Quantitative Analysis of l-Arginine, Dimethylated Arginine Derivatives, l-Citrulline, and Dimethylamine in Human Serum Using Liquid Chromatography–Mass Spectrometric Method

Abstract

Nitric oxide (NO) is a signaling molecule involved in the regulation of many biological processes, such as vascular and airway tone regulation, angiogenesis, inflammatory response and immunity, nerve signal transmission, and insulin synthesis [ 1 ]. Nitric oxide is produced during the two-step oxidation of l -arginine to l -citrulline, catalyzed by nitric oxide synthase (NOS). There are three isoforms of the enzyme, two of which are calcium dependent and expressed constitutively—neuronal and endothelial NOS (nNOS/NOS I and eNOS/NOS III, respectively)—and one inducible and calcium independent (iNOS/NOS II). Nitric oxide synthase activity is regulated by both negative feedback [ 2 , 3 ] and availability of l -arginine [ 4 ]. l -Arginine is a semi-essential amino acid obtained either from the diet or synthesized endogenously from L-citrulline. l -Citrulline, in turn, can be derived from ornithine in the catabolism of proline or glutamine and glutamate, or from l -arginine via arginine–citrulline pathway. l -Citrulline can also be obtained during the degradation of asymmetric dimethylarginine (ADMA), the process catalyzed by dimethylarginine dimethylaminohydrolase (DDAH), yielding dimethylamine (DMA) as a coproduct [ 5 ]. Asymmetric dimethylarginine is constitutively synthesized during the turnover of methylated proteins. In addition to being a potential source of l -citrulline, ADMA acts as endogenous inhibitor of several NOS isoforms, including nNOS, and regulates l -arginine availability by controlling the transport of amino acid across the cell membranes [ 6 , 7 , 8 ]. Symmetric dimethylarginine (SDMA)—ADMA regioisomer—is considered only as a weak intermediate NOS inhibitor and is competing with l -arginine for transport across cell membranes [ 9 ]. SDMA has been recognized as an effective marker of renal function [ 10 ]. There are also growing number evidences supporting participation of SDMA in the development of inflammation and atherosclerosis.