Graphenoxide Cross-Linker Based Potentiometric Biosensor Design for Sarcosine Determination.

BACKGROUND: Sarcosine, also known as N-methyl glycine, is a natural amino acid that is an intermediate and by product in glycine synthesis and degradation. Recently found in many peptides, sarcosine has been researched as a newly accepted prostate cancer marker. The increased concentration of sarcosine in blood serum and the urine showed that malignancy of measured prostate cancer cells is active. OBJECTIVE: In this article, we aimed to design a potentiometric biosensor for detection of sarcosine with a low detection limit, high selectivity, short response time, wide linear range, and satisfactory long-term stability. METHODS: In this article, we developed a new Graphene oxide (GFOX) photosensitive cross-linker based potentiometric biosensor based on the AmiNoAcid (monomer) Decorated and Light Underpinning Conjugation Approach (ANADOLUCA) method. The functional groups determined using Raman, FT-IR, XPS analyzes, and surface characterization, the morphology of synthesized GFOX photosensitive cross-linker were determined by TEM and AFM studies. Then, the performance of the GFOX based potentiometric biosensor has been evaluated. RESULTS: When the usage of the developed GFOX doped potentiometric biosensor against sarcosine determination, it was found that 10-4 mM sarcosine was determined in 60 seconds in the solution. In addition, the detection limit of the GFOX doped potentiometric biosensor was found to be 9.45x10-7 mM, and the linear potentiometric biosensor was found to be in the concentration range of 10-1 to 10-5 mM. The selectivity studies of the developed potentiometric biosensor were investigated using glycine solutions, and it was determined that GFOX doped potentiometric biosensor was more selective against sarcosine. Besides this, a reusability test using 10-3 mM sarcosine solution showed that reproducible studies were performed without the loss of potential of designed potentiometric biosensor and no loss of sensitivity. CONCLUSION: After applying the framework, we get a new potentiometric biosensor for sarcosine determination. GFOX photosensitive cross-linker was used in designing potentiometric biosensors, and this increased the stability and efficiency of the biosensor. Therefore, the developed potentiometric biosensor for sarcosine determination could be easily used for the early diagnosis of prostate cancer.

Glycine/glycine N-methyltransferase/sarcosine axis mediates benzene-induced hematotoxicity.

Benzene, an important and widely used industrial chemical, is the cause of different types of blood disorders. However, the mechanisms of benzene-induced hematotoxicity are still unclear. This study aimed to explore the effects of benzene on metabolism, especially in amino acid metabolism, in human peripheral blood B lymphocyte cells (AHH-1 cells) treated with 1,4-benzoquinone (1,4-BQ) and in benzene-exposed population based on the un-targeted and targeted metabolomics platforms. The results showed that 1,4-BQ disturbed the metabolic activity, such as arginine biosynthesis, citrate cycle, glycine, serine, and threonine metabolism pathways, and significantly upregulated the ratio of sarcosine/glycine in vitro. Meanwhile, the targeted metabolomics further showed that the ratio of sarcosine/glycine was also increased in the benzene exposure population. Notably, the expression of glycine N-methyltransferase (GNMT), an enzyme catalyzing the transformation of glycine to sarcosine, was upregulated both in 1,4-BQ treated AHH-1 cells and benzene-exposed workers. These results imply that the glycine/GNMT/sarcosine axis was involved in benzene-induced hematotoxicity. Such evidence will help to develop a better understanding of the underlying mechanism of benzene-induced hematotoxicity at the level of amino acid metabolism.

Integrated metabolomic analysis and cytokine profiling define clusters of immuno-metabolic correlation in new-onset psoriasis.

