Globimetula braunii is a hemi-parasitic plant used in African ethnomedicine for the management of microbial infections, rheumatic pain and tumors amongst others. We report the isolation and characterization of eight compounds with their antioxidant and antimicrobial activities. The air-dried powdered leaf was macerated in EtOH/H20 (4:1). The extract was solvent-partitioned into n-hexane, EtOAc, n-BuOH and aqueous fractions. The fractions were screened for their antioxidant properties, using DPPH, FRAP, TAC and FIC assays. Antimicrobial analysis was performed using the micro-broth dilution method. The active EtOAc fraction was purified for its putative compounds on a repeated silica gel column chromatography monitored with TLC-bioautography. The isolated compounds were characterized using spectroscopic methods of UV, FT-IR, NMR and MS. Eight compounds (1–8) were isolated and characterized as 13,27-cycloursane (1), phyllanthone (2), globraunone (3), three phenolics: methyl 3,5-dihydroxy-4-methoxybenzoate (4), methyl 3-methyl-4-hydroxybenzoate (5) and guaiacol (6), as well as two phenol derivatives: 4-formaldehyde phenone (7) and 6-methoxy-2H-inden-5-ol (8). The study identified 4 and 6 as natural antioxidant compounds with potential as antimicrobial agents. Globimetula braunii (Engl.) Van Tiegh. (Family Loranthaceae) is a hemi-parasitic and epiphytic plant that derives water and mineral nutrients from its hosts by means of a specialized root system called “haustorium” + at m/z 410.0, consistent with the molecular formula C30H50 [M − C23H29]+ at m/z 105.3; 1H-NMR  δ ppm: 0.75 , 0.89 , 0.98 , 1.03 , 1.07 , 1.20 , 1.28 ; 13C-NMR  δ ppm: 39.27 (C-1), 22.30 (C-2), 29.71 (C-3), 30.02 (C-4), 59.51 (C-5), 18.26 (C-6), 41.32 (C-7), 39.72 (C-8), 58.25 (C-9), 37.47 (C-10), 30.52 (C-11), 41.55 (C-12), 38.32 (C-13), 42.16 (C-14), 36.03 (C-15), 32.46 (C-16), 53.12 (C-17), 28.19 (C-18), 42.82 (C-19), 35.04 (C-20), 35.65 (C-21), 32.80 (C-22), 32.11 (C-23), 31.80 (C-24), 20.27 (C-25), 17.96 (C-26), 35.36 (C-27), 14.68 (C-28), 18.68 (C-29), 6.84 (C-30).Compound 2 (20 mg): 13,27-cycloursan-3-one (Phyllanthone) isolated as white amorphous powder, m.p. 154–155 °C; ESI-MS: [M]+ at m/z 424.3 consistent with the molecular formula C30H48O, [M − CH2]+ at m/z 410.0, [M − C23H29]+ at m/z 105.3; UV (CHCl3) λmax: 270.50 nm; IR (KBr) υmax cm−1: 2948.3 (SP3 C-H), 2836.5 (SP3 C-H), 1681.0 (C=O of ketone); 1H-NMR  δ ppm: 0.75 , 0.89 , 0.98 , 1.03 , 1.07 , 1.20 , 1.28 ; 13C-NMR  δ ppm: 39.27 (C-1), 22.30 (C-2), 213.18 (C-3), 30.02 (C-4), 59.51 (C-5), 18.26 (C-6), 41.32 (C-7), 39.72 (C-8), 58.25 (C-9), 37.47 (C-10), 30.52 (C-11), 41.55 (C-12), 38.32 (C-13), 42.16 (C-14), 36.03 (C-15), 32.46 (C-16), 53.12 (C-17), 28.19 (C-18), 42.82 (C-19), 35.04 (C-20), 35.65 (C-21), 32.80 (C-22), 32.11 (C-23), 31.80 (C-24), 20.27 (C-25), 17.96 (C-26), 35.36 (C-27), 14.68 (C-28), 18.68 (C-29), 6.84 (C-30).Compound 3 (28 mg): Globraunone, isolated as white amorphous powder, m.p. 220–222 °C; ESI-MS: [M]+ at m/z 554.2, consistent with the molecular formula C37H62O3, base peak M+ at m/z 554.2, [M + H]+ at m/z 555.0, [M − CH3]+ at m/z 539.1, [M − C7H13O3]+ at m/z 409.9, [M − C14H25O3] + at m/z 313.6; UV (CHCl3) λmax: 229.00 nm, 282.00 nm; IR (KBr) υmax cm−1: 3332.2 , 2974.4 (SP3 C-H), 1656.8 , 1381.0–1274.7 (C–O). 