Asymmetric and symmetric dimethylarginines are toxic non-coded amino acids. They are formed by post-translational modifications and play multifunctional roles in some human diseases. Their determination in human blood plasma is performed using capillary electrophoresis with contactless conductivity detection. The separations are performed in a capillary covered with covalently bonded PAMAPTAC polymer, which generates anionic electroosmotic flow and the separation takes place in the counter-current regime. The background electrolyte is a 750 mM aqueous solution of acetic acid with pH 2.45. The plasma samples for analysis are treated by the addition of acetonitrile and injected into the capillary in a large volume, reaching 94.5% of the total volume of the capillary, and subsequently subjected to electrophoretic stacking. The attained LODs are 16 nm for ADMA and 22 nM for SDMA. The electrophoretic resolution of both isomers has a value of 5.3. The developed method is sufficiently sensitive for the determination of plasmatic levels of ADMA and SDMA. The determination does not require derivatization and the individual steps in the electrophoretic stacking are fully automated. The determined plasmatic levels for healthy individuals vary in the range 0.36-0.62 µM for ADMA and 0.32-0.70 µM for SDMA.

Department of Hygiene 3rd Faculty of Medicine Charles University Ruská 87 100 00 Prague 10 Czech Republic

Institute of Organic Chemistry and Biochemistry The Czech Academy of Sciences Flemingovo n 2 166 10 Prague 6 Czech Republic

Laboratoire des IMRCP UMR 5623 University Toulouse 3 Paul Sabatier 31062 Toulouse France

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Gary J.D., Clarke S. RNA and protein interactions modulated by protein arginine methylation. Prog. Nucleic Acid Res. Mol. Biol. 1998;61:65–131. doi: 10.1016/s0079-6603(08)60825-9. PubMed DOI

Bedford M.T., Clarke S.G. Protein Arginine Methylation in Mammals: Who, What, and Why. Mol. Cell. 2009;33:1–13. doi: 10.1016/j.molcel.2008.12.013. PubMed DOI PMC

Carlson S.M., Gozani O. Emerging Technologies to Map the Protein Methylome. J. Mol. Biol. 2014;426:3350–3362. doi: 10.1016/j.jmb.2014.04.024. PubMed DOI PMC

Ogawa T., Kimoto M., Sasaoka K. Occurrence of a new enzyme catalyzing the direct conversion of NG,NG-dimethyl-L-arginine to L-citrulline in rats. Biochem. Biophys. Res. Commun. 1987;148:671–677. doi: 10.1016/0006-291X(87)90929-6. PubMed DOI

Achan V., Broadhead M., Malaki M., Whitley G., Leiper J., MacAllister R., Vallance P. Asymmetric dimethylarginine causes hypertension and cardiac dysfunction in humans and is actively metabolized by dimethylarginine dimethylaminohydrolase. Arterioscler. Thromb. Vasc. Biol. 2003;23:1455–1459. doi: 10.1161/01.ATV.0000081742.92006.59. PubMed DOI

Vallance P., Leone A., Calver A., Collier J., Moncada S. Accumulation of an endogenous inhibitor of nitric-oxide synthesis in chronic-renal-failure. Lancet. 1992;339:572–575. PubMed

Marescau B., Nagels G., Possemiers I., DeBroe M.E., Becaus I., Billiouw J.M., Lornoy W., DeDeyn P.P. Guanidino compounds in serum and urine of nondialyzed patients with chronic renal insufficiency. Metab. Clin. Exp. 1997;46:1024–1031. doi: 10.1016/S0026-0495(97)90273-0. PubMed DOI

Blackwell S. The biochemistry, measurement and current clinical significance of asymmetric dimethylarginine. Ann. Clin. Biochem. 2010;47:17–28. doi: 10.1258/acb.2009.009196. PubMed DOI

Siroen M.P.C., Teerlink T., Nijveldt R.J., Prins H.A., Richir M.C., van Leeuwen P.A.M. The clinical significance of asymmetric dimethylarginine. Annu. Rev. Nutr. 2006;26:203–228. doi: 10.1146/annurev.nutr.26.061505.111320. PubMed DOI

Tain Y.L., Hsu C.N. Interplay between Oxidative Stress and Nutrient Sensing Signaling in the Developmental Origins of Cardiovascular Disease. Int. J. Mol. Sci. 2017;18:841. doi: 10.3390/ijms18040841. PubMed DOI PMC

Cooke J.P. The pivotal role of nitric oxide for vascular health. Can. J. Cardiol. 2004;20:7B–15B. PubMed

Steyers C.M., Miller F.J. Endothelial Dysfunction in Chronic Inflammatory Diseases. Int. J. Mol. Sci. 2014;15:11324–11349. doi: 10.3390/ijms150711324. PubMed DOI PMC

