; p = 0.03, respectively). In conclusion, our findings indicate that metabolite testing can identify cows at risk of SCM as early as 8 and 4 wks before parturition. Validation of the two identified metabolite panels is warranted to implement these predictive biomarkers, facilitate early intervention strategies, and improve dairy cow management to mitigate the impact of SCM. Further research is needed to confirm the efficacy and applicability of these biomarkers in practical farm settings.

Urine Metabolite Profiles after the Consumption of a Low- and a High-Digestible Protein Meal, and Comparison of Urine Normalization Techniques.

In the context of dietary transition toward plant proteins, it is necessary to ensure protein security in populations. It would thus be of interest to identify biomarkers of altered protein digestibility in populations. We examined the association between urinary metabolites and the acute intake of low- or highly digestible protein in healthy volunteers. The urine samples were collected before and 9 h after the ingestion of a meal containing either no protein, zein (low-digestible) or whey protein isolate (highly digestible). The liquid chromatography-high resolution mass spectrometry metabolomics approach was used for the profiling of the urinary metabolites. For the standardization of metabolomics data sets, osmolality-based, standard normal variates (SNV) and probabilistic quotient normalization (PQN) techniques were used. The ANOVA-based factorial method, AComDim_ICA, was used for chemometrics analysis. The osmolality adjustment has a beneficial effect and the subsequent mathematical normalization improves the chemometric analysis further. Some changes in the urinary metabolomes were observed 9 h after the meal in the three groups. However, there was no difference in the urine metabolome between groups. No biomarker of protein digestibility can be identified after the ingestion of a single meal, even when marked differences in the digestion efficiency of protein have been observed.

P324 Urine metabolite profile characterizes clinical activity and disease extent in patients with inflammatory bowel disease

Abstract Background Inflammatory Bowel Disease (IBD), which comprises Crohn's Disease (CD) and Ulcerative Colitis (UC), is a disorder in which complex interactions between genetic, environmental, and microbial factors trigger alterations in the immune intestinal response. On the other hand, nutrient catabolism from the diet carried out by the microbiota results in the formation of metabolites that also impact immune responses on the intestinal mucosa. Considering that IBD patients have altered microbiota compared to healthy controls (HC) and that the urinary metabolic profile of an individual is the result of a combination of genetic, dietary, and microbial components, we propose the hypothesis that the urinary metabolic profile distinguishes IBD patients from HC. The aim of the study was to compare metabolomic analysis of urine samples from HC and IBD patients. Methods 27 patients were included [HC (n=10), UC (n=10), and CD (n=7)]. Clinical IBD activity was evaluated with partial Mayo score (UC) and Harvey-Bradshaw index (CD). Metabolites in urine were measured using gas chromatography-mass spectrometry (GC-MS). Metabolomic analysis and machine learning were performed using the MetaboAnalyst software. PLS-DA was used for multivariate analysis, and clustering separation was performed using hierarchical clustering. To identify and interpret patterns of metabolite concentration changes in a biologically meaningful way we perform Over Representation Analysis. Results In total, 62 organic acids were detected. Comparing UC and HC revealed 11 significantly altered metabolites (9 increased and 2 decreased). Main increased metabolites comprised lactic acid (p=0.0015), pyroglutamic acid (p=0.0147), isocitric acid (p=0.0325), and phosphoric acid (p=0.0091). Interestingly, the value of adipic and suberic acids were higher in remission vs. active patients. Furthermore, adipic acid was higher in UC patients with proctitis compared to left-side colitis/extensive colitis. Comparing CD and HC revealed only 2 significantly altered metabolites, glycolic acid (p=0.0214), and lactic acid (p=0.0111). Oxalic acid was more concentrated in CD patient with perianal disease. Finally, propionic and phosphoric acid were able to discriminate UC from CD patients. The most affected metabolic pathways were related to energy metabolism and oxidative stress, including the Warburg effect (p=0.0016), gluconeogenesis (p=0.0106), pyruvate metabolism (p=0.0197) and glutathione metabolism (p=0.0039) (Fig.1). Conclusion We found that the urinary metabolic profile of IBD patients differs from the HC, particularly for UC patients. These metabolites profile could be used to improve diagnosis and monitoring in patients with IBD.