The association between the metabolic profile and inflammatory cytokines in psoriasis is poorly understood. We analyzed the metabolic and cytokine/chemokine profiles in serum and skin from patients with new-onset psoriasis and healthy subjects (n = 7/group) by HR-MAS NMR and Bio-Plex immunoassay. Immuno-metabolic correlation matrix was analyzed in skin and serum to identify a potential immune-metabolic signature. Metabolomics analysis showed a significant increase in ascorbate and a decrease in scyllo-inositol, and a trend towards an increase in eight other metabolites in psoriatic skin. In serum, there was a significant increase of dimethylglycine and isoleucine. In parallel, psoriatic skin exhibited an increase of early inflammatory cytokines (IL-6, IL-8, TNF-α, IL-1ß) and correlation analysis highlighted some major clusters of immune-metabolic correlations. A cluster comprising scyllo-inositol and lysine showed correlations with T-cell cytokines; a cluster comprising serine and taurine showed a negative correlation with early inflammatory cytokines (IL-6, G-CSF, CCL3). A strong positive correlation was enlightened between glutathione and inflammatory cytokines/angiogenesis promoters of psoriasis. The integration of metabolic and immune data indicated a molecular signature constituted by IL-6, IL1-ra, DMG, CCL4, Ile, Gly and IL-8, which could discriminate patients and healthy subjects and could represent a candidate tool in the diagnosis of new-onset psoriasis.

Associations of circulating choline and its related metabolites with cardiometabolic biomarkers: an international pooled analysis.

BACKGROUND: Choline is an essential nutrient; however, the associations of choline and its related metabolites with cardiometabolic risk remain unclear. OBJECTIVE: We examined the associations of circulating choline, betaine, carnitine, and dimethylglycine (DMG) with cardiometabolic biomarkers and their potential dietary and nondietary determinants. METHODS: The cross-sectional analyses included 32,853 participants from 17 studies, who were free of cancer, cardiovascular diseases, chronic kidney diseases, and inflammatory bowel disease. In each study, metabolites and biomarkers were log-transformed and standardized by means and SDs, and linear regression coefficients (ß) and 95% CIs were estimated with adjustments for potential confounders. Study-specific results were combined by random-effects meta-analyses. A false discovery rate <0.05 was considered significant. RESULTS: We observed moderate positive associations of circulating choline, carnitine, and DMG with creatinine [ß (95% CI): 0.136 (0.084, 0.188), 0.106 (0.045, 0.168), and 0.128 (0.087, 0.169), respectively, for each SD increase in biomarkers on the log scale], carnitine with triglycerides (ß = 0.076; 95% CI: 0.042, 0.109), homocysteine (ß = 0.064; 95% CI: 0.033, 0.095), and LDL cholesterol (ß = 0.055; 95% CI: 0.013, 0.096), DMG with homocysteine (ß = 0.068; 95% CI: 0.023, 0.114), insulin (ß = 0.068; 95% CI: 0.043, 0.093), and IL-6 (ß = 0.060; 95% CI: 0.027, 0.094), but moderate inverse associations of betaine with triglycerides (ß = -0.146; 95% CI: -0.188, -0.104), insulin (ß = -0.106; 95% CI: -0.130, -0.082), homocysteine (ß = -0.097; 95% CI: -0.149, -0.045), and total cholesterol (ß = -0.074; 95% CI: -0.102, -0.047). In the whole pooled population, no dietary factor was associated with circulating choline; red meat intake was associated with circulating carnitine [ß = 0.092 (0.042, 0.142) for a 1 serving/d increase], whereas plant protein was associated with circulating betaine [ß = 0.249 (0.110, 0.388) for a 5% energy increase]. Demographics, lifestyle, and metabolic disease history showed differential associations with these metabolites. CONCLUSIONS: Circulating choline, carnitine, and DMG were associated with unfavorable cardiometabolic risk profiles, whereas circulating betaine was associated with a favorable cardiometabolic risk profile. Future prospective studies are needed to examine the associations of these metabolites with incident cardiovascular events.

Oxytocin-induced increase in N,N-dimethylglycine and time course of changes in oxytocin efficacy for autism social core symptoms.

