Computational approach to design of aptamers to the receptor binding domain of sars-cov-2

The aim of the research. In this work, in silico selection of DNA-aptamers to the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein was performed using molecular modeling methods. Material and methods. A new computational approach to aptamer in silico selection is based on a cycle of simulations, including the stages of molecular modeling, molecular docking, molecular dynamic simulations, and quantum chemical calculations. To verify the obtained calculated results flow cytometry, fluorescence polarization, and small-angle X-ray scattering methods were applied. Results. An initial library consisted of 256 16-mer oligonucleotides was modeled. Based on molecular docking results, the only one aptamer (Apt16) was selected from the library as a starting aptamer to the RBD protein. For Apt16/RBD complex, molecular dynamic and quantum chemical calculations revealed the pairs of nucleotides and amino acids whose contribution to the binding between aptamer and RBD is the largest. Taking into account these data, Apt16 was subjected to the structure modifications in order to increase the binding with the RBD. Thus, a new aptamer Apt25 was designed. The procedure of 1) aptamer structure modeling/modification, 2) molecular docking, 3) molecular dynamic simulations, 4) quantum chemical calculations was performed sev-eral times. As a result, four aptamers (Apt16, Apt25, Apt27, Apt31) to the RBD were designed in silico without any preliminary experimental data. Binding of the each modeled aptamer to the RBD was studied in terms of interactions between residues in protein and nucleotides in the aptamers. Based on the simulation results, the strongest binding with the RBD was predicted for two Apt27 and Apt31aptamers. The calculated results are in good agreement with experimental data obtained by flow cytometry, fluorescence polarization, and small-angle X-ray scattering methods. Conclusion. The proposed computational approach to selection and refinement of aptamers is universal and can be used for wide range of molecular ligands and targets. Key words

Applicability of Capillary Gas-Liquid Chromatography for Determination of Parabens in Pharmaceutical Analysis

The traditional gas-liquid chromatography (GLC) method using packed columns is still used in pharmaceutical analysis for determination of parabens, despite the fact that this technique has a number of serious drawbacks.The aim of the study was to develop a more effective capillary GLC method for determination of parabens in active pharmaceutical ingredients and finished pharmaceutical products.Materials and methods: the study was performed using Agilent 6890N and Agilent 7890B systems with flame-ionisation detectors. The systems were equipped with Agilent 7683B and Agilent G4513A autosamplers, respectively. The following columns were used in the study: ZB-1 15 m х 0.32 mm х 0.25 pm, DB-1 30 m х 0.32 mm х 3.0 pm, Cp-Sil 5-CB 30 m х 0.32 mm х 3.0 pm.Results: the authors developed a method for methylparaben and propylparaben determination using capillary column GLC. The chromatographic parameters (chromatographic system performance, reproducibility of peak areas, peak asymmetry) were determined for both capillary and packed column GLC. The authors outlined the prospects for simultaneous determination of several compounds using the proposed method: a four-component mixture containing methyl-, ethyl-, propyl-, and butylparaben was separated in 9 minutes. The authors used Loma Lux Psoriasis to perform partial validation of the test method. They determined the linearity range and the limit of quantitation for methylparaben and propylparaben, and verified accuracy and intermediate precision of the test method.Conclusions: the results of the study allowed for selection of optimal chromatographic conditions for rapid and high-precision determination of methylparaben and propylparaben in medicinal products. The developed method is recommended for control of the content of these compounds in medicinal products

Computational approach to design of aptamers to the receptor binding domain of SARS-CoV-2

The aim of the research. In this work, in silico selection of DNA-aptamers to the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein was performed using molecular modeling methods. Material and methods. A new computational approach to aptamer in silico selection is based on a cycle of simulations, including the stages of molecular modeling, molecular docking, molecular dynamic simulations, and quantum chemical calculations. To verify the obtained calculated results flow cytometry, fluorescence polarization, and small-angle X-ray scattering methods were applied. Results. An initial library consisted of 256 16-mer oligonucleotides was modeled. Based on molecular docking results, the only one aptamer (Apt16) was selected from the library as a starting aptamer to the RBD protein. For Apt16/RBD complex, molecular dynamic and quantum chemical calculations revealed the pairs of nucleotides and amino acids whose contribution to the binding between aptamer and RBD is the largest. Taking into account these data, Apt16 was subjected to the structure modifications in order to increase the binding with the RBD. Thus, a new aptamer Apt25 was designed. The procedure of 1) aptamer structure modeling/modification, 2) molecular docking, 3) molecular dynamic simulations, 4) quantum chemical calculations was performed sev-eral times. As a result, four aptamers (Apt16, Apt25, Apt27, Apt31) to the RBD were designed in silico without any preliminary experimental data. Binding of the each modeled aptamer to the RBD was studied in terms of interactions between residues in protein and nucleotides in the aptamers. Based on the simulation results, the strongest binding with the RBD was predicted for two Apt27 and Apt31aptamers. The calculated results are in good agreement with experimental data obtained by flow cytometry, fluorescence polarization, and small-angle X-ray scattering methods. Conclusion. The proposed computational approach to selection and refinement of aptamers is universal and can be used for wide range of molecular ligands and targets.nonPeerReviewe

