Natural Protein Photon Upconversion Supramolecular Assemblies for Background-Free Biosensing.

The diagnosis of disease biomarkers is crucial for the identification, monitoring, and prognostic assessment of malignant disease. However, biological samples with autofluorescence, complex components, and heterogeneity pose major challenges to reliable biosensing. Here, we report the self-assembly of natural proteins and the triplet-triplet annihilation upconversion (TTA-UC) pair to form upconverted protein clusters (∼8.2 ± 1.1 nm), which were further assembled into photon upconversion supramolecular assemblies (PUSA). This PUSA exhibited unique features, including a small size (∼44.1 ± 4.1 nm), oxygen tolerance, superior biocompatibility, and easy storage via lyophilization, all of which are long sought after for photon upconversion materials. Further, we have revealed that the steric hindrance of the annihilator suppresses the stacking of the annihilator in PUSA, which is vital for maintaining the water dispersibility and enhancing the upconversion performance of PUSA. In conjunction with sarcosine oxidase, this near infrared (NIR)-excitable PUSA nanoprobe could perform background-free biosensing of urinary sarcosine, which is a common biomarker for prostatic carcinoma (PCa). More importantly, this nanoprobe not only allows for qualitative identification of urinary samples from PCa patients by the unaided eye under NIR-light-emitting diode (LED) illumination but also quantifies the concentration of urinary sarcosine. These remarkable findings have propelled photon upconversion materials to a new evolutionary stage and expedited the progress of upconversion biosensing in clinical diagnostics.

Facile one-step targeted immobilization of an enzyme based on silane emulsion self-assembled molecularly imprinted polymers for visual sensors.

Immobilized enzymes play significant roles in many practical applications. However, the enzymes need to be purified before immobilization by conventional immobilizing methods, and the purification process is expensive, laborious, complicated and results in a decrease of the enzymatic activity. So, we present a novel method by a facile one-step targeted immobilization of an enzyme without a purification process from complex samples. For this purpose, a novel molecularly imprinted polymer was prepared via a silane emulsion self-assembly method using boric acid-modified Fe3O4 nanoparticles as magnetic nuclei, horseradish peroxidase as a template, 3-aminopropyltriethoxysilane as a functional monomer and tetraethyl orthosilicate as a crosslinking agent. The molecularly imprinted polymers were characterized using a scanning electron microscope, X-ray photoelectron spectroscope, vibrating sample magnetometer and X-ray diffractometer. The as-prepared and characterized materials were employed to immobilize horseradish peroxidase from a crude extract of horseradish. Moreover, the immobilized horseradish peroxidase was employed to develop visual sensors for the detection of glucose and sarcosine. This study demonstrated that the molecularly imprinted polymers prepared via the silane emulsion self-assembly method can facilely immobilize horseradish peroxidase from a crude extract of horseradish without any purification process. The developed visual method based on the immobilized horseradish peroxidase shows great potential applications for the visual detection of glucose and sarcosine.

An electrochemical sarcosine sensor based on biomimetic recognition.

A nonenzymatic electrochemical sensor is described for the prostate cancer biomarker sarcosine (Sar). Riboflavin was employed to mimic the active center of the enzyme sarcosine oxidase for constructing the biomimetic sensor. The use of riboflavon (Rf) avoids the disadvantages of an enzymatic sensor, such as high cost and poor stability. A glassy carbon electrode (GCE) was modified with a graphene-chitosan (GR) composite and further modified with gold-platinum bimetallic nanoparticles in a polypyrrole (PPy) matrix in order to enhance the catalytic activity of the enzyme mimic. Finally, Rf was electrodeposited on the surface of the AuPt-PPy/GR-modified GCE. Under optimized conditions, the GCE provided high sensitivity and selectivity for Sar at around 0.61 V. Response covers the 2.5-600 µM concentration range, and the detection limit is 0.68 µM. The method was successfully applied to the determination of Sar in spiked urine with 98.0%-103.2% recovery. Graphical abstract Schematic presentation of the fabrication of the Rf/AuPt-PPy/GR/GCE surface and the measurement principle by differential pulse voltammetry (DPV).

