Vol. 19 No. 1(73) (2021)

Published: 2021-03-29

Original Researches

  • The preparative synthetic approach to 4-(trifluoromethoxy)piperidine and 4-(trifluoromethoxymethyl)piperidine

    Ivan G. Logvinenko, Violetta G. Dolovanyuk, Ivan S. Kondratov

    Aim. To develop a convenient synthetic approach for the preparation of multigram amounts of 4-(trifluoromethoxy)-piperidine and 4-(trifluoromethoxymethyl)piperidine – promising building blocks for medicinal chemistry.
    Results and discussion. 4-(Trifluoromethoxy)piperidine (8.4 g) and 4-(trifluoromethoxymethyl)piperidine (12.9 g) were synthesized in 5 stages starting from 4-hydroxypiperidine (the overall yield 40 %) and 4-(hydroxymethyl)piperidine (the overall yield 13.5 %), respectively.
    Experimental part. The first stage of the synthetic strategy was acylation of 4-hydroxypiperidine with benzoyl chloride. N-benzoyl-4-hydroxypiperidine obtained was transformed to N-benzoyl-4-(trifluoromethoxy)piperidine in two stages using the Hiyama method (the synthesis of the corresponding S-methyl xanthate with the subsequent desulfurization/fluorination using N-bromosuccinimide and Olah’s reagent). Then the N-benzoyl group was reduced to benzyl one, which was removed using 1-chloroethyl chloroformate. The similar approach was applied to the synthesis of 4-(trifluoromethoxymethyl)piperidine starting from 4-(hydroxymethyl)piperidine. The structure and composition of the compounds synthesized were confrmed by 1Н, 13C and 19F NMR spectroscopy,
    mass-spectrometry and elemental analysis.
    Conclusions. The synthetic approach developed is a convenient method for the multigram preparation of
    4-(trifluoromethoxy)piperidine and 4-(trifluoromethoxymethyl)piperidine and can be used for the synthesis of other secondary amines containing the CF3O-group.
    Key words: fluorination; trifluoromethoxy group; xanthate; piperidine; protection group

    DOI: https://doi.org/10.24959/ophcj.21.222669
  • Convenient approaches to the synthesis of 6-amino- and 6-oxoimidazo[4,5-b]pyrazolo[3,4-e]pyridines

    G. G. Yakovenko, M. V. Vovk

    Aim. To develop convenient approaches to the synthesis of 6-amino- and 6-oxoimidazo[4,5-b]pyrazolo[3,4-e]pyridines as promising biologically active scaffolds.
    Results and discussion. It has been found that cyclocondensation of N-Boc-4-aminopyrazole-5-carbaldehydes with creatinine can be used as an effective method for obtaining 6-aminoimidazo[4,5-b]pyrazolo[3,4-e]pyridines previously unknown. For the synthesis of their 6-oxoanalogs, the reaction of 5-aminopyrazolo[4,3-b]pyridine-6-carboxylic acids used in a modifed Curtius rearrangement with diphenylphosphorylazide was successful. This method was implemented through the stage of the intermediate aminoisocyanates formation.
    Experimental part. The reaction of N-Boc-4-aminopyrazole-5-carbaldehydes with creatinine in the presence of pyrrolidine as a catalyst in refluxing acetic acid allowed to obtain 6-aminoimidazo[4,5-b]pirazolo[3,4-e]pyridines with the yields of 54 – 70 %. The structure of the compounds synthesized was proven by spectral measurements. In the 1H NMR spectra there were singlets of H-3 (7.63 – 7.88 ppm) and H-8 (7.87 – 8.26 ppm) protons, as well as broad singlets of the NH2 group in the range of 7.05 – 7.21 ppm. Heating of 5-aminopyrazolo[4,3-b]pyridine-6-carboxylic acids with triethylamine and diphenylphosphorylazide in dioxane for 6 hours gave 1-substituted imidazo[4,5-b]pyrazolo[3,4-е]pyridine-6(5Н)-ones with the yields of 67 – 80 %. The IR-spectra of the compounds synthesized were characterized by the absorption bands of the C=O (1705 – 1708 cm-1) and NH (3275 – 3281 cm-1) groups. 1H NMR-spectra were characterized by singlets of H-3 and H-8 protons in the intervals of 7.43 – 8.08 ppm and 7.92 – 8.32 ppm respectively, as well as by two broad singlets of NH-protons in the ranges of 10.90 – 11.12 ppm and 11.25 – 11.37 ppm.
    Conclusions. Effective approaches to the synthesis of new promising heterocyclic systems of 6-amino- and
    6-oxoimidazo[4,5-b]pirazolo[3,4-e]pyridines have been developed. Cyclocondensations of N-Boc-4-aminopyrazole-5-carbaldehydes with creatinine and 5-aminopyrazolo[4,3-b]pyridine-6-carboxylic acids with diphenylphosphorylazide have been proven to be convenient ways to obtain these compounds with good yields.
    Key words: N-Boc-4-aminopyrazole-5-carbaldehyde; creatinine; 5-aminopyrazolo[4,3-b]pyridine-
    6-carboxylic acid; diphenylphosphorylazide; 6-amino(oxo)imidazo[4,5-b]pyrazolo[3,4-e]pyridines;

