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Introduction: Asymmetric dimethylarginine (ADMA) competitively inhibits nitric oxide synthase (NOS). Its levels in specimens from murine models and asthmatic patients are related to inflammation and oxidative stress. Patients with cystic fibrosis(CF) reportedly have higher arginase activity, lower NO production and NOS expression than healthy controls., Objective: The objective was to assess the role of ADMA and related metabolites as disease biomarkers in exhaled breath condensate (EBC) of pediatric CF patients, compared with age-matched healthy controls (HC)., Methods: A longitudinal design was conceived and 34 CF patients (21 stable, 13 at the onset of exacerbation) and 16 HC were enrolled. All CF patients underwent clinical examination, spirometry and EBC collection at enrolment; the same tests were performed also after an antibiotic course in those patients with exacerbation. Metabolites levels in EBC were measured with an ultra-performance liquid chromatography and tandem mass spectrometry technique., Results: All CF patients had ADMA levels (expressed as ratio to tyrosine) similar to those in HC (median 0.0112, IQR 0.0103-0.0120 and median 0.0114, IQR 0.0090-0.0128, respectively; P = 0.983), while a significant increase in the citrulline/tyrosine ratio was found in CF patients (median 0.6419, IQR 0.5738-0.6899 in CF vs median 0.4176, IQR 0.2986-0.5082 in HC; P = 0.00003). No differences in ADMA levels emerged between stable patients and those with exacerbation., Conclusion: ADMA and related aminoacids were measured simultaneously for the first time in EBC from CF patients. Higher citrulline/tyrosine ratios were found in CF children with normal ADMA levels, suggesting a dysregulated ADMA metabolism in these patients., (© 2016 John Wiley & Sons Ltd.)

Background: Exhaled breath condensate (EBC) is a biofluid collected non invasively that, enabling the measurement of several biomarkers, has proven useful in the study of airway inflammatory diseases, including asthma, COPD and cystic fibrosis. To the best of our knowledge, there is no previous report of any analytical method to detect ADMA in EBC., Objectives: Aim of this work was to develop an online sample trapping and enrichment system, coupled with an UPLC-MS/MS method, for simultaneous quantification of seven metabolites related to "Arginine-ADMA cycle", using the isotopic dilution., Methods: Butylated EBC samples were trapped in an online cartridge, washed before and after each injection with cleanup solution to remove matrix components and switched inline into the high pressure analytical column. Multiple reaction monitoring in positive mode was used for analyte quantification by tandem mass spectrometry., Results: Validation studies were performed in EBC to examine accuracy, precision and robustness of the method. For each compound, the calibration curves showed a coefficient of correlation (r(2)) greater than 0.992. Accuracy (%Bias) was <3% except for NMMA and H-Arg (<20%), intra- and inter-assay precision (expressed as CV%) were within ±20% and recovery ranged from 97.1 to 102.8% for all analytes. Inter-day variability analysis on 20 EBC of adult subjects did not demonstrate any significant variation of quantitative data for each metabolite. ADMA and SDMA mean concentrations (μmolL(-1)), measured in EBC samples of asthmatic adolescents are significantly increased (p<0.0001) than in normal controls (0.0040±0.0021 vs. 0.0012±0.0005 and 0.0020±0.0015 vs. 0.0002±0.0001, respectively), as well the ADMA/Tyr (0.34±0.09 vs. 0.12±0.02, p<0.0001) and the SDMA/Tyr ratio (0.10±0.04 vs. 0.015±0.004, p<0.0001)., Conclusions: The proposed method features simple specimen preparation, maintenance of an excellent peak shape of all metabolites and reduced matrix effects as well mass spectrometer noise. Moreover, the possibility to perform different cycles of enrichment, using large injection volumes, compensated for the low concentration of analytes contained in EBC, leading to a good analytical sensitivity. Preliminary data obtained from asthmatic and healthy adolescents, demonstrated that the analytical method applied to EBC seems suitable not only for research purposes, but also for clinical routinely analysis., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Background: Asymmetric dimethylarginine (ADMA), an endogenous nitric oxide (NO) formation inhibitor, has emerged as a promising biomarker of NO-associated endothelial dysfunction in cardiovascular diseases as well in chronic renal failure. The interest in potentially fundamental role of this metabolite, in basic and clinical research, led to the development of numerous analytical methods for the quantitative determination of ADMA and dimethylarginines in biological systems, notably plasma, serum and urine., Objectives: The aim of this work was to present a simple, fast and accurate UPLC-tandem-MS-based method for the simultaneous determination and quantification of arginine, ADMA, SDMA, NMMA, homo-arginine and citrulline. This method is designed for high sample throughput of only 10 μL of human plasma, serum or urine., Methods: The analysis time is reduced to 1.9 min by an ultrahigh-performance liquid chromatography run coupled with electrospray ionization (ESI) in the positive mode tandem mass spectrometry detection., Results: