phkineticaФармакокинетика и ФармакодинамикаPharmacokinetics and Pharmacodynamics2587-78362686-8830ООО «Издательство ОКИ»10.37489/2587-7836-2023-3-28-38phkinetica-379Research ArticleДОКЛИНИЧЕСКИЕ ИССЛЕДОВАНИЯ ФАРМАКОКИНЕТИКИPRECLINICAL PHARMACOKINETIC STUDIESКоличественное определение циклического гуанозинмонофосфата (ц-ГМФ) в тканях крыс с помощью жидкостной хроматографии и тандемной масс-спектрометрииQuantitative determination of cyclic guanosine monoposphate (c-GMP) in rat tissues using liquid chromatography and tandem mass spectrometryhttps://orcid.org/0000-0002-1792-7414ПоповН. С.PopovN. S.

Попов Никита Сергеевич - к. фарм. н., зав. научно-исследовательской лабораторией, доцент кафедры фармакологии и клинической фармакологии

Тверь

Nikita S. Popov - PhD, Cand. Sci. Pharm., Head of Research Laboratory, Associate Professor, Department of Pharmacology and Clinical Pharmacology

Tver

ns.popov@mail.ruhttps://orcid.org/0000-0002-5744-7060БалабаньянВ. Ю.BalabanyanV. Yu.

Балабаньян Вадим Юрьевич -д. фарм. н., доцент, в. н. с., лаборатория трансляционной медицины, факультет фундаментальной медицины

Москва

Vadim Yu. Balabanyan - Dr. Sci. (Pharm), Associate Professor, Leading Researcher, Laboratory of Translational Medicine, Faculty of Fundamental Medicine

Moscow

bal.pharm@mail.ruКолгинаН. Ю.KolginaN. Yu.

Колгина Наталья Юрьевна - к. м. н., заведующая кафедрой фармакологии и клинической фармакологии

Тверь

Natalya Yu. Kolgina - PhD, Cand. Sci. Med., Head of Department of Pharmacology and Clinical Pharmacology

Tver

natali_jbk@mail.ruПетровГ. А.PetrovG. A.

Петров Геннадий Анатольевич - к. м. н., доцент кафедры фармакологии и клинической фармакологии

Тверь

Gennady A. Petrov - PhD, Cand. Sci. Med., Associate Professor, Department of Pharmacology and Clinical Pharmacology

Tver

petrov_gennadiy@mail.ruhttps://orcid.org/0000-0003-3780-5534ДонсковС. А.DonskovS. A.

Донсков Сергей Александрович - к. с.-х. н., доцент кафедры гистологии, эмбриологии и цитологии

Тверь

Sergey A. Donskov - PhD, Cand. Sci. Agricult., Associate Professor, Department of Histology, Embryology and Cytology

Tver

donskov_s@mail.ruhttps://orcid.org/0000-0003-1995-2661АтаджановИ. Б.AtadzhanovI. B.

Атаджанов Ильяс Борисович - препаратор кафедры гистологии, эмбриологии и цитологии

Тверь

Ilyas B. Atadzhanov - preparator of the Department of Histology, Embryology and Cytology

Tver

atadzanov.ilyas@yandex.ruГБОУ ВО «Тверской государственный медицинский университет» Министерства здравоохранения Российской Федерации (ФГБОУ ВО Тверской ГМУ Минздрава России)РоссияFSBEI HE Tver SMU MOH RussiaRussian FederationФГБОУ ВО «Московский государственный университет им. М.В. Ломоносова»РоссияLomonosov Moscow State University,Russian Federation202303112023032838Copyright © Попов Н.С., Балабаньян В.Ю., Колгина Н.Ю., Петров Г.А., Донсков С.А., Атаджанов И.Б., 20232023Попов Н.С., Балабаньян В.Ю., Колгина Н.Ю., Петров Г.А., Донсков С.А., Атаджанов И.Б.Popov N.S., Balabanyan V.Y., Kolgina N.Y., Petrov G.A., Donskov S.A., Atadzhanov I.B.Данная работа распространяется под лицензией Creative Commons Attribution 4.0.This work is licensed under a Creative Commons Attribution 4.0 License.https://www.pharmacokinetica.ru/jour/article/view/379

Циклический 3′,5′-гуанозинмонофосфат является вторичным внутриклеточным мессенджером, который играет ключевую роль во многих физиологических процессах. Количественное определение уровня ц-ГМФ в тканях лабораторных животных является актуальной задачей экспериментальной фармакологии и физиологии.