Возможность применения капиллярной газожидкостной хроматографии в фармацевтическом анализе при определении парабенов

The traditional gas-liquid chromatography (GLC) method using packed columns is still used in pharmaceutical analysis for determination of parabens, despite the fact that this technique has a number of serious drawbacks.The aim of the study was to develop a more effective capillary GLC method for determination of parabens in active pharmaceutical ingredients and finished pharmaceutical products.Materials and methods: the study was performed using Agilent 6890N and Agilent 7890B systems with flame-ionisation detectors. The systems were equipped with Agilent 7683B and Agilent G4513A autosamplers, respectively. The following columns were used in the study: ZB-1 15 m х 0.32 mm х 0.25 pm, DB-1 30 m х 0.32 mm х 3.0 pm, Cp-Sil 5-CB 30 m х 0.32 mm х 3.0 pm.Results: the authors developed a method for methylparaben and propylparaben determination using capillary column GLC. The chromatographic parameters (chromatographic system performance, reproducibility of peak areas, peak asymmetry) were determined for both capillary and packed column GLC. The authors outlined the prospects for simultaneous determination of several compounds using the proposed method: a four-component mixture containing methyl-, ethyl-, propyl-, and butylparaben was separated in 9 minutes. The authors used Loma Lux Psoriasis to perform partial validation of the test method. They determined the linearity range and the limit of quantitation for methylparaben and propylparaben, and verified accuracy and intermediate precision of the test method.Conclusions: the results of the study allowed for selection of optimal chromatographic conditions for rapid and high-precision determination of methylparaben and propylparaben in medicinal products. The developed method is recommended for control of the content of these compounds in medicinal products.В практике фармацевтического анализа для определения парабенов применяется имеющий ряд серьезных недостатков метод классической газожидкостной хроматографии (ГЖХ) с использованием насадочных колонок.Цель работы: разработать методику более эффективного определения парабенов в фармацевтических субстанциях и лекарственных средствах методом капиллярной ГЖХ.Материалы и методы: исследования проводились на газовых хроматографах Agilent 6890N и Agilent 7890B с пламенно-ионизационным детектором, оснащенных автосамплерами Agilent 7683B и Agilent G4513A соответственно. В работе использованы хроматографические колонки ZB-1 15 м х 0,32 мм х 0,25 мкм, DB-1 30 м х 0,32 мм х 3,0 мкм, CP-Sil 5-CB 30 м х 0,32 мм х 3,0 мкм.Результаты: разработана методика определения метилпарабена и пропилпарабена с использованием метода газожидкостной хроматографии с капиллярной колонкой. Определены параметры (эффективность хроматографической системы, воспроизводимость площадей пиков, асимметрия пиков) хроматографического определения парабенов методом капиллярной ГЖХ и методом ГЖХ с насадочной колонкой. Обозначены перспективы единовременного определения нескольких соединений с помощью предложенной методики: за 9 минут была разделена четырехкомпонентная смесь метил-, этил-, пропил- и бутилпарабена. Проведена частичная валидация методики на примере лекарственного средства «Лома Люкс Псориасис». Установлен диапазон линейности методики, предел количественного определения метилпарабена и пропилпарабена, подтверждены правильность и прецизионность (сходимость и внутрилабораторная воспроизводимость).Выводы: проведенные испытания позволили выбрать оптимальные хроматографические условия для быстрого и прецизионного определения метилпарабена и пропилпарабена в лекарственных средствах. Разработанную методику рекомендовано использовать для контроля содержания данных соединений в средствах медицинского применения