    DOI: https://doi.org/10.24959/ophcj.21.224583
  • The determination of the phytochemical composition of the Altabor substance

    S. Yu. Sheiko, A. S. Shalamay

    The development of medicines based on alder cone extracts led to the introduction of Altan and Altabor
    medicines into medical practice. The technology of extraction cake from cones has made it possible to obtain extracts with different therapeutic properties.
    Aim. To develop an effective method for studying the qualitative composition of the Altabor substance and
    determine the quantitative content of its components.
    Results and discussion. The Altabor substance is a complex mixture of ellagitannins containing more than 70 components. The main components of the extract (or their isomers) were determined by mass spectrometry and by comparing the retention times with the literature data. Gallic, ellagic, valoneic acids dilactone were conclusively determined by adding reference standards of these acids to the extract. The substance contains the following compounds: 2,3-hexahydroxydiphenoyl-(α/β)-glucose tr1 = 0.55 min, tr2 = 0.89 min (α and β isomers), 4,6-O-[(S)-valeonyl]-D-glucose (isomer) tr = 0.64 min, gallic acid tr = 1.198 min, pedunculagin tr1 = 3.63 min, tr2 = 4.62 min
    (α and β isomers), proecoxin A (isomer) tr = 4.78 min, valoneic acid dilactone tr = 6.19 min, ellagic acid pentoside (isomer) tr = 7.07 min, ellagic acid tr = 7.335 min.
    Experimental part. The composition analysis was performed using an Agilent 1200 chromatograph with a UV detector, a G6140 mass detector, an Alltech 3300 light scattering detector (ELSD), as well as the Agilent ChemStation Rev.B.04.03 software. The molecular weights of the compounds in the extract were determined using the method of mass spectrometry of ESI-electrospray ionization. The determination of the components was performed using an ultraviolet detector at a wavelength of 280 nm. The column was Rapid Resolution HT Cartige, 4.6 × 30 mm, 1.8 μm, Zorbax SB-C18.
    Conclusions. A new effective method of analysis of the Altabor substance has been developed; it allows
    determining the qualitative and quantitative content of its structural components. The method gives the possibility to control the process of obtaining the Altabor substance, study the dependence of its composition on the conditions of its obtaining, batch number, place, time of the natural raw material collection, and study the composition of other pharmaceutical substances, the plant raw material containing tannins. The advantage of the method is
    the short time (up to 10 min) of analysis using high-performance liquid chromatography at high resolution.
    Key words: gallotannins; ellagitannins; Altabor; chemical composition; structure; HPLC (high-performance liquid chromatography)

    DOI: https://doi.org/10.24959/ophcj.21.202081
  • A novel luminol-based chemiluminescence method for detecting acetylcholine