Flavonoid Metabolites in Serum and Urine after the Ingestion of Selected Tropical Fruits.

The serum concentration and urinary excretion of flavonoids after the ingestion of guava, pineapple, and pomelo were determined using liquid chromatography-mass spectroscopy (LC-MS/MS). Each group of healthy volunteers was given 200 g of fresh fruit after overnight fasting and a 24-h flavonoid-free diet. The results demonstrate that only the glucuronic-conjugated metabolites of luteolin, quercetin, kaempferol, and myricetin were detected after fruit ingestion. The metabolites were first detected after 2 h, with the time to maximum concentration (Tmax) at 6 h. The most abundant metabolites for guava, pineapple, and pomelo were the glucuronide metabolites of quercetin (AUC0-8 5.4 ± 1.3 μg·h/mL), kaempferol (AUC0-8 9.9 ± 2.3 μg·h/mL), and luteolin (AUC0-8 6.4 ± 1.1 μg·h/mL), respectively. The flavonoids found in the 24-h urinary excretions were glucuronic- and mainly sulfate-conjugated metabolites. Quercetin metabolites were the most abundant after guava and pineapple ingestion, accounting for 900 and 700 μg, respectively. Luteolin metabolites were the most abundant after pomelo ingestion, accounting for 450 μg. The serum and urinary metabolite profiles suggested that guava and pineapple are good sources of quercetin, pineapple is a good source of kaempferol, and pomelo is a good source of luteolin. The study of flavonoid profiles may provide information for the selection of fruits as functional foods for their health benefits to help with various health conditions.

Interaction between a Sulfated Polysaccharide from Sea Cucumber and Gut Microbiota Influences the Fat Metabolism in Rats.

Due to its significant physiological effects, a sulfated polysaccharide has been considered an important nutrient of sea cucumber, but its metabolism in vivo is still unclear. The present study investigated the metabolism of a sea cucumber sulfated polysaccharide (SCSP) in rats and its influence on the metabolite profiles. The quantification by HPLC-MS/MS revealed that the blood level of SCSP achieved a maximum of 54.0 ± 4.8 μg/mL at 2 h after gavage, almost no SCSP was excreted through urine, and 55.4 ± 29.8% of SCSP was eliminated through feces within 24 h. These results prove the utilization of SCSP by gut microbiota, and a further microbiota sequencing analysis indicated that the SCSP utilization in the gut was positively correlated with Muribaculaceae and Clostridia_UCG-014. In addition, the non-targeted metabolomic analysis demonstrated the significant effects of SCSP administration on the metabolite profiles of blood, urine, and feces. It is worth noting that the SCSP supplement decreased palmitic acid, stearic acid, and oleic acid in blood and urine while increasing stearic acid, linoleic acid, and γ-linolenic acid in feces, suggesting the inhibition of fat absorption and the enhancement of fat excretion by SCSP, respectively. The present study shed light on the metabolism in vivo and the influence on the fat metabolism of SCSP.

Development of a single mobile phase for LC-IM-MS-based discovery lipidomics and metabolic phenotyping: Application to methapyrilene hepatotoxicity in the rat