Bidirectional Electrochemiluminescence Color Switch: An Application in Detecting Multimarkers of Prostate Cancer.

A selective excitation of [Ir(df-ppy)2(pic)] and [Ru(bpy)3]2+ through tuning the electrode potential is reported in this work. Bidirectional color change from blue-green to red could be observed along with increase and decrease of the potential, which was ascribed to the dual-potential excitation property of [Ir(df-ppy)2(pic)]. Similar to the three-electrode system, selective excitation of ECL could be achieved at the anode of the bipolar electrode (BPE). Both increase and decrease of the faradic reactions at the cathode of the BPE could induce ECL reporting color at the other pole switched from blue-green to red. We applied a closed BPE device for the bioanalysis of multicolor ECL since the organic solvent containing electrochemiluminophores could be separated from the bioanalytes. On the basis of BPE arrays coupled with the ECL switch, the detection of three biomarkers of prostate cancer, PSA, microRNA-141, and sarcosine were integrated in a same device. The cutoff values of the biomarkers could be recognized directly by the naked eye. Such a device holds great potential in the early diagnosis of prostate cancer.

A fluorescent probe based on a Tb3+/Cu2+ co-functionalized MOF for urinary sarcosine detection.

A novel luminescent probe based on a Tb3+/Cu2+ heterometallic metal-organic framework (MOF) was first designed for beamed monitoring of urinary sarcosine, a differential metabolite that can indicate the progression of prostate cancer (PCa). The fluorescent probe presented high selectivity towards sarcosine in urine. It also displayed good sensitivity with a comparatively low detection limit and a fast response to sarcosine within 5 min. Moreover, such high selectivity and sensitivity towards sarcosine is not subject to interference from other coexisting species in urine. At the same time, this fluorescent material also demonstrated the possibility for recycling. The excellent sensing performance of this Ln-MOF (lanthanide MOF) enables it to be further employed as a serviceable tool for PCa diagnosis and monitoring.

Three-phase solvent bar liquid-phase microextraction combined with high-performance liquid chromatography to determine sarcosine in human urine.

Sarcosine is a potential prostate cancer marker. In this study, we developed a method of three-phase solvent bar liquid-phase microextraction combined with high-performance liquid chromatography to determine sarcosine after derivatization with 4-dimethylarminoazobenzene-4-sulfonyl chloride from human urine. The effects of different extraction conditions on extraction efficiency were investigated and optimized. Under optimum experimental conditions, a calibration graph exhibited linearity over the range of 0.05-25 µmol/L with a correlation coefficient (r2 ) of 0.9990. The enrichment factor was 168, and the detection limit was 0.02 µmol/L. The method was successfully used to analyze sarcosine in human urine and non-invasive detection, and good spiked recoveries ranging from 90.5 to 93.6% were obtained. The proposed method exhibited high sensitivity, high enrichment factor, good precision, and a simple setup. It may contribute to the early accurate diagnosis and the progression monitoring of prostatic carcinoma.

The Urinary Sarcosine/Creatinine Ratio is a Potential Diagnostic and Prognostic Marker in Prostate Cancer.

BACKGROUND The aim of this study was to evaluate the role of the urinary sarcosine/creatinine ratio in the diagnosis and prognosis of prostate cancer, using a sarcosine oxidase assay. MATERIAL AND METHODS Urine samples were obtained from 203 patients with benign prostate hyperplasia (BPH) and 209 patients with prostate cancer. Levels of urinary sarcosine were measured using the sarcosine oxidase method. The urinary sarcosine/creatinine ratios were compared between the group of patients with BPH and the patients with prostate cancer. In the two patients groups, the urinary sarcosine/creatinine ratio was compared with the measurement of serum prostate-specific antigen (PSA) and the free/total (F/T) PSA ratio. Correlations between of the urinary sarcosine/creatinine ratio and the Gleason grade and stage of prostate cancer were analyzed. RESULTS There was a significant difference between the urinary sarcosine/creatinine ratio in the BPH group and prostate cancer group (P<0.01), which was independent of serum PSA. The receiver-operating characteristic (ROC) curve, and area under the curve (AUC) for the urinary sarcosine/creatinine ratio was significantly higher compared with the serum PSA and the F/T PSA ratio. There was a significant difference in the urinary sarcosine/creatinine ratio in patients with prostate cancer with Gleason score ≤6, 7, and ≥8, and between patients with metastatic and non-metastatic prostate cancer. CONCLUSIONS The urinary sarcosine/creatinine ratio was a diagnostic indicator of prostate cancer, for patients with a serum PSA level <10 ng/ml, and correlated with the Gleason score and with the presence of metastases (stage) of prostate cancer.