    M. Ye. Blazheyevskіy, O. V. Koval’ska, K. V. Dynnyk

    Aim. To develop а new simple non-enzymatic method for the determination of acetylcholine (ACh) by the chemiluminescent reaction of luminol under conditions of the enzymatic hydrolysis of acetylcholine (pH 8.5).
    Experimental part. The method proposed is based on the perhydrolysis reaction of ACh by the excess of
    hydrogen peroxide with the formation of peracetic acid. The latter was further determined by the activation effect of the luminol chemiluminescent oxidation reaction in the presence of hydrogen peroxide. The analytical signal was the summary luminescence (Σ) registered within certain time.
    Results and discussion. The pH range of the analytically applicable system was from 8.2 to 8.5. The effect of ACh + H2O2 incubation period on the reaction progress was also studied. The increase of the incubation period enhanced the sensitivity of the method (the limit of detection (LOD)), but because of practical reasons (especially the detection speed) and practical experience the incubation period was set to 30 min. The linear dependence was observed in the acetylcholine chloride concentration range of (0.8 – 2.8) × 10-4 mol/L. While determining acetylcholine chloride in the concentration range of (1.1 – 2.2) × 10-4 mol/L the relative standard deviation (RSD) did not
    exceed 3 % ((X – μ) × 100 %/μ = –0.5…+0.5 %). The Limit of Quantitation (LOQ, 10S) was 7.7 × 10-5 mol/L.
    Conclusions. A new non-enzymatic kinetic method for the chemiluminescent determination of ACh in aqueous solutions and the pharmaceutical formulation Acetylcholinchlorid Injeel® has been proposed. This method is simple, fast, inexpensive, and thus appropriate for the routine ACh quality control in the laboratories of hospitals, pharmaceutical industries and research institutions.
    Key words: acetylcholine; chemiluminescence method

    DOI: https://doi.org/10.24959/ophcj.21.224212
  • Development of methods for standardization of Crocus sativus (saffron) stigmas for inclusion in the draft monograph of the State Pharmacopoeia of Ukraine

    O. O. Mykhailenko, A. G. Kotov, E. E. Kotova, L. M. Sira, V. M. Kovalyov, V. A. Georgiants

    Crocus sativus L. from the Iridaceae family is a medicinal and edible plant that has recently been actively cultivated in Ukraine. Saffron spice is crocus flower stigmas exhibiting a wide range of the pharmacological activity due to its three main bioactive compounds: crocin, picrocrocin and safranal. The quality of this raw material is regulated
    by various normative documents, but there is no monograph in the State Pharmacopoeia of Ukraine (SPhU).