BACKGROUND: Oxytocin is expected as a novel therapeutic agent for autism spectrum disorder (ASD) core symptoms. However, previous results on the efficacy of repeated administrations of oxytocin are controversial. Recently, we reported time-course changes in the efficacy of the neuropeptide underlying the controversial effects of repeated administration; however, the underlying mechanisms remained unknown. METHODS: The current study explored metabolites representing the molecular mechanisms of oxytocin's efficacy using high-throughput metabolomics analysis on plasma collected before and after 6-week repeated intranasal administration of oxytocin (48 IU/day) or placebo in adult males with ASD (N = 106) who participated in a multi-center, parallel-group, double-blind, placebo-controlled, randomized controlled trial. RESULTS: Among the 35 metabolites measured, a significant increase in N,N-dimethylglycine was detected in the subjects administered oxytocin compared with those given placebo at a medium effect size (false discovery rate (FDR) corrected P = 0.043, d = 0.74, N = 83). Furthermore, subgroup analyses of the participants displaying a prominent time-course change in oxytocin efficacy revealed a significant effect of oxytocin on N,N-dimethylglycine levels with a large effect size (PFDR = 0.004, d = 1.13, N = 60). The increase in N,N-dimethylglycine was significantly correlated with oxytocin-induced clinical changes, assessed as changes in quantifiable characteristics of autistic facial expression, including both of improvements between baseline and 2 weeks (PFDR = 0.006, r = - 0.485, N = 43) and deteriorations between 2 and 4 weeks (PFDR = 0.032, r = 0.415, N = 37). LIMITATIONS: The metabolites changes caused by oxytocin administration were quantified using peripheral blood and therefore may not directly reflect central nervous system changes. CONCLUSION: Our findings demonstrate an association of N,N-dimethylglycine upregulation with the time-course change in the efficacy of oxytocin on autistic social deficits. Furthermore, the current findings support the involvement of the N-methyl-D-aspartate receptor and neural plasticity to the time-course change in oxytocin's efficacy. TRIAL REGISTRATION: A multi-center, parallel-group, placebo-controlled, double-blind, confirmatory trial of intranasal oxytocin in participants with autism spectrum disorders (the date registered: 30 October 2014; UMIN Clinical Trials Registry: https://upload.umin.ac.jp/cgi-open-bin/ctr_e/ctr_view.cgi?recptno=R000017703 ) (UMIN000015264).

Choline Metabolism and Risk of Atrial Fibrillation and Heart Failure in the PREDIMED Study.

BACKGROUND: Few studies have examined the associations of trimethylamine-N-oxide (TMAO) and its precursors (choline, betaine, dimethylglycine, and L-carnitine) with the risk of atrial fibrillation (AF) and heart failure (HF). This study sought to investigate these associations. METHODS: Prospective associations of these metabolites with incident AF and HF were examined among participants at high cardiovascular risk in the PREDIMED study (PREvención con DIeta MEDiterránea) after follow-up for about 10 years. Two nested case-control studies were conducted, including 509 AF incident cases matched to 618 controls and 326 HF incident cases matched to 426 controls. Plasma levels of TMAO and its precursors were semi-quantitatively profiled with liquid chromatography tandem mass spectrometry. Odds ratios were estimated with multivariable conditional logistic regression models. RESULTS: After adjustment for classical risk factors and accounting for multiple testing, participants in the highest quartile vs. the lowest quartile of baseline choline and betaine levels had a higher risk of AF [OR (95% CI): 1.85 (1.30-2.63) and 1.57 (1.09-2.24), respectively]. The corresponding OR for AF for extreme quartiles of dimethylglycine was 1.39 (0.99-1.96). One SD increase in log-transformed dimethylglycine was positively associated with AF risk (OR, 1.17; 1.03-1.33). The corresponding ORs for HF for extreme quartiles of choline, betaine, and dimethylglycine were 2.51 (1.57-4.03), 1.65 (1.00-2.71) and 1.65 (1.04-2.61), respectively. TMAO and L-carnitine levels were not associated with AF or HF. CONCLUSIONS: Our findings support the role of the choline metabolic pathway in the pathogenesis of AF and HF.