A Rapid Method for the Detection of Sarcosine Using SPIONs/Au/CS/SOX/NPs for Prostate Cancer Sensing.

BACKGROUND: Sarcosine is an amino acid that is formed by methylation of glycine and is present in trace amounts in the body. Increased sarcosine concentrations in blood plasma and urine are manifested in sarcosinemia and in some other diseases such as prostate cancer. For this purpose, sarcosine detection using the nanomedicine approach was proposed. In this study, we have prepared superparamagnetic iron oxide nanoparticles (SPIONs) with different modified surface area. Nanoparticles (NPs) were modified by chitosan (CS), and sarcosine oxidase (SOX). SPIONs without any modification were taken as controls. Methods and Results: The obtained NPs were characterized by physicochemical methods. The size of the NPs determined by the dynamic light scattering method was as follows: SPIONs/Au/NPs (100⁻300 nm), SPIONs/Au/CS/NPs (300⁻700 nm), and SPIONs/Au/CS/SOX/NPs (600⁻1500 nm). The amount of CS deposited on the NP surface was found to be 48 mg/mL for SPIONs/Au/CS/NPs and 39 mg/mL for SPIONs/Au/CS/SOX/NPs, and repeatability varied around 10%. Pseudo-peroxidase activity of NPs was verified using sarcosine, horseradish peroxidase (HRP) and 3,3',5,5'-tetramethylbenzidine (TMB) as a substrate. For TMB, all NPs tested evinced substantial pseudo-peroxidase activity at 650 nm. The concentration of SPIONs/Au/CS/SOX/NPs in the reaction mixture was optimized to 0⁻40 mg/mL. Trinder reaction for sarcosine detection was set up at 510 nm at an optimal reaction temperature of 37 °C and pH 8.0. The course of the reaction was linear for 150 min. The smallest amount of NPs that was able to detect sarcosine was 0.2 mg/well (200 µL of total volume) with the linear dependence y = 0.0011x - 0.0001 and the correlation coefficient r = 0.9992, relative standard deviation (RSD) 6.35%, limit of detection (LOD) 5 µM. The suggested method was further validated for artificial urine analysis (r = 0.99, RSD 21.35%, LOD 18 µM). The calculation between the detected and applied concentrations showed a high correlation coefficient (r = 0.99). NPs were tested for toxicity and no significant growth inhibition was observed in any model system (S. cerevisiae, S. aureus, E. coli). The hemolytic activity of the prepared NPs was similar to that of the phosphate buffered saline (PBS) control. The reaction system was further tested on real urine specimens. Conclusion: The proposed detection system allows the analysis of sarcosine at micromolar concentrations and to monitor changes in its levels as a potential prostate cancer marker. The whole system is suitable for low-cost miniaturization and point-of-care testing technology and diagnostic systems. This system is simple, inexpensive, and convenient for screening tests and telemedicine applications.

Is N,N-dimethylglycine N-oxide a choline and betaine metabolite?

Choline metabolism is by oxidation to betaine, which is demethylated to N,N-dimethylglycine; dimethylglycine is oxidatively demethylated to sarcosine. This pathway is important for osmoregulation and as a source of methyl groups. We asked whether another metabolite was involved. We synthesized the N-oxide of dimethylglycine (DMGO) by oxidizing dimethylglycine with peracetic acid, and measured DMGO in human plasma and urine by HPLC-MS/MS with positive ion detection, using two chromatography procedures, based on ion exchange and HILIC separations. The molecular ion DMGOH+ (m/z=120) yielded four significant fragments (m/z=103, 102, 58 and 42). The suspected DMGO peak in human body fluids showed all these fragments, and co-chromatographed with added standard DMGO in both HPLC systems. Typical plasma concentrations of DMGO are under 1 µmol/l. They may be lower in metabolic syndrome patients. Urine concentrations are higher, and DMGO has a higher fractional clearance than dimethylglycine, betaine and choline. It was present in all of over 80 human urine and plasma samples assayed. Plasma DMGO concentrations correlate with plasma DMG concentrations, with betaine and choline concentrations, with the osmolyte myo-inositol, and strongly with urinary DMGO excretion. We conclude that DMGO is probably a normal human metabolite.

