LIQUID CHROMATOGRAPHY-TANDEM MASS SPECTROMETRY (HPLS-MS-MS) AS METHOD FOR SEPARATION AND IDENTIFICATION OF NEW ANTI-INFLAMMATORY AGENT FROM DERIVATIVES OF THIADIAZOLILAMIDE

DOI: https://doi.org/10.29296/25877313-2018-04-03
Issue: 
4
Year: 
2018

M.A. Demidova Dr.Sc. (Med.), Professor, Head of Department, Tver State Medical University N.S. Popov Assistant, Department of Pharmacy, Tver State Medical University E-mail: ns.popov@mail.ru A.S. Malygin Ordinator, Department of Pharmacology and Clinical Pharmacology, Tver State Medical University E-mail: education@tvgmu.ru

Objective. Development of the HPLC-MS / MS technique for the separation and identification of new anti-inflammatory drugs from the thiadiazolamide group. Material and methods. Non-steroid anti-inflammatory drugs from the group of thiadiazolylamide derivatives – acexazolamide, pyridazolamide and nitrobenzolamide were the objects of this study. Chromatography was performed using an Agilent 1260 Infinity II chromatograph (Agilent InfinityLab Poroshell 120 analytical columns EC-C18 2.7 μm 4.6 × 100 mm, Agilent Zorbax Eclipse Plus C18 5 μm 4.6 × 150 mm). As a mobile phase, a mixtures of acetonitrile, methanol, deionized water and formic acid was used in different ratios in isocratic or gradient modes. Identification of new anti-inflammatory agents from the thiadiazolamide group was carried out mass-spectrometrically using a triple quadrupole mass spectrometer AB Sciex QTrap 3200 MD with an electrospray ion source (Turbo V with a TurboIonSpray probe). Results. a high performance liquid chromatography technique with tandem mass spectrometry (HPLC-MS / MS) was developed to separate and identify new anti-inflammatory agents from the thiadiazolamide group (Zorbax Eclipse Plus C18 analytical column 5 μm 4.6 × 150 mm, gradient elution: 1-2 , 5 min 30% acetonitrile, 0.6 ml/min 2.5 - 12 min 70% acetonitrile, 1.2 ml/min, analysis time 3 minutes, mass spectrometric detection). The retention time of acexazolamide, pyridazolamide and nitrobenzolamide was 2.05, 2.22 and 2.70 minutes respectively. The identification of thiadiazolylamides was carried out by mass spectrometry. Detection conditions: negative polarization, electrospray voltage - 5500.0 V, decasterization potential of acexazolamide, pyridazolamide and nitrobenzenamide -170,0; -140.5, -280.0 V at a curtain gas pressure of 20.0 psi and a spray gas of 40.0 psi, a rate of 10 μl/min. The input potential for all ions was -4.5 V, the scan range was 200-300 Da. The values of MRM transitions with negative polarization were for azexazolamide m/z 282.8 → 73.1 m/z 282.8 → 128 and m /z 282.8 → 196.1; for nitrobenzenamide m/z 276.9 → 189.9, m/z 276.9 → 144.2 and m/z 276.9 → 122.1; for pyridazolamide m/z 232.9 → 146.2, m/z 232.9 → 78.0 and m/z 232.9 → 102.9. Conclusion. The developed HPLC-MS / MS technique allows the detection of new non-steroid anti-inflammatory drugs from the group of thiadiazolamide derivatives in one sample without their separation

Keywords: 
HPLS-MS/MS
chromatography
mass-spectrometry
thiadiazolilamides
non-steroid anti-inflammatory drug

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