Comprehensive metabolomic analysis of first-trimester serum identifies biomarkers of early-onset hypertensive disorder of pregnancy.

Hypertensive disorders of pregnancy (HDP) lead to the death of approximately 30,000 women annually, and the identification of biomarkers to predict their onset before symptom occurrence is crucial. Here, we aimed to identify the first-trimester maternal serum biomarkers for predicting early-onset HDP via a comprehensive metabolomic analysis. This study was conducted by the Fukushima Regional Center as an adjunct study to the Japan Environment and Children's Study. The study comprised 12 patients with early-onset HDP and 12 control subjects with healthy pregnancy whose medical background information was matched with that of the patients by propensity-score matching. Capillary electrophoresis and mass spectrometry-based quantitative analysis of charged metabolites were performed with the first-trimester maternal serum samples. Welch's t-test was used to analyse metabolite peak areas in the two groups. A total of 166 charged metabolites were identified. The peak area of N-dimethylglycine and S-methylcysteine was significantly higher in the first-trimester serum of patients with early-onset HDP than in the controls. Conversely, the peak area of munic acid was significantly decreased in the serum of patients with early-onset HDP. Although we identified potential biomarkers for the prediction and diagnosis of early-onset HDP, no clear marker was identified because of a low statistical power.

Plasma Sarcosine Measured by Gas Chromatography-Mass Spectrometry Distinguishes Prostatic Intraepithelial Neoplasia and Prostate Cancer from Benign Prostate Hyperplasia.

OBJECTIVE: Sarcosine was postulated in 2009 as a biomarker for prostate cancer (PCa). Here, we assess plasma sarcosine as a biomarker that is complementary to prostate-specific antigen (PSA). METHODS: Plasma sarcosine was measured using gas chromatography-mass spectrometry (GC-MS) in adults classified as noncancerous controls (with benign prostate hyperplasia [BPH], n = 36), with prostatic intraepithelial neoplasia (PIN, n = 16), or with PCa (n = 27). Diagnostic accuracy was assessed using receiver operating characteristic curve analysis. RESULTS: Plasma sarcosine levels were higher in the PCa (2.0 µM [1.3-3.3 µM], P <.01) and the PIN (1.9 µM [1.2-6.5 µM], P <.001) groups than in the BPH (0.9 µM [0.6-1.4 µM]) group. Plasma sarcosine had "good" and "very good" discriminative capability to detect PIN (area under the curve [AUC], 0.734) and PCa (AUC, 0.833) versus BPH, respectively. The use of PSA and sarcosine together improved the overall diagnostic accuracy to detect PIN and PCa versus BPH. CONCLUSION: Plasma sarcosine measured by GC-MS had "good" and "very good" classification performance for distinguishing PIN and PCa, respectively, relative to noncancerous patients diagnosed with BPH.

Natural Choline from Egg Yolk Phospholipids Is More Efficiently Absorbed Compared with Choline Bitartrate; Outcomes of A Randomized Trial in Healthy Adults.

Choline is a vitamin-like essential nutrient, important throughout one's lifespan. Therefore, choline salts are added to infant formula, supplements and functional foods. However, if choline is present in a natural form, e.g. bound to phospholipids, it may be more efficiently absorbed. The study's aim was to evaluate if choline uptake is improved after consumption of an egg yolk phospholipid drink, containing 3 g of phospholipid bound choline, compared to a control drink with 3 g of choline bitartrate. We performed a randomized, double blind, cross-over trial with 18 participants. Plasma choline, betaine and dimethylglycine concentrations were determined before and up to six hours after consumption of the drinks. The plasma choline response, as determined by the incremental area under the curve, was four times higher after consumption of the egg yolk phospholipid drink compared with the control drink (p < 0.01). Similar outcomes were also observed for choline's main metabolites, betaine (p < 0.01) and dimethylglycine (p = 0.01). Consumption of natural choline from egg yolk phospholipids improved choline absorption compared to consumption of chemically produced choline bitartrate. This information is of relevance for the food industry, instead of adding choline-salts, adding choline from egg yolk phospholipids can improve choline uptake and positively impact health.