Biomarkers in localized prostate cancer.

Biomarkers can improve prostate cancer diagnosis and treatment. Accuracy of prostate-specific antigen (PSA) for early diagnosis of prostate cancer is not satisfactory, as it is an organ- but not cancer-specific biomarker, and it can be improved by using models that incorporate PSA along with other test results, such as prostate cancer antigen 3, the molecular forms of PSA (proPSA, benign PSA and intact PSA), as well as kallikreins. Recent reports suggest that new tools may be provided by metabolomic studies as shown by preliminary data on sarcosine. Additional molecular biomarkers have been identified by the use of genomics, proteomics and metabolomics. We review the most relevant biomarkers for early diagnosis and management of localized prostate cancer.

Influence of Long-Distance Bicycle Riding on Serum/Urinary Biomarkers of Prostate Cancer.

Herein, we present a study focused on the determination of the influence of long-distance (53 km) bicycle riding on levels of chosen biochemical urinary and serum prostate cancer (PCa) biomarkers total prostate-specific antigen (tPSA), free PSA (fPSA) and sarcosine. Fourteen healthy participants with no evidence of prostate diseases, in the age range from 49-57 years with a median of 52 years, underwent physical exercise (mean race time of 150 ± 20 min, elevation increase of 472 m) and pre- and post-ride blood/urine sampling. It was found that bicycle riding resulted in elevated serum uric acid (p = 0.001, median 271.76 vs. 308.44 µmol/L pre- and post-ride, respectively), lactate (p = 0.01, median 2.98 vs. 4.8 mmol/L) and C-reactive protein (p = 0.01, 0.0-0.01 mg/L). It is noteworthy that our work supports the studies demonstrating an increased PSA after mechanical manipulation of the prostate. The subjects exhibited either significantly higher post-ride tPSA (p = 0.002, median 0.69 vs. 1.1 ng/mL pre- and post-ride, respectively) and fPSA (p = 0.028, median 0.25 vs. 0.35 ng/mL). Contrary to that, sarcosine levels were not significantly affected by physical exercise (p = 0.20, median 1.64 vs. 1.92 µmol/mL for serum sarcosine, and p = 0.15, median 0.02 µmol/mmol of creatinine vs. 0.01 µmol/mmol of creatinine for urinary sarcosine). Taken together, our pilot study provides the first evidence that the potential biomarker of PCa-sarcosine does not have a drawback by means of a bicycle riding-induced false positivity, as was shown in the case of PSA.

Discovery and validation of urinary metabotypes for the diagnosis of hepatocellular carcinoma in West Africans.