The method was validated in plasma, serum and urine. Correlation coefficients (r(2)) of the calibration curves in all matrices considered ranged from 0.9810 to 0.9993. Inter- and intra-assay precision, accuracy, recovery and carry-over were evaluated for validation. The LOD was 0.01 μM for all compounds in water, plasma and serum and 0.1 μM in urine. The LOQ was 0.05 μM for ADMA, SDMA, NMMA and H-Arg and 0.5 μM for Arg and Cit in water, plasma and serum; while in urine was 0.1 μM for ADMA, SDMA, NMMA and H-Arg and 0.5 μM for Arg and Cit. The precision was ranged from 1% to 15% expressed as CV% and the accuracy (bias %) was <±7% for all added concentrations with the exception of NMMA (-10%). ADMA mean plasma levels, measured in healthy adults and newborns, were in accord with literature data published: (M±SD) 0.56±0.10 μM and 0.84±0.21 μM, respectively, showing that ADMA levels in plasma decreased with age. In serum we have similar data (0.54±0.18 μM and 1.14±0.36 μM), while in neonatal urine ADMA was 11.98±7.13 μmol mmol(-1) creatinine., Conclusions: Data from calibration curves and method validation reveal that the method is accurate and precise. The fast run time, the feasibility of high sample throughput and the small amount of sample required make this method very suitable for routine analysis in the clinical setting., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Background and aim: The pathogenesis of the pain that occurs in episodic migraine attack is due to the activation of the trigeminal system's first neuron receptors located on vessel wall. The release from the endothelium of nitric oxide, a product of arginine metabolism, causes vasodilation and stretching of the vascular trigeminal system and promotes pain. It is unknown whether this same metabolic event is involved in the pain accompanying chronic migraine. To understand the possible role of arginine in the pathogenesis of chronic migraine patients, we evaluated the metabolism of arginine in plasma of chronic migraine and control subjects. Methods: We evaluated the metabolism of arginine in a group of patients affected by chronic migraine. Quantification of arginine, ornithine, citrulline, monomethyl arginine (NMMA), dimethylarginines (ADMA, SDMA), and tyramine was performed by ultra-performance liquid chromatography coupled with a triple quadrupole mass spectrometer. Results: Chronic migraine patients showed low plasma levels of arginine, significantly elevated levels of ornithine, ADMA, and NMMA whereas the levels of citrulline and SDMA were in the range of controls. Conclusions: The elevated levels of ADMA and NMMA, inhibitors of nitric oxide synthase, suggest that the metabolism of arginine may be inhibited with a possible reduction of NO release in the circulation of chronic patients. This suggests that the origin of pain may not be related to the vasodilation of trigeminal vascular system that occurs in episodic migraine patients. [ABSTRACT FROM AUTHOR]

Abstract: Background: Asymmetric dimethylarginine (ADMA), an endogenous nitric oxide (NO) formation inhibitor, has emerged as a promising biomarker of NO-associated endothelial dysfunction in cardiovascular diseases as well in chronic renal failure. The interest in potentially fundamental role of this metabolite, in basic and clinical research, led to the development of numerous analytical methods for the quantitative determination of ADMA and dimethylarginines in biological systems, notably plasma, serum and urine. Objectives: The aim of this work was to present a simple, fast and accurate UPLC–tandem-MS-based method for the simultaneous determination and quantification of arginine, ADMA, SDMA, NMMA, homo-arginine and citrulline. This method is designed for high sample throughput of only 10μL of human plasma, serum or urine. Methods: The analysis time is reduced to 1.9min by an ultrahigh-performance liquid chromatography run coupled with electrospray ionization (ESI) in the positive mode tandem mass spectrometry detection. Results: The method was validated in plasma, serum and urine. Correlation coefficients (r 2) of the calibration curves in all matrices considered ranged from 0.9810 to 0.9993. Inter- and intra-assay precision, accuracy, recovery and carry-over were evaluated for validation. The LOD was 0.01μM for all compounds in water, plasma and serum and 0.1μM in urine. The LOQ was 0.05μM for ADMA, SDMA, NMMA and H-Arg and 0.5μM for Arg and Cit in water, plasma and serum; while in urine was 0.1μM for ADMA, SDMA, NMMA and H-Arg and 0.5μM for Arg and Cit. The precision was ranged from 1% to 15% expressed as CV% and the accuracy (bias %) was <±7% for all added concentrations with the exception of NMMA (−10%). ADMA mean plasma levels, measured in healthy adults and newborns, were in accord with literature data published: (M±SD) 0.56±0.10μM and 0.84±0.21μM, respectively, showing that ADMA levels in plasma decreased with age. In serum we have similar data (0.54±0.18μM and 1.14±0.36μM), while in neonatal urine ADMA was 11.98±7.13μmolmmol−1 creatinine. Conclusions: Data from calibration curves and method validation reveal that the method is accurate and precise. The fast run time, the feasibility of high sample throughput and the small amount of sample required make this method very suitable for routine analysis in the clinical setting. [Copyright &y& Elsevier]