Serum Betaine and Dimethylglycine Are Higher in South Asian Compared with European Pregnant Women in Canada, with Betaine and Total Homocysteine Inversely Associated in Early and Midpregnancy, Independent of Ethnicity.

BACKGROUND: As a methyl donor required in the folate-vitamin B-12 independent remethylation of total homocysteine (tHcy) to methionine, betaine is critical for fetal development. Pregnant South Asian women living in Canada had a higher reported prevalence of low vitamin B-12 status compared with Europeans; betaine concentrations in this population are unknown. OBJECTIVES: We aimed to compare serum betaine concentrations between South Asian and European pregnant women, and to determine the relation between betaine and tHcy concentrations in early pregnancy. METHODS: A retrospective cohort study was conducted using biobanked serum samples of 723 apparently healthy pregnant women of South Asian (50%) and European ethnicity residing in British Columbia, Canada. Betaine, dimethylglycine (DMG), tHcy, and related metabolites were quantified in samples collected in the first (8-13 weeks of gestation) and second (14-20 weeks of gestation) trimesters. The relation between betaine and tHcy concentrations was assessed using a generalized regression model adjusted for weeks of gestation, ethnicity, prepregnancy BMI, maternal age, neonatal sex, parity, total vitamin B-12, folate, pyridoxal 5'-phosphate, and methionine concentrations. RESULTS: Median serum concentrations of betaine and its metabolite DMG were higher in South Asian women in the first (19.8 [IQR: 16.3-25.0] and 1.55 [IQR: 1.30-1.96] $\mu {\rm mol/L} $, respectively) and second trimesters (16.1 [IQR: 12.9-19.8] and 1.42 [IQR: 1.14-1.81] $\mu {\rm mol/L} $, respectively) compared with European women (17.6 [IQR: 13.7-22.6] and 1.38 [IQR: 1.12-1.77] $\mu {\rm mol/L} $, respectively) and (12.9 [IQR: 10.6-16.7] and 1.19 [IQR: 0.97-1.52] $\mu {\rm mol/L} $, respectively; all P values < 0.0001). Betaine was inversely associated with tHcy concentration (ß = -0.0208; 95% CI: -0.0341, -0.00742; P = 0.002). Additionally, total vitamin B-12 was associated with tHcy concentration (ß = -0.0312; 95% CI: -0.0401, -0.0224), after adjusting for confounding factors. CONCLUSIONS: Pregnant South Asian women residing in Canada had higher betaine and DMG concentrations, compared with women of European ethnicity, while betaine and total vitamin B-12 predicted tHcy independent of ethnicity. Our results emphasize the role of betaine, as methyl donor, in the remethylation of tHcy in a folate-replete population.

Amperometric sarcosine biosensor based on hollow magnetic Pt-Fe3O4@C nanospheres.

Sarcosine is a recently identified biomarker for prostate cancer. However, the rapid detection methods for sarcosine are relatively lack because of the low concentration and the presence of complicated interfering substances in serum or urine. In this manuscript, hollow nanospheres of Fe3O4 was synthesized and used as carrier to disperse Pt (Pt) nanoparticles. In order to achieve excellent electron transfer ability, we use polyaniline to coat Pt-Fe3O4 nanoparticles, and pyrolyze the polyaniline to carbon (C). Thus, hollow magnetic Pt-Fe3O4@C nanocomposites with good electron transfer ability are formed. The Pt-Fe3O4@C nanocomposites have high catalytic activity and stability. The nanocomposites were immobilized on glassy carbon electrode (GCE) to construct a nonenzyme hydrogen peroxide (H2O2) sensor (Pt-Fe3O4@C/GCE). We further construct a sensitive sarcosine biosensor by immobilizing sarcosine oxidase (SOx) on the Pt-Fe3O4@C/GCE. The high catalytic activity and good biocompatibility of Pt-Fe3O4@C nanocomposites greatly retained the bioactivity of immobilized SOx, and the prepared sarcosine biosensor has good electrocatalytic performance towards sarcosine. It has a linear detection range between 0.5 and 60 µM with a limit of detection (LOD) of 0.43 µM (the signal to noise ratio is 3), and the sensitivity is 3.45 nA µM-1 (48.8 nA µM-1 cm-2), which has the potential to be used for rapid screening of prostate cancer.

