I.S. Mamedov - Ph.D. (Med.), Director, Center for Implementation of Innovative Medical and Pharmaceutical Technologies, Pirogov Russian National Research Medical University (RNRMU), E-mail: is_mamedov@mail.ru
P.B. Glagovsky - Physician of Clinical Laboratory Diagnostics, Pirogov Russian National Research Medical University (RNRMU). E-mail: glagovsky@gmail.com
I.V. Zolkina - Senior Researcher, Research Laboratory of General Pathology, Moscow Research Institute of Pediatrics and Child Surgery. E-mail: zolkina_ira@mail.ru
V.S. Sukhorukov - Dr. Sc. (Med.), Professor, Head of the Research Laboratory of General Pathology, Moscow Research Institute of Pediatrics and Child Surgery. E-mail: vsukhorukov@pedklin.ru
V.I. Vasin - Senior Laboratory Assistant, General Surgery Department № 1, Medical faculty, Pirogov Russian National Research Medical University. E-mail: vitalikv25@mail.ru

Methods for determining levels of catecholamines and serotonin metabolites in the urine should be very sensitive and accurate, allowing to identify and trace the dynamics of changes in these values. These requirements are met, such methods of determining joint as HPLC and GC-MS. For these purposes, ELISA sufficiently informative. Objective: to study the possibility of applying the new algorithm to diagnose various diseases by determining serotonin and catecholamine metabolites in urine using a new high-tech methods, and evaluate the effectiveness of the joint determination of these compounds. In the course of the study samples for analysis served daily urine of patients diagnosed with the disease. After collection of daily urine volume in the total of the plastic container of about 100 ml were selected biomaterial indicating diuresis. As the preservative used 6N hydrochloric acid. After acidification, the shelf life of the samples in a refrigerator at 2 - 8 °C does not exceed 5 days, so prolonged storage, samples were frozen at - 20 °C. Selected biomaterial subjected to quantitative analysis by gas chromatography and HPLC. Gas chromatography combined with mass spectrometric detection (GC-MS, GC-MS). The process of sample preparation comprised the selective solvent extraction end metabolite catecholamines and serotonin, and concentration and derivatization at 80 °C. As an internal standard used methyl testosterone. Determination of metabolites of epinephrine, norepinephrine, serotonin and dopamine - homovanillic acid (HVA), vanililmandelic acid (VMA) and 5-hydroxyindole acetic acid (5-HIAA) was carried out by gas chromatography with mass spectrometric detection company Shimadzu GCMS QP-5050A, equipped with autoinjector AOC-20i ( Russian State Medical University, Department of Clinical Laboratory Diagnostics). For each sample chromatograms were obtained on which by computer data processing system CLASS-VPTM Chromatography Data System for gas chromatography-mass spectrometer determined concentration of the metabolite in mg/day. All data were subjected to statistical processing. In the analysis, as the standards used homovanillic, vanililmandelic acid, 5-hydroxyindoleacetic acid. Served as an internal standard iso- vanililmandelic acid. All compounds were purchased from Sigma-Aldrich . Stock solutions of test compounds at the concentrations of 100 mg/l was prepared by diluting 10-3M hydrochloric acid. Working solutions were prepared from 10-fold diluted standard solutions. Analysis of samples was performed by liquid chromatography tandem mass spectrometer Agilent Technologies 6410 Triple Quad LC / MS with autosampler , consisting of dual gradient pump Agilent 1200 series, vacuum degasser, an analytical column Zorbax Eclipse XDB8-C18 (4.0×150 mm, particle size of 5 um) , the temperature controller equipped. The data obtained were analyzed using Mass Hunter software for the analysis and identification of the chromatographic peaks. Ultimately, the concentration of a metabolite calculated in urine as mg/day. Figure 2 shows a chromatogram of the urine obtained by HPLC. In the study described procedures performed chromatographic analysis of metabolites of catecholamines and serotonin in approximately 100 children suspected of various diseases. Based on the urine of the patients were determined reference values of catecholamines, serotonin and their main metabolites (see Table 1). In three cases, the diagnosis was confirmed neuroblastoma in children, and HVA and VMA levels in the urine which were above the age normal range on average 3 times. In 5 children had confirmed the diagnosis of pheochromocytoma, and in three cases, the disease was diagnosed in the early stages in the subclinical phase. The obtained results allow us to choose the method of gas chromatography with mass spectrometric detection, as the most sensitive method. The developed technique reduces analysis time and allows you to analyze up to 100 samples per day. Thus, it is possible to successfully use this method for screening assays for the presence of neuroblastoma, pheochromocytoma, carcinoid syndrome and other disorders, while considerably facilitating the search of modern diagnostic methods for medical practitioners.