The untargeted global profiling of endogenous metabolites and lipids has the potential to increase knowledge and understanding in many areas of biology. LC-MS/MS is a key technology for such analyses however, several different LC methodologies, using different mobile phase compositions, are required to cover the diversity in polarity and analyte structure encountered in biological samples. Most notably many lipid screening methods make use of isopropanol (IPA) as a major component of mobile phases employed for comprehensive lipidomic profiling. In order to increase laboratory efficiency, and minimize opportunities for errors, a suite of methods, based on a single acetonitrile (ACN)-aqueous buffer mobile phase combination, has been developed. This mobile phase can be used for hydrophobic interaction liquid chromatography on an amide stationary phase (for polar analytes), reversed-phase (RP) LC analysis on a C8 stationary phase (for moderately polar-non-polar compounds) and RPLC using a CSH phenyl-hexyl bonded column (for lipids). All of these sub 10 minute separations had good throughput and reproducibility with CV's of analyte response <25 % whilst eliminating the need for complex mobile phase preparation and the use of IPA as an organic modifier for lipidomics. Advantages of removing IPA and replacing it with the ACN-based method were a 58 % increase in peak capacity for lipids, with improved resolution for the di- and triglycerides and cholesterol esters compared to current methods. Compared to the IPA-containing solvent system the ACN-based mobile phase also resulted in a 61 % increase in lipid feature detection. The utility of this “universal” mobile phase approach was demonstrated by its application to a rat toxicology study investigating the consequences of methapyrilene administration through on the endogenous metabolite profiles of plasma and urine. Methapyrilene and its metabolites were also profiled in these samples.

Altered dietary behaviour during pregnancy impacts systemic metabolic phenotypes.

Evidence suggests consumption of a Mediterranean diet (MD) can positively impact both maternal and offspring health, potentially mediated by a beneficial effect on inflammatory pathways. We aimed to apply metabolic profiling of serum and urine samples to assess differences between women who were stratified into high and low alignment to a MD throughout pregnancy and investigate the relationship of the diet to inflammatory markers. From the ORIGINS cohort, 51 pregnant women were stratified for persistent high and low alignment to a MD, based on validated MD questionnaires. 1H Nuclear Magnetic Resonance (NMR) spectroscopy was used to investigate the urine and serum metabolite profiles of these women at 36 weeks of pregnancy. The relationship between diet, metabolite profile and inflammatory status was investigated. There were clear differences in both the food choice and metabolic profiles of women who self-reported concordance to a high (HMDA) and low (LMDA) Mediterranean diet, indicating that alignment with the MD was associated with a specific metabolic phenotype during pregnancy. Reduced meat intake and higher vegetable intake in the HMDA group was supported by increased levels of urinary hippurate (p = 0.044) and lower creatine (p = 0.047) levels. Serum concentrations of the NMR spectroscopic inflammatory biomarkers GlycA (p = 0.020) and GlycB (p = 0.016) were significantly lower in the HDMA group and were negatively associated with serum acetate, histidine and isoleucine (p < 0.05) suggesting a greater level of plant-based nutrients in the diet. Serum branched chain and aromatic amino acids were positively associated with the HMDA group while both urinary and serum creatine, urine creatinine and dimethylamine were positively associated with the LMDA group. Metabolic phenotypes of pregnant women who had a high alignment with the MD were significantly different from pregnant women who had a poor alignment with the MD. The metabolite profiles aligned with reported food intake. Differences were most significant biomarkers of systemic inflammation and selected gut-microbial metabolites. This research expands our understanding of the mechanisms driving health outcomes during the perinatal period and provides additional biomarkers for investigation in pregnant women to assess potential health risks.

Identification and validation of diagnostic biomarkers for intrahepatic cholestasis of pregnancy based on untargeted and targeted metabolomics analyses of urine metabolite profiles