Amperometric sarcosine biosensor with strong anti-interference capabilities based on mesoporous organic-inorganic hybrid materials.

Amperometric enzyme biosensors are some of the simplest and cheapest types of medical devices used in the rapid detection of biomarkers that have been developed in the past fifty years. When the concentrations of biomarkers are at micromoles per liter, such as for sarcosine, which was recently discovered as a biomarker for prostate cancer, the response signal of the interferences is huge, and the biosensor is hard to satisfy the requirements of practical applications. In this manuscript, we describe a strategy for synthesizing a surface electronegative organic-inorganic hybrid mesoporous material, which could reduce the interference signal much better than Nafion and Chitosan. We verify that the surface potential of the carrier nanomaterial plays an important role in excluding anionic interferences. We also prepare a sensitive (16.35 µA mM-1), low LOD (0.13 µM) and wide linear range (1-70 µM) amperometric sarcosine biosensor with excellent anti-interference properties. This mesoporous material provides a bio-composite platform for the development of simple amperometric biosensors for detecting micromoles per liter of analytes in serum or urine.

Quantitative analysis of sarcosine with special emphasis on biosensors: a review.

The quantitative determination of sarcosine is of great importance in clinical chemistry, food and fermentation industries. Elevated sarcosine levels are associated with Alzheimer, dementia, prostate cancer, colorectal cancer, stomach cancer and sarcosinemia. This review summarizes the various methods for quantitative analysis of sarcosine with special emphasis on various strategies of biosensors and their analytical performance. The current bio sensing methods have overcome the drawbacks of conventional methods. Sarcosine biosensors work optimally at pH 7.0 to 8.0 in the linear range of 0.1 to 100 µM within 2 to 17 s and between 25 and 37 °C, within a limit of detection (LOD) between 0.008 and 500 mM. The formulated biosensors can be reused within a stability period of 3-180 days. Future research could be focused to modify existing sarcosine biosensors, leading to simple, reliable, and economical sensors ideally suited for point-of-care treatment. Clinical significance Elevated sarcosine levels are associated with prostate and colorectal cancer, Alzheimer, dementia, stomach cancer and sarcosinemia. Quantitative determination of sarcosine is of great importance in clinical chemistry as well as food and fermentation industries. Attempts made in development of sarcosine biosensors have been reviewed with their advantages and disadvantages, so that scientist and clinicians can improvise the methods of developing more potent sarcosine biosensor applicable in multitudinous fields. This is the first comprehensive review which compares the various immobilization methods, sensing principles, strategies used in biosensors and their analytical performance in detail.

Maternal betaine status, but not that of choline or methionine, is inversely associated with infant birth weight.

Maternal one-carbon metabolism during pregnancy is crucial for fetal development and programming by DNA methylation. However, evidence on one-carbon biomarkers other than folate is lacking. We, therefore, investigated whether maternal plasma methyl donors, that is, choline, betaine and methionine, are associated with birth outcomes. Blood samples were obtained from 115 women during gestation (median 26·3 weeks, 90 % range 22·7-33·0 weeks). Plasma choline, betaine, methionine and dimethylglycine were measured using HPLC-tandem MS. Multivariate linear and logistic regression models were used to estimate the association between plasma biomarkers and birth weight, birth length, the risk of small-for-gestational-age and large-for-gestational-age (LGA). Higher level of maternal betaine was associated with lower birth weight (-130·3 (95 % CI -244·8, -15·9) per 1 sd increment for log-transformed betaine). Higher maternal methionine was associated with lower risk of LGA, and adjusted OR, with 95 % CI for 1 sd increase in methionine concentration was 0·44 (95 % CI 0·21, 0·89). Stratified analyses according to infant sex or maternal plasma homocysteine status showed that reduction in birth weight in relation to maternal betaine was only limited to male infants or to who had higher maternal homocysteine status (≥5·1 µmol/l). Higher maternal betaine status was associated with reduced birth weight. Maternal methionine was inversely associated with LGA risk. These findings are needed to be replicated in future larger studies.

