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Determination of Guanidinoacetate and Creatine in Urine and Plasma by Liquid Chromatography–Tandem Mass Spectrometry

2004; American Association for Clinical Chemistry; Volume: 50; Issue: 8 Linguagem: Inglês

10.1373/clinchem.2004.034538

1530-8561

S. Cognat, David Cheillan, Monique Piraud, Birthe Roos, Cornelis Jakobs, Christine Vianey‐Saban,

Glycogen Storage Diseases and Myoclonus

In the last 10 years, three new inborn errors of creatine metabolism and transport, called creatine deficiency syndromes (CDS), have been described (1). These are deficiencies of arginine:glycine amidinotransferase (EC 2.1.4.1), S -adenosyl-l-methionine:guanidinoacetate N -methyltransferase (GAMT; EC 2.1.1.2), and creatine transporter. Patients with CDS have mental retardation, severe speech disturbance, and depending on the disorder, epilepsy and/or extrapyramidal signs. Biochemical detection of CDS relies on the determination of two main metabolites in biological fluids: guanidinoacetate (GAA) and creatine. Few patients with CDS have been reported to date, probably because of underdiagnosis, most likely attributable to the limited availability of quantitative GAA and creatine assays. This underdiagnosis is particularly harmful because some patients improve with oral creatine supplementation. Several analytical methods for GAA and creatine have been described, including use of the Sakaguchi reaction (2), stable-isotope-dilution gas chromatography–mass spectrometry (GC/MS) (3)(4), and HPLC (5). Recently, a liquid chromatography–tandem mass spectrometry (LC/MS/MS) method has been reported for analysis of these metabolites in plasma and dried blood on filter cards (6). Here we describe the validation of LC/MS/MS for the simultaneous quantification of GAA and creatine in urine and plasma and compare the method with a stable-isotope-dilution GC/MS method. To investigate the molecular fragmentation and to prepare calibrators, we prepared 5 mmol/L stock solutions and successive dilutions of creatine, GAA (Sigma-Aldrich), N -methyl-d3-creatine (CDN Isotopes), and [13C2]GAA (Dr. H. Ten Brink, VU University Medical Center, Amsterdam, The Netherlands) in distilled water. We evaluated MS/MS conditions in the positive-ion mode by infusion of aqueous solutions into the API 2000 tandem mass spectrometer (Sciex Applied Biosystems) as described previously (7). Nitrogen was used as both the curtain and collision gas. The transitions chosen for the quantification of GAA, creatine, d3-creatine, and [13 …