UNLABELLED: There is no clinically applicable biomarker for surveillance of hepatocellular carcinoma (HCC), because the sensitivity of serum alpha-fetoprotein (AFP) is too low for this purpose. Here, we determined the diagnostic performance of a panel of urinary metabolites of HCC patients from West Africa. Urine samples were collected from Nigerian and Gambian patients recruited on the case-control platform of the Prevention of Liver Fibrosis and Cancer in Africa (PROLIFICA) program. Urinary proton nuclear magnetic resonance ((1) H-NMR) spectroscopy was used to metabolically phenotype 290 subjects: 63 with HCC; 32 with cirrhosis (Cir); 107 with noncirrhotic liver disease (DC); and 88 normal control (NC) healthy volunteers. Urine samples from a further cohort of 463 subjects (141 HCC, 56 Cir, 178 DC, and 88 NC) were analyzed, the results of which validated the initial cohort. The urinary metabotype of patients with HCC was distinct from those with Cir, DC, and NC with areas under the receiver operating characteristic (AUROC) curves of 0.86 (0.78-0.94), 0.93 (0.89-0.97), and 0.89 (0.80-0.98) in the training set and 0.81 (0.73-0.89), 0.96 (0.94-0.99), and 0.90 (0.85-0.96), respectively, in the validation cohort. A urinary metabolite panel, comprising inosine, indole-3-acetate, galactose, and an N-acetylated amino acid (NAA), showed a high sensitivity (86.9% [75.8-94.2]) and specificity (90.3% [74.2-98.0]) in the discrimination of HCC from cirrhosis, a finding that was corroborated in a validation cohort (AUROC: urinary panel = 0.72; AFP = 0.58). Metabolites that were significantly increased in urine of HCC patients, and which correlated with clinical stage of HCC, were NAA, dimethylglycine, 1-methylnicotinamide, methionine, acetylcarnitine, 2-oxoglutarate, choline, and creatine. CONCLUSION: The urinary metabotyping of this West African cohort identified and validated a metabolite panel that diagnostically outperforms serum AFP.

An electrochemiluminescence-supramolecular approach to sarcosine detection for early diagnosis of prostate cancer.

Monitoring Prostate Cancer (PCa) biomarkers is an efficient way to diagnosis this disease early, since it improves the therapeutic success rate and suppresses PCa patient mortality: for this reason a powerful analytical technique such as electrochemiluminescence (ECL) is already used for this application, but its widespread usability is still hampered by the high cost of commercial ECL equipment. We describe an innovative approach for the selective and sensitive detection of the PCa biomarker sarcosine, obtained by a synergistic ECL-supramolecular approach, in which the free base form of sarcosine acts as co-reagent in a Ru(bpy)3(2+)-ECL process. We used magnetic micro-beads decorated with a supramolecular tetraphosphonate cavitand (Tiiii) for the selective capture of sarcosine hydrochloride in a complex matrix like urine. Sarcosine determination was then obtained with ECL measurements thanks to the complexation properties of Tiiii, with a protocol involving simple pH changes - to drive the capture-release process of sarcosine from the receptor - and magnetic micro-bead technology. With this approach we were able to measure sarcosine in the µM to mM window, a concentration range that encompasses the diagnostic urinary value of sarcosine in healthy subjects and PCa patients, respectively. These results indicate how this ECL-supramolecular approach is extremely promising for the detection of sarcosine and for PCa diagnosis and monitoring, and for the development of portable and more affordable devices.

Metabolomic profiles delineate potential role for sarcosine in prostate cancer progression.

Multiple, complex molecular events characterize cancer development and progression. Deciphering the molecular networks that distinguish organ-confined disease from metastatic disease may lead to the identification of critical biomarkers for cancer invasion and disease aggressiveness. Although gene and protein expression have been extensively profiled in human tumours, little is known about the global metabolomic alterations that characterize neoplastic progression. Using a combination of high-throughput liquid-and-gas-chromatography-based mass spectrometry, we profiled more than 1,126 metabolites across 262 clinical samples related to prostate cancer (42 tissues and 110 each of urine and plasma). These unbiased metabolomic profiles were able to distinguish benign prostate, clinically localized prostate cancer and metastatic disease. Sarcosine, an N-methyl derivative of the amino acid glycine, was identified as a differential metabolite that was highly increased during prostate cancer progression to metastasis and can be detected non-invasively in urine. Sarcosine levels were also increased in invasive prostate cancer cell lines relative to benign prostate epithelial cells. Knockdown of glycine-N-methyl transferase, the enzyme that generates sarcosine from glycine, attenuated prostate cancer invasion. Addition of exogenous sarcosine or knockdown of the enzyme that leads to sarcosine degradation, sarcosine dehydrogenase, induced an invasive phenotype in benign prostate epithelial cells. Androgen receptor and the ERG gene fusion product coordinately regulate components of the sarcosine pathway. Here, by profiling the metabolomic alterations of prostate cancer progression, we reveal sarcosine as a potentially important metabolic intermediary of cancer cell invasion and aggressivity.