Association between one-carbon metabolism indices and DNA methylation status in maternal and cord blood.

One-carbon metabolism is essential for multiple cellular processes and can be assessed by the concentration of folate metabolites in the blood. One-carbon metabolites serve as methyl donors that are required for epigenetic regulation. Deficiencies in these metabolites are associated with a variety of poor health outcomes, including adverse pregnancy complications. DNA methylation is known to vary with one-carbon metabolite concentration, and therefore may modulate the risk of adverse pregnancy outcomes. This study addresses changes in one-carbon indices over pregnancy and the relationship between maternal and child DNA methylation and metabolite concentrations by leveraging data from 24 mother-infant dyads. Five of the 13 metabolites measured from maternal blood and methylation levels of 993 CpG sites changed over the course of pregnancy. In dyads, maternal and fetal one-carbon concentrations were highly correlated, both early in pregnancy and at delivery. The 993 CpG sites whose methylation levels changed over pregnancy in maternal blood were also investigated for associations with metabolite concentrations in infant blood at delivery, where five CpG sites were associated with the concentration of at least one metabolite. Identification of CpG sites that change over pregnancy may result in better characterization of genes and pathways involved in maintaining a healthy, term pregnancy.

Sarcosine Is Uniquely Modulated by Aging and Dietary Restriction in Rodents and Humans.

A hallmark of aging is a decline in metabolic homeostasis, which is attenuated by dietary restriction (DR). However, the interaction of aging and DR with the metabolome is not well understood. We report that DR is a stronger modulator of the rat metabolome than age in plasma and tissues. A comparative metabolomic screen in rodents and humans identified circulating sarcosine as being similarly reduced with aging and increased by DR, while sarcosine is also elevated in long-lived Ames dwarf mice. Pathway analysis in aged sarcosine-replete rats identify this biogenic amine as an integral node in the metabolome network. Finally, we show that sarcosine can activate autophagy in cultured cells and enhances autophagic flux in vivo, suggesting a potential role in autophagy induction by DR. Thus, these data identify circulating sarcosine as a biomarker of aging and DR in mammalians and may contribute to age-related alterations in the metabolome and in proteostasis.

Glucose and Fat Tolerance Tests Induce Differential Responses in Plasma Choline Metabolites in Healthy Subjects.

Plasma choline shows associations with plasma glucose and lipids. We studied changes of choline metabolites after oral glucose tolerance test (OGTT) and fat tolerance test (OFTT). Eighteen healthy subjects (mean age 54.3 years; BMI 26.8 kg/m²) underwent 2 tests. First, OFTT (80 g fat) was applied and blood was collected at baseline and 4 h after OFTT. Seven days later, 75 g glucose was applied and blood was collected at baseline and 2 h after OGTT. Plasma concentrations of choline, betaine, trimethylamine N-oxide (TMAO), dimethylglycine, S-adenosylmethionine (SAM), lipids and glucose were measured. After OFTT, plasma choline declined (10.6 to 9.2 µmol/L; p = 0.004), betaine declined (33.4 to 31.7 µmol/L; p = 0.003), TMAO slightly increased (4.1 to 5.6 µmol/L; p = 0.105), glucose declined (5.39 to 4.98 mmol/L; p < 0.001), and triglycerides increased (1.27 to 2.53 mmol/L; p < 0.001). After OGTT, plasma choline increased (10.1 to 11.1 µmol/L; p < 0.001), TMAO declined (4.0 to 3.5 µmol/L; p = 0.029), dimethylglycine declined (2.0 to 1.7 µmol/L; p = 0.005), SAM declined (103 to 96 nmol/L; p = 0.041), but betaine, glucose, and SAM were unchanged. In conclusion, OFTT lowered plasma betaine and choline and caused heterogeneous changes in plasma TMAO. OGTT reduced the flow of methyl groups and plasma TMAO.