Grosse G.M., Schwedhelm E., Worthmann H., Choe C.U. Arginine Derivatives in Cerebrovascular Diseases: Mechanisms and Clinical Implications. Int. J. Mol. Sci. 2020;21:1798. doi: 10.3390/ijms21051798. PubMed DOI PMC

Boger R.H., Bode-Boger S.M., Szuba A., Tsao P.S., Chan J.R., Tangphao O., Blaschke T.F., Cooke J.P. Asymmetric dimethylarginine (ADMA): A novel risk factor for endothelial dysfunction—Its role in hypercholesterolemia. Circulation. 1998;98:1842–1847. doi: 10.1161/01.CIR.98.18.1842. PubMed DOI

Sydow K., Mondon C.E., Cooke J.P. Insulin resistance: Potential role of the endogenous nitric oxide synthase inhibitor ADMA. Vasc. Med. 2005;10:S35–S43. doi: 10.1177/1358836X0501000106. PubMed DOI

Jacobi J., Tsao P.S. Asymmetrical dimethylarginine in renal disease: Limits of variation or variation limits? Am. J. Nephrol. 2008;28:224–237. doi: 10.1159/000110092. PubMed DOI PMC

Rysz J., Gluba-Brzozka A., Franczyk B., Jablonowski Z., Cialkowska-Rysz A. Novel Biomarkers in the Diagnosis of Chronic Kidney Disease and the Prediction of Its Outcome. Int. J. Mol. Sci. 2017;18:1702. doi: 10.3390/ijms18081702. PubMed DOI PMC

Surdacki A., Nowicki M., Sandmann J., Tsikas D., Boeger R.H., Bode-Boeger S.M., Kruszelnicka-Kwiatkowska O., Kokot F., Dubiel J.S., Froelich J.C. Reduced urinary excretion of nitric oxide metabolites and increased plasma levels of asymmetric dimethylarginine in men with essential hypertension. J. Cardiovasc. Pharmacol. 1999;33:652–658. doi: 10.1097/00005344-199904000-00020. PubMed DOI

Tain Y.L., Hsu C.N. Targeting on Asymmetric Dimethylarginine-Related Nitric Oxide-Reactive Oxygen Species Imbalance to Reprogram the Development of Hypertension. Int. J. Mol. Sci. 2016;17:2020. doi: 10.3390/ijms17122020. PubMed DOI PMC

Cardounel A.J., Cui H.M., Samouilov A., Johnson W., Kearns P., Tsai A.L., Berka V., Zweier J.L. Evidence for the pathophysiological role of endogenous methylarginines in regulation of endothelial NO production and vascular function. J. Biol. Chem. 2007;282:879–887. doi: 10.1074/jbc.M603606200. PubMed DOI

Bode-Boger S.M., Scalera F., Kielstein J.T., Martens-Lobenhoffer J., Breithardt G., Fobker M., Reinecke H. Symmetrical dimethylarginine: A new combined parameter for renal function and extent of coronary artery disease. J. Am. Soc. Nephrol. 2006;17:1128–1134. doi: 10.1681/ASN.2005101119. PubMed DOI

Zhang W.Z., Kaye D.M. Simultaneous determination of arginine and seven metabolites in plasma by reversed-phase liquid chromatography with a time-controlled ortho-phthaldialdehyde precolumn derivatization. Anal. Biochem. 2004;326:87–92. doi: 10.1016/j.ab.2003.11.006. PubMed DOI

Marra M., Bonfigli A.R., Testa R., Testa I., Gambini A., Coppa G. High-performance liquid chromatographic assay of asymmetric dimethylarginine, symmetric dimethylarginine, and arginine in human plasma by derivatization with naphthalene-2,3-dicarboxaldehyde. Anal. Biochem. 2003;318:13–17. doi: 10.1016/S0003-2697(03)00157-X. PubMed DOI

Heresztyn T., Worthley M.I., Horowitz J.D. Determination of L-arginine and N-G,N-G- and N-G,N-G ‘-dimethyl-L-arginine in plasma by liquid chromatography as AccQ-Fluor (TM) fluorescent derivatives. J. Chromatogr. B. 2004;805:325–329. doi: 10.1016/j.jchromb.2004.03.020. PubMed DOI

Nonaka S., Tsunoda M., Imai K., Funatsu T. High-performance liquid chromatographic assay of N-G-monomethyl-L-arginine, N-G-dimethyl-L-arginine, and (NNG)-N-G-dimethyl-L-arginine using 4-fluoro-7-nitro-2,1,3-benzoxaldiazole as a fluorescent reagent. J. Chromatogr. A. 2005;1066:41–45. doi: 10.1016/j.chroma.2005.01.052. PubMed DOI