BackgroundIntrahepatic cholestasis of pregnancy (ICP) is a prevalent pregnancy-specific complication that presents with maternal itching and elevated serum bile acid levels. ICP is associated with unfavorable pregnancy outcomes, severely decreasing the pregnant woman’s quality of life. Timely identification of ICP is crucial for effective management and improved outcomes.MethodsWe collected urine samples from 8 patients with ICP and 8 healthy individuals. We used Liquid Chromatography-Mass Spectrometry (LC-MS) to detect metabolite expression levels, then conducted a series of bioinformatic analyses to explore the potential biological meanings of differentially expressed metabolites, and preliminarily discovered several candidate biomarkers. To validate these candidate biomarkers, we performed Gas Chromatography-Mass Spectrometry (GC-MS) detection and analyzed their diagnostic values using receiver operating characteristic (ROC) curve.ResultsUntargeted metabolomics data showed that 6129 positive peaks and 6218 negative peaks were extracted from each specimen. OPLS-DA analysis and the heat map for cluster analysis showed satisfactory capability in discriminating ICP specimens from controls. Subsequent analysis extracted 64 significantly differentially expressed metabolites, which could be potential biomarkers for diagnosis of ICP. Based on the KEGG enrichment analyses, six candidate biomarkers were preliminarily identified. Two most promising biomarkers (3-hydroxypropionic acid and uracil) were validated by targeted metabolomics analyses with the area under the curve (AUC) of 0.920 and 0.850 respectively.ConclusionBased on preliminary screening from untargeted metabolomics and subsequent validation through targeted metabolomics, 3-hydroxypropionic acid and uracil were identified as promising diagnostic biomarkers for ICP.

Sulforaphane Bioavailability in Healthy Subjects Fed a Single Serving of Fresh Broccoli Microgreens.

Cruciferous vegetable consumption is associated with numerous health benefits attributed to the phytochemical sulforaphane (SFN) that exerts antioxidant and chemopreventive properties, among other bioactive compounds. Broccoli sprouts, rich in SFN precursor glucoraphanin (GRN), have been investigated in numerous clinical trials. Broccoli microgreens are similarly rich in GRN but have remained largely unexplored. The goal of this study was to examine SFN bioavailability and the microbiome profile in subjects fed a single serving of fresh broccoli microgreens. Eleven subjects participated in a broccoli microgreens feeding study. Broccoli microgreens GRN and SFN contents and stability were measured. Urine and stool SFN metabolite profiles and microbiome composition were examined. Broccoli microgreens had similar GRN content to values previously reported for broccoli sprouts, which was stable over time. Urine SFN metabolite profiles in broccoli microgreens-fed subjects were similar to those reported previously in broccoli sprouts-fed subjects, including the detection of SFN-nitriles. We also reported the detection of SFN metabolites in stool samples for the first time. A single serving of broccoli microgreens did not significantly alter microbiome composition. We showed in this study that broccoli microgreens are a significant source of SFN. Our work provides the foundation for future studies to establish the health benefits of broccoli microgreens consumption.

Urinary Metabolite Profiling to Non-Invasively Monitor the Omega-3 Index: An Exploratory Secondary Analysis of a Randomized Clinical Trial in Young Adults.

The Omega-3 Index (O3I) reflects eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) content in erythrocytes. While the O3I is associated with numerous health outcomes, its widespread use is limited. We investigated whether urinary metabolites could be used to non-invasively monitor the O3I in an exploratory analysis of a previous placebo-controlled, parallel arm randomized clinical trial in males and females (n = 88) who consumed either ~3 g/d olive oil (OO; control), EPA, or DHA for 12 weeks. Fasted blood and first-void urine samples were collected at baseline and following supplementation, and they were analyzed via gas chromatography and multisegment injection-capillary electrophoresis-mass spectrometry (MSI-CE-MS), respectively. We tentatively identified S-carboxypropylcysteamine (CPCA) as a novel urinary biomarker reflecting O3I status, which increased following both EPA and DHA (p < 0.001), but not OO supplementation, and was positively correlated to the O3I (R = 0.30, p < 0.001). Additionally, an unknown dianion increased following DHA supplementation, but not EPA or OO. In ROC curve analyses, CPCA outperformed all other urinary metabolites in distinguishing both between OO and EPA or DHA supplementation groups (AUC > 80.0%), whereas the unknown dianion performed best in discriminating OO from DHA alone (AUC = 93.6%). Candidate urinary biomarkers of the O3I were identified that lay the foundation for a non-invasive assessment of omega-3 status.