An integrated portable system for single chip simultaneous measurement of multiple disease associated metabolites.

Metabolites, the small molecules that underpin life, can act as indicators of the physiological state of the body when their abundance varies, offering routes to diagnosis of many diseases. The ability to assay for multiple metabolites simultaneously will underpin a new generation of precision diagnostic tools. Here, we report the development of a handheld device based on complementary metal oxide semiconductor (CMOS) technology with multiple isolated micro-well reaction zones and integrated optical sensing allowing simultaneous enzyme-based assays of multiple metabolites (choline, xanthine, sarcosine and cholesterol) associated with multiple diseases. These metabolites were measured in clinically relevant concentration range with minimum concentrations measured: 25 µM for choline, 100 µM for xanthine, 1.25 µM for sarcosine and 50 µM for cholesterol. Linking the device to an Android-based user interface allows for quantification of metabolites in serum and urine within 2 min of applying samples to the device. The quantitative performance of the device was validated by comparison to accredited tests for cholesterol and glucose.

Construction and application of amperometric sarcosine biosensor based on SOxNPs/AuE for determination of prostate cancer.

An improved amperometric sarcosine biosensor was constructed based on covalent immobilization of sarcosine oxidase nanoparticles (SOxNPs) onto gold electrode (AuE). The SOxNPs/AuE was characterized by scanning electron microscopy (SEM), fourier transform infrared (FTIR) spectroscopy and electrochemical impedance spectroscopy (EIS) at different stages of its construction. The biosensor worked optimally within 2 s at a potential of 1.0 V, against Ag/AgCl, pH 6.5 and 35 °C. A linear relationship was observed between sarcosine concentration range, 0.1-100 µM and the biosensor response i.e. current in mA under optimum conditions. The biosensor offered a low detection limit of 0.01 µM and gratifying storage stability. The SOxNPs/AuE was unaffected by a number of serum substances at their physiological concentrations. The biosensor measured sarcosine level in sera collected from persons suffering from prostate cancer (mean13.5 µM, n = 8), which was significantly higher (p < 0.01) than those in apparently healthy persons (mean 2.2 µM, n = 8). The SOxNPs/Au electrode was reused 300- times during the span of 180 days, with only 10% loss in its initial activity while being stored dry at 4 °C.

Influence of collection tubes during quantitative targeted metabolomics studies in human blood samples.

BACKGROUND: Plasma and serum are the most widely used matrices in clinical studies. However, some variability in absolute concentrations of metabolites are likely to be observed in these collection tubes matrices. METHODS: We analyzed 189 metabolites using the same protocol for quantitative targeted metabolomics (LC-MS/MS AbsoluteIDQ p180 Kit Biocrates) in three types of samples, serum, plasma EDTA and citrate, of 80 subjects from the Cooperative Health Research In South Tyrol cohort (40 healthy elderly and 40 healthy young). RESULTS: The concentration levels were higher in serum than citrate and EDTA, in particular for amino acids and biogenic amines. The average Pearson's correlation coefficients were however always higher than 0.7 for these two classes of metabolites. We could also demonstrate that blank EDTA vacutainer tubes contain a significant amount of sarcosine. Finally, we compared the metabolome of young people against elderly subjects and found that the highest number of metabolites significantly changing with age was detected in serum. CONCLUSION: Serum samples provide higher sensitivity for biomarker discovery studies. Due to the presence of spurious amount of sarcosine in vacutainer EDTA tubes, plasma EDTA is not suitable for studies requiring accurate quantification of sarcosine.