Teerlink T., Nijveldt R.J., de Jong S., van Leeuwen P.A.M. Determination of arginine, asymmetric dimethylarginine, and symmetric dimethylarginine in human plasma and other biological samples by high-performance liquid chromatography. Anal. Biochem. 2002;303:131–137. doi: 10.1006/abio.2001.5575. PubMed DOI

Shin S., Fung S.M., Mohan S., Fung H.L. Simultaneous bioanalysis of L-arginine, L-citrulline, and dimethylarginines by LC-MS/MS. J. Chromatogr. B. 2011;879:467–474. doi: 10.1016/j.jchromb.2011.01.006. PubMed DOI PMC

Schwedhelm E., Maas R., Tan-Andresen J., Schulze F., Riederer U., Boger R.H. High-throughput liquid chromatographic-tandem mass spectrometric determination of arginine and dimethylated arginine derivatives in human and mouse plasma. J. Chromatogr. B. 2007;851:211–219. doi: 10.1016/j.jchromb.2006.11.052. PubMed DOI

Sotgia S., Zinellu A., Paliogiannis P., Pinna G.A., Mangoni A.A., Milanesi L., Carru C. A diethylpyrocarbonate-based derivatization method for the LC-MS/MS measurement of plasma arginine and its chemically related metabolites and analogs. Clin. Chim. Acta. 2019;492:29–36. doi: 10.1016/j.cca.2019.02.004. PubMed DOI

Teerlink T. HPLC analysis of ADMA and other methylated L-arginine analogs in biological fluids. J. Chromatogr. B. 2007;851:21–29. doi: 10.1016/j.jchromb.2006.07.024. PubMed DOI

Martens-Lobenhoffer J., Bode-Boger S.M. Mass spectrometric quantification of L-arginine and its pathway related substances in biofluids: The road to maturity. J. Chromatogr. B. 2014;964:89–102. doi: 10.1016/j.jchromb.2013.10.030. PubMed DOI

Tsikas D. A critical review and discussion of analytical methods in the L-arginine/nitric oxide area of basic and clinical research. Anal. Biochem. 2008;379:139–163. doi: 10.1016/j.ab.2008.04.018. PubMed DOI

Martens-Lobenhoffer J., Bode-Boger S.A. Chromatographic-mass spectrometric methods for the quantification of L-arginine and its methylated metabolites in biological fluids. J. Chromatogr. B. 2007;851:30–41. doi: 10.1016/j.jchromb.2006.07.038. PubMed DOI

Schwedhelm E. Quantification of ADMA: Analytical approaches. Vasc. Med. 2005;10:S89–S95. doi: 10.1177/1358836X0501000113. PubMed DOI

Albsmeier J., Schwedhelm E., Schulze F., Kastner M., Boger R.H. Determination of N-G,N-G-dimethyl-L-arginine, an endogenous NO synthase inhibitor, by gas chromatography-mass spectrometry. J. Chromatogr. B. 2004;809:59–65. doi: 10.1016/j.jchromb.2004.06.008. PubMed DOI

Schulze F., Wesemann R., Schwedhelm E., Sydow K., Albsmeier J., Cooke J.P., Boger R.H. Determination of asymmetric dimethylarginine (ADMA) using a novel ELISA assay. Clin. Chem. Lab. Med. 2004;42:1377–1383. doi: 10.1515/CCLM.2004.257. PubMed DOI

Causse E., Siri N., Arnal J.F., Bayle C., Malatray P., Valdiguie P., Salvayre R., Couderc F. Determination of asymmetrical dimethylarginine by capillary electrophoresis-laser-induced fluorescence. J. Chromatogr. B. 2000;741:77–83. doi: 10.1016/S0378-4347(00)00034-7. PubMed DOI

Zinellu A., Sotgia S., Zinellu E., Pinna A., Carta F., Gaspa L., Deiana L., Carru C. High-throughput CZE-UV determination of airginine and dimethylated arginines in human plasma. Electrophoresis. 2007;28:1942–1948. doi: 10.1002/elps.200600534. PubMed DOI

Zinellu A., Sotgia S., Porcu P., Casu M.A., Bivona G., Chessa R., Deiana L., Carru C. Carotid restenosis is associated with plasma ADMA concentrations in carotid endarterectomy patients. Clin. Chem. Lab. Med. 2011;49:897–901. doi: 10.1515/CCLM.2011.121. PubMed DOI

Zinellu A., Sotgia S., Scanu B., Deiana L., Carru C. Determination of protein-incorporated methylated arginine reference values in healthy subjects whole blood and evaluation of factors affecting protein methylation. Clin. Biochem. 2008;41:1218–1223. doi: 10.1016/j.clinbiochem.2008.07.011. PubMed DOI

Zinellu A., Sotgia S., Usai M.F., Pintus G., Deiana L., Carru C. Improved method for plasma ADMA, SDMA, and arginine quantification by field-amplified sample injection capillary electrophoresis UV detection. Anal. Bioanal. Chem. 2011;399:1815–1821. doi: 10.1007/s00216-010-4580-0. PubMed DOI