On-line determination of sarcosine in biological fluids utilizing dummy molecularly imprinted polymers in microextraction by packed sorbent.

Several years ago, sarcosine received attention as a prostate-cancer marker. Prostate cancer is one of the most widespread types of tumor diseases in men. The prostate-specific antigen is normally used as a marker, and it can only be detected in blood with a sensitivity of approximately 80%. In the present study, dummy molecularly imprinted polymers in microextraction by packed sorbent with on-line liquid chromatography coupled to tandem mass spectrometry was used for the determination of sarcosine in human plasma and urine samples. The polymer network glycine was used for the dummy molecularly imprinted polymers. The selectivity of the method was evaluated using similar prostate-cancer biomarkers. In addition, various parameters affecting the extraction performance were investigated. The method limits of detection and quantification in the plasma and urine were 1.0 and 3.0 ng/mL, respectively. The values of the coefficient of determination were over 0.99 for all runs in the studied concentration range (3.0-10 000 ng/mL). The method recovery was 87 and 89% in plasma and urine, respectively. The intraday and interday precisions of sarcosine in the plasma and urine samples were in the ranges of 4.0-7.1, 3.0-6.3, 2.9-4.7, and 5.0-6.7, respectively.

Exclusive recognition of sarcosine in water and urine by a cavitand-functionalized silicon surface.

A supramolecular approach for the specific detection of sarcosine, recently linked to the occurrence of aggressive prostate cancer forms, has been developed. A hybrid active surface was prepared by the covalent anchoring on Si substrates of a tetraphosphonate cavitand as supramolecular receptor and it was proven able to recognize sarcosine from its nonmethylated precursor, glycine, in water and urine. The entire complexation process has been investigated in the solid state, in solution, and at the solid-liquid interface to determine and weight all the factors responsible of the observed specificity. The final outcome is a Si-based active surface capable of binding exclusively sarcosine. The complete selectivity of the cavitand-decorated surface under these stringent conditions represents a critical step forward in the use of these materials for the specific detection of sarcosine and related metabolites in biological fluids.

Supramolecular sensing with phosphonate cavitands.

Molecular recognition is a recurrent theme in chemical sensing because of the importance of selectivity for sensor performances. The popularity of molecular recognition in chemical sensing has resulted from the progress made in mastering weak interactions, which has enabled the design of synthetic receptors according to the analyte to be detected. However, the availability of a large pool of modular synthetic receptors so far has not had a significant impact on sensors used in the real world. This technological gap has emerged because of the difficulties in transferring the intrinsic molecular recognition properties of a given receptor from solution to interfaces and in finding high fidelity transduction modes for the recognition event. This Account focuses on the ways to overcome these two bottlenecks, and we recount our recent efforts to produce highly selective supramolecular sensors using phosphonate cavitands as receptors. Through two examples, we present an overview of the different operating strategies that are implemented depending on whether the interface is vapor-solid or liquid-solid. First we describe the selective detection of short chain aliphatic alcohols in the vapor phase. In this example, we solved a key issue common to all sensors for organic vapors: the dissection of the specific interaction (between cavitand and the alcohol) from ubiquitous nonspecific dispersion interactions (between the analytes and interferents in the solid layer). We removed responses resulting from the nonspecific interactions of the analytes with interferents by directly connecting the recognition event at the interface to the transduction mechanism (photoinduced charge transfer). The second example addresses the specific detection of sarcosine in urine. Recent research has suggested that sarcosine can serve as reliable biomarker of the aggressive forms of prostate cancer. Tetraphosphonate cavitands can complex N-methyl ammonium salts with impressive selectivity in solution, and we used this property as a starting point. The sensor implementation requires that we first graft the cavitand onto silicon and gold surfaces as monolayers. The exclusive recognition of sarcosine by these supramolecular sensors originates from their operation in aqueous environments, where synergistic multiple interactions with the phosphonate cavitand are possible only for N-methyl ammonium derivatives. We couple that selectivity with detection modes that probe the strength of the complexation either directly (microcantilever) or via exchange with molecules that have comparable affinity for the cavity (fluorescence dye displacement).