Цель — разработка методики количественного определения циклического гуанозинмонофосфата в различных тканях крыс с применением высокоэффективной жидкостной хроматографии с масс-спектрометрическим детектированием.

Методы

Методы. Выделение ц-ГМФ осуществляли путём гомогенизации биоматериала с деионизированной водой. Экстракцию ц-ГМФ из гомогенатов проводили с помощью метанола, в качестве внутреннего стандарта использовали ацикловир. Детектирование ц-ГМФ и ацикловира осуществляли с помощью масс-спектрометра Sciex QTrap 3200MD, хроматографическое разделение проводили с использованием ВЭЖХ Agilent Technologies 1260 Infinity II. В качестве элюента использовали метанол и деионизированную воду.

Результаты

Результаты. Детекцию ц-ГМФ осуществляли на основании MRM переходов m/z 346,2/152,1; 346,2/135,1, хроматографическое определение ц-ГМФ проводили в обращённо-фазовом режиме на колонке Agilent InfinityLab Poroshell 120 EC-C18 4,6×100 мм, 2,7 мкм. Время удерживания ц-ГМФ и ацикловира составило 7,85 и 7,45 минут, соответственно, при общей продолжительности хроматографического анализа 12 минут. Аналитический диапазон методики определения ц-ГМФ в гомогенатах составил 0,5–1000,0 пмоль/мл. Для апробации методики был проведён анализ содержания ц-ГМФ в тканях интактных крыс Wistar.

Заключение

Заключение. Разработанная биоаналитическая ВЭЖХ-МС/МС-методика количественного определения ц-ГМФ полностью соответствует валидационным требованиям. Метрологические характеристики методики позволяют с высокой точностью оценить содержание ц-ГМФ в различных тканях крыс.

Relevance

Relevance. Cyclic 3′,5′-guanosine monophosphate is a secondary intracellular messenger that plays a key role in many physiological processes.

Quantitative determination of the level of c-GMP in the tissues of laboratory animals is an urgent task of experimental pharmacology and physiology.

Purpose of the study

Purpose of the study. Development of a method for the quantitative determination of cyclic guanosine monophosphate in various tissues of rats using high performance liquid chromatography with mass spectrometric detection.

Methods

Methods. The biomaterial was homogenized with deionized water. Extraction of c-GMP from homogenates was performed with methanol, acyclovir was used as an internal standard. Detection of c-GMP and acyclovir was performed using a Sciex QTrap 3200MD mass spectrometer, chromatographic separation was performed using an Agilent Technologies 1260 Infinity II HPLC. The mobile phase was methanol and deionized water.

Results

Results. Detection of c-GMP was performed by MRM transitions m/z 346.2/152.1; 346.2/135.1, chromatographic determination of c-GMP was performed in reverse phase mode on an Agilent InfinityLab Poroshell 120 EC-C18 4.6×100 mm, 2.7 µm column. The retention time of c-GMP and acyclovir was 7.85 and 7.45 minutes, respectively, the total duration of the chromatographic analysis was 12 minutes. The analytical range of the procedure for determining c-GMP in homogenates was 0.5–1000.0 pmol/ml. The content of c-GMP in the tissues of intact Wistar rats was analyzed using the developed method.

Conclusion

Conclusion. The developed bioanalytical HPLC-MS/MS method for the quantitative determination of c-GMP fully complies with the validation requirements. The metrological characteristics of the method make it possible to estimate the content of c-GMP in various tissues of rats with high accuracy.

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The authors declare that there are no conflicts of interest present.