Berlinguer F., Porcu C., Molle G., Cabiddu A., Dattena M., Gallus M., Pasciu V., Succu S., Sotgiu F.D., Paliogiannis P., et al. Circulating Concentrations of Key Regulators of Nitric Oxide Production in Undernourished Sheep Carrying Single and Multiple Fetuses. Animals. 2020;10:65. doi: 10.3390/ani10010065. PubMed DOI PMC

Desiderio C., Rossetti D.V., Messana I., Giardina B., Castagnola M. Analysis of arginine and methylated metabolites in human plasma by field amplified sample injection capillary electrophoresis tandem mass spectrometry. Electrophoresis. 2010;31:1894–1902. doi: 10.1002/elps.200900690. PubMed DOI

Linz T.H., Snyder C.M., Lunte S.M. Optimization of the Separation of NDA-Derivatized Methylarginines by Capillary and Microchip Electrophoresis. JALA. 2012;17:24–31. doi: 10.1177/2211068211424551. PubMed DOI PMC

Linz T.H., Lunte S.M. Heat-assisted extraction for the determination of methylarginines in serum by CE. Electrophoresis. 2013;34:1693–1700. doi: 10.1002/elps.201200567. PubMed DOI PMC

Hauser P.C., Kubáň P. Capacitively coupled contactless conductivity detection for analytical techniques—Developments from 2018 to 2020. J. Chromatogr. A. 2020;1632:19. doi: 10.1016/j.chroma.2020.461616. PubMed DOI

Tůma P. Determination of amino acids by capillary and microchip electrophoresis with contactless conductivity detection—Theory, instrumentation and applications. Talanta. 2021;224 doi: 10.1016/j.talanta.2020.121922. PubMed DOI

Tůma P., Samcová E., Štulík K. Contactless conductivity detection in capillary electrophoresis employing capillaries with very low inner diameters. Electroanalysis. 2011;23:1870–1874. doi: 10.1002/elan.201100264. DOI

Tůma P., Hložek T., Sommerová B., Koval D. Large volume sample stacking of antiepileptic drugs in counter current electrophoresis performed in PAMAPTAC coated capillary. Talanta. 2021;221:8. doi: 10.1016/j.talanta.2020.121626. PubMed DOI

Tůma P., Koval D., Sommerová B., Vaculín S. Separation of anaesthetic ketamine and its derivates in PAMAPTAC coated capillaries with tuneable counter-current electroosmotic flow. Talanta. 2020;217:8. doi: 10.1016/j.talanta.2020.121094. PubMed DOI

Tůma P., Sommerová B., Vaculín Š. Rapid electrophoretic monitoring of the anaesthetic ketamine and its metabolite norketamine in rat blood using a contactless conductivity detector to study the pharmacokinetics. J. Sep. Sci. 2019;42:2062–2068. doi: 10.1002/jssc.201900116. PubMed DOI

Tůma P. Frequency-tuned contactless conductivity detector for the electrophoretic separation of clinical samples in capillaries with very small internal dimensions. J. Sep. Sci. 2017;40:940–947. doi: 10.1002/jssc.201601213. PubMed DOI

El-Debs R., Marechal A., Dugas V., Demesmay C. Photopolymerization of acrylamide as a new functionalization way of silica monoliths for hydrophilic interaction chromatography and coated silica capillaries for capillary electrophoresis. J. Chromatogr. A. 2014;1326:89–95. doi: 10.1016/j.chroma.2013.12.038. PubMed DOI

El-Debs R., Dugas V., Demesmay C. Photografting as a versatile, localizable, and single-step surface functionalization of silica-based monoliths dedicated to microscale separation techniques. J. Sep. Sci. 2013;36:993–1001. doi: 10.1002/jssc.201200878. PubMed DOI

Shihabi Z.K. Transient pseudo-isotachophoresis for sample concentration in capillary electrophoresis. Electrophoresis. 2002;23:1612–1617. doi: 10.1002/1522-2683(200206)23:11<1612::AID-ELPS1612>3.0.CO;2-3. PubMed DOI

Shihabi Z.K. Stacking of weakly cationic compounds by acetonitrile for capillary electrophoresis. J. Chromatogr. A. 1998;817:25–30. doi: 10.1016/S0021-9673(98)00312-4. DOI

Shihabi Z.K. Stacking in capillary zone electrophoresis. J. Chromatogr. A. 2000;902:107–117. doi: 10.1016/S0021-9673(00)00743-3. PubMed DOI

Křivánková L., Pantůčková P., Boček P. Isotachophoresis in zone electrophoresis. J. Chromatogr. A. 1999;838:55–70. doi: 10.1016/S0021-9673(99)00169-7. PubMed DOI