    Aim. To perform a comparative analysis of the monograph “Saffron for homoeopatic preparations” from the Ph. Eur. 9.0, “Saffron. Croci stigma” from the Deutscher arzneimittel codex (DAC) and “Spices – Saffron (Crocus sativus L.)” from the ISO 3632 by the following indicators: description, identifcation (microscopy and TLC), impurities, loss on drying, total ash and quantifcation of crocin, picrocrocin and safranal by UV-Vis-spectrophotometry to clarify the possibility of harmonizing the requirements of the national legal framework for Crocus stigmas with
    the Ph. Eur. 9.0, DAC and ISO 3632. The results of the analysis will be taken into account in the development of the monograph of the national part of the SPhU “Crocus stigmas (saffron)”.
    Results and discussion. The quality indicators of Crocus stigmas and their standardization determined
    by the monograph in the Ph. Eur. 9.0, DAC and ISO 3632 on the raw material studied have been analyzed; some differences in the regulated quality indicators of the raw material have been found. The results of our own research have shown that the samples of the Ukrainian Crocus (saffron) meet all these requirements. The results of the macro- and microscopic examination of the raw material are given. The chromatographic examination has
    been performed in accordance with the Ph. Eur. 9.0 and DAC using crocin as a standard compound; the quantitative determination of crocin, picrocrocin and safranal in 8 samples of Crocus stigmas has been performed by UV-Vis-spectrophotometry in accordance with the ISO 3632. It has been proposed to introduce the following indicators to determine the specifc absorption rate: for crocin (C44H64O24; Mw – 976.70) not less than 180, for safranal (C10H14O; Mw – 150.22) – not less than 30, for picrocrocin (C16H26O7; Mw – 330.37) – not less than 60 calculated with reference to the dried raw material.
    Experimental part. For analysis the dried stigmas of Crocus sativus (saffron) collected in the village
    Lyubimivka, Kherson region, Ukraine (2016-2018), as well as commercial samples of saffron from Morocco,
    Azerbaijan, Spain were used. The analysis of the leading normative documents containing monographs “Crocus stigmas”, among them the Eur. Ph. 9.0, DAC and ISO 3632, was performed. The studies included description of the appearance of the whole Crocus stigmas; loss on drying and the mass fraction of volatile compounds (105 °C, 16 h); the mass fraction of total ash. The amount of picrocrocin, safranal and crocin was measured by direct reading of the absorption of 1 % aqueous solution of stigmas at 257 nm, 330 nm and 440 nm, respectively, using
    a UV-Vis-spectrophotometer.
    Conclusions. The analysis of the requirements to quality of the medicinal raw material – stigmas of Crocus sativus – has been performed based on the monographs: “Saffron for homoeopatic preparations” from the Ph. Eur. 9.0, “Saffron. Croci stigma” from the Deutscher arzneimittel codex and “Spices – Saffron (Crocus sativus L.)” from the ISO 3632. The main criteria for standardization of the raw material have been determined. It has been proposed to introduce the monograph to the national part of the SPhU “Crocus stigmas (saffron)” according to the following requirements: identifcation (macro- and microscopic signs; thin-layer chromatography (crocin); the quantitative
    determination (the content of crocin, picrocrocin and safranal) adapted to the ISO 3632, performed by UV-Vis-spectrophotometry; impurities; loss on drying; total ash.
    Key words: standardization; State Pharmacopoeia of Ukraine; Crocus sativus stigma; saffron; crocin

    DOI: https://doi.org/10.24959/ophcj.21.179933
  • The study of polyphenolic compounds of Pimpinella anisum herb and determination of their antioxidant activity

    U. Umarov, S. V. Kolisnyk, O. V. Kolisnyk, M. Fatkhullaeva, N. K. Chinibekova, M. M. Khamdamov

    Aim. To study the qualitative composition and quantitative content of the polyphenolic compounds in Pimpinella anisum herb and determine their antioxidant activity.
    Results and discussion. Among the polyphenolic compounds contained in anise herb, chlorogenic acid
    (4.409 mg/g) predominates. Signifcant amounts of catechins (3.104 mg/g), apigenin derivatives (3.077 mg/g) and luteolin (1.864 mg/g) also accumulate in the herb. Minor amounts of myricetin (0.105 mg/g) and naringenin (0.019 mg/g) derivatives, rutin (0.189 mg/g), quercetin (0.028 mg/g), apigenin (0.009 mg/g) and hesperetin (0.002 mg/g) are present. According to the research results, the antioxidant activity of polyphenolic compounds of anise herb with reference to ascorbic acid was found to be 67.76 ± 0.05 mmol/g. Rutin exhibited the antioxidant activity at the level of 3979.59 ± 0.08 mmol/g.
    Experimental part. Pimpinella anisum herb collected during the flowering stage in the summer of 2019 in
    the Kharkiv region (Ukraine) was used for analysis. The analysis of 70 % ethanolic extract from anise herb was performed by high performance liquid chromatography using a Prominence LC-20 Shimadzu chromatographic system (Japan) with a SPD-20AV spectrophotometric detector, an Agilent Technologies Microsorb-MV-150 column (reversedphase, C18 modifed silica gel, length – 150 mm, diameter – 4.6 mm, particles size – 5 μm). Identifcation of substances in the extract was carried out by comparing the retention time and the spectral characteristics of the test substances with the same characteristics of the reference standards. The antioxidant activity was determined by the potentiometric method (pH meter – Hanna 2550, with redox electrode EZDO PO50) with reference to ascorbic acid.
    Conclusions. The qualitative composition and quantitative content of polyphenolic compounds in the ethanolic extract of anise herb have been determined by high performance liquid chromatography. The total content of polyphenolic compounds is 17.576 mg/g. The antioxidant activity of polyphenolic compounds of anise herb with reference to ascorbic acid has been found to be 67.76 ± 0.05 mmol/g.
    Key words: polyphenolic compounds; anise; herb; high performance liquid chromatography; antioxidant activity