Extreme urinary betaine losses in type 2 diabetes combined with bezafibrate treatment are associated with losses of dimethylglycine and choline but not with increased losses of other osmolytes.

PURPOSE: Betaine deficiency is a probable cardiovascular risk factor and a cause of elevated homocysteine. Urinary betaine excretion is increased by fibrate treatment, and is also often elevated in diabetes. Does fibrate further increase betaine excretion in diabetes, and does it affect the plasma concentrations and excretions of related metabolites and of other osmolytes? METHODS: Samples from a previous study of type 2 diabetes were selected if participants were taking bezafibrate (n = 32). These samples were compared with participants matched for age and gender and not on a fibrate (comparator group, n = 64). Betaine, related metabolites, and osmolytes were measured in plasma and urine samples from these 96 participants. RESULTS: Median urinary betaine excretion in those on bezafibrate was 5-fold higher than in the comparator group (p < 0.001), itself 3.5-fold higher than the median reported for healthy populations. In the bezafibrate group, median dimethylglycine excretion was higher (9-fold, p < 0.001). Excretions of choline, and of the osmolytes myo-inositol, taurine and glycerophosphorylcholine, were not significantly different between groups. Some participants excreted more betaine than usual dietary intakes. Several betaine fractional clearances were >100 %. Betaine excretion correlated with excretions of the osmolytes myo-inositol and glycerophosphorylcholine, and also with the excretion of choline and N,N-dimethylglycine, but it was inconclusive whether these relationships were affected by bezafibrate therapy. CONCLUSIONS: Increased urinary betaine excretions in type 2 diabetes are further increased by fibrate treatment, sometimes to more than their dietary intake. Concurrent betaine supplementation may be beneficial.

Preconcentration based on paramagnetic microparticles for the separation of sarcosine using hydrophilic interaction liquid chromatography coupled with coulometric detection.

Sarcosine has been identified as a potential prostate cancer marker. To provide determination of this compound, a number of methods are developing. In this study, we optimized a method for its separation by hydrophilic interaction LC with electrochemical detection (ED). Due to the fact that mobile phases commonly used for this type of separation altered the LODs measured by electrochemical detectors, we applied postcolumn dosing of buffer suitable for ED. The optimized conditions were mobile phase A acetonitrile, mobile phase B water in the ratio A/B 70:30, with postcolumn addition of mobile phase C (200 mM phosphate buffer pH 9). The optimal mixing ratio was A + B/C 1:1 with a flow rate of 0.80 mL/min (0.40 + 0.40 mL/min) and detection potential of 1000 mV. Due to the optimization of the parameters for effective separation, which had to meet the optimal parameters of ED, we reached a good resolution for separation also with a good LOD (100 nM). In addition, we successfully carried out sarcosine analysis bound on our modified paramagnetic microparticles with the ability to preconcentrate sarcosine isolated from artificial urine.

Sensitive determination of the potential biomarker sarcosine for prostate cancer by LC-MS with N,N'-dicyclohexylcarbodiimide derivatization.

Sarcosine, a potential biomarker of prostate cancer, has drawn great attention in recent years. However, controversial research keeps arising about its role as a biomarker that might come from the two isomers (α-alanine and ß-alanine) of sarcosine due to their same molecular weight and similar properties, which could interfere with the accurate detection of sarcosine. In this study, a simple and sensitive method was developed for the detection of sarcosine and the two isomers by LC with ion-trap MS through a novel derivatization reagent N,N'-dicyclohexylcarbodiimide. N,N'-Dicyclohexylcarbodiimide is usually considered as a condensation reagent, however, it was directly used as a derivatization reagent through a rearrangement side reaction in this study. The proposed method not only improved the chromatographic retention behavior of sarcosine and its two isomers, which was a benefit to their separation, but also dramatically enhanced the detection sensitivity of sarcosine, which was more favorable for real sample analysis. The factors affecting the productivity of the derivatization reaction, such as reaction time and amount of derivatization reagent, were systematically optimized. The method shows good linearity (R(2) > 0.99), sensitivity with LODs of sarcosine as low as 1 ng/mL, and repeatability with the RSD < 6.07%. The developed method was applied to the analysis of urine.