    DOI: https://doi.org/10.24959/ophcj.21.226276
  • The synthesis of 4-amino-5-(quinolin-2-yl)-4H-1,2,4-triazole-3-thiol and its interaction with aldehydes

    D. М. Zozulynets, A. G. Kaplaushenko, A. S. Korzhova

    Aim. To synthesize 4-amino-5-(quinolin-2-yl)-4H-1,2,4-triazole-3-thiol and study its reactivity in the reaction
    with aldehydes.
    Results and discussion. 4-Amino-5-(quinolin-2-yl)-4H-1,2,4-triazole-3-thiol was synthesized, and a number of 4-(ethyl, aryl)idenamino derivatives were obtained on its basis.
    Experimental part. Using a series of four successive reactions based on quinoline-2-carboxylic acid, 4-amino-5-(quinolin-2-yl)-4H-1,2,4-triazole-3-thiol was synthesized; its interaction with aldehydes allowed to obtain a number of 4-(ethyl, aryl)idenamino derivatives. The structure of all compounds synthesized was confrmed by IR and 1H NMR spectroscopy, as well as by elemental analysis, and their purity by thin layer chromatography.
    Conclusions. 4-Amino-5-(quinolin-2-yl)-4H-1,2,4-triazole-3-thiol has been synthesized. It has been found that its interaction with aldehydes leads to the formation of new 4-((ethyl, aryl)idenamino)-5-(quinolin-2-yl)-4H-1,2,4-triazole-3-thiols.
    Key words: 5-(quinolin-2-yl)-1,2,4-triazole-3-thiol; 4-arylidenamino derivatives; quinaldic acid; biological

    DOI: https://doi.org/10.24959/ophcj.21.188137
  • Validation of the alkalimetry method for the quantitative determination of free organic acids in raspberry leaves

    O. Yu. Maslov, S. V. Kolisnyk, T. A. Kostina, Z. V. Shovkova, E. Yu. Ahmedov, M. A. Komisarenko

     Organic acids are a large group of biologically active compounds that perform important functions in the plant organism. Moreover, all plants, regardless of the species and family, contain organic acids to a small or large extent as organic acids belong to intermediate metabolites arising from the oxidation of proteins and amino acids, fats and carbohydrates.
    Aim. To validate the method of alkalimetry proposed with potentiometric detection of the end-point for the quantitative determination of free organic acids in raspberry leaves.
    Results and discussion. The method proposed was validated according to the International Conference on Harmonization (ICH) guidelines. The linearity was in the concentration range of 40 – 200 % (r2 = 0.9991). The percentage of recovery was found to be in the range of 98.77 – 102.48 %. The repeatability and intermediate precision were 1.58 % and 1.74 %, respectively. The method is accurate and reliable, with the relative standard deviation of less than 2 %.
    Experimental part. Leaves of raspberry were collected in the Kharkiv region during the period of full ripening. A Hanna 2550 pH-meter with a HI 1131P potentiometric electrode was used for alkalimetric titration of free organic acids. The titration was carried out using a microburette with Class A accuracy.
    Conclusions. The alkalimetry method for the quantitative determination of free organic acids in raspberry leaves has been proposed and validated according to the following parameters: specifcity, linearity, accuracy, repeatability, intermediate precision, robustness. It has been confrmed that the method is simple, reliable, accurate and cost-effective.
    Key words: raspberry; leaves; free organic acids; alkalimetry; validation

    DOI: https://doi.org/10.24959/ophcj.21.226278