1H-NMR  δ ppm: 0.75 , 0.89 , 0.97 , 1.02 , 1.07 , 1.20 , 1.28 ; 13C-NMR  δ ppm: 39.27 (C-1), 22.30 (C-2), 213.19 (C-3), 30.02 (C-4), 59.51 (C-5), 18.26 (C-6), 41.32 (C-7), 39.72 (C-8), 58.25 (C-9), 37.47 (C-10), 30.52 (C-11), 41.55 (C-12), 38.32 (C-13), 42.16 (C-14), 36.03 (C-15), 32.46 (C-16), 53.12 (C-17), 28.19 (C-18), 42.82 (C-19), 35.04 (C-20), 35.65 (C-21), 32.80 (C-22), 32.11 (C-23), 20.27 (C-25), 17.96 (C-26), 35.36 (C-27), 14.68 (C-28), 18.68 (C-29), 6.84 (C-30). C-24 Side Chain: 41.74 (C-1′), 72.77 (C-2′), 30.66 (C-3′), 15.80 (C-4′), 35.57 (C-5′), 35.21 (C-6′), 31.80 (C-7′).Compound 4 (1.22 g): methyl 3,5-dihydroxy-4-methoxybenzoate, isolated as ash amorphous powder, m.p. 160–161 °C; ESI-MS: m/z 198.0 [M]+ consistent with the molecular formula C9H10O5, loss of methoxy at m/z 167.3 [M − 31]+, m/z 154.3 [M − 44]+, loss of methyl ethanoate at m/z 135.2 [M − 61]+, loss of both methoxy and methyl ethanoate at m/z 107.1 [M − 91]+; UV-Vis (MeOH) λmax: 210 nm, 232 nm, 253.0 nm; 1H-NMR:  δ ppm: 3.85 , 3.91 , 4.89 , 7.36 ; 13C-NMR:  δ ppm: 55.26 (C-4-methoxy), 59.72 (C-1-methoxy), 106.80 , 125.72 (C-1), 142.41 (C-4), 152.90 , 168.05 (C-1-ester).Compound 5 (26 mg): methyl 3-methyl-4-hydroxybenzoate, isolated as yellow semi-solid; ESI-MS: [M + H]+ at m/z 167.7, consistent with the molecular formula C9H10O3, base peak M+ at m/z 149.2, [M − 61]+ at m/z 105.2; UV-Vis (MeOH) λmax: 212.50 nm, 242.00 nm, 266.50 nm; IR (KBr) υmax cm−1: 3385.0 , 2939.0 (C-H stretch), 1715.0 , 1586.0 (C=C aromatic), 1336.3–1123.8 ; 1H-NMR:  δ ppm: 2.05 , 3.90 , 4.88 , 6.83 , 7.21 , 7.46 ; 13C-NMR:  δ ppm: 20.35 , 55.38 , 110.74 (C-2), 114.30 (C-5), 116.27 (C-6), 122.43 (C-1), 144.62 (C-3), 150.09 (C-4), 168.74 (C-1-carbonyl ester).Compound 6 (29 mg): Guaiacol, isolated as a reddish-brown semisolid, ESI-MS: m/z 125.1 [M + H]+ consistent with the molecular formula C7H8O2; UV-Vis (MeOH) λmax: 214.5 nm, 241.5 nm, 271.0 nm; IR (KBr) υmax cm−1: 3339.7 , 1638.2 , 1094.0 ; 1H-NMR:  δ ppm: 3.91 , 4.87 , 6.73 , 6.98 , 7.10 ; 13C-NMR:  δ ppm: 55.34 (C-6a), 102.52 (C-4), 108.17 (C-5), 114.83 (C-3), 129.39 (C-6), 144.55 (C-2), 147.65 (C-1).Compound 7 (20 mg): 4-methyl-4-formaldehyde phenone, isolated as brown semi-solid; ESI-MS: m/z 136.0 [M] + consistent with the molecular formula C8H8O2; m/z 119.2 [M − 17]+, m/z 106.8 [M − 29]+, m/z 104.3 [M − 32]+; UV-Vis (MeOH) λmax: 234.5 nm, 254.5 nm, 278.50 nm, 291.5 nm; IR (KBr) υmax cm−1: 2926.0 (C-H stretch), 1716.4 (C=O), 1459.3 (C=C aromatic); 1H-NMR:  δ ppm: 1.81 , 7.21 , 7.74 , 8.48 ; 13C-NMR:  δ ppm: 22.29 (C-4b), 54.45 (C-4), 125.56 , 127.19 , 176.60 (C-4a), 199.17 (C-1).Compound 8 (26 mg): 6-methoxy-2H-inden-5-ol, isolated as yellow semi-solid; ESI-MS: m/z 162.0 [M]+ consistent with the molecular formula C10H10O2; loss of -OH at m/z 145.0 [M − 17]+, loss of -OCH3 at m/z 131.0 [M − 31]+, m/z 114.3 [M − 48]+; UV-Vis (MeOH) λmax: 212 nm, 242 nm, 266 nm; IR (KBr) υmax cm−1: 3367.6 (Phenolic OH), 2931.6 (SP3 − CH), 1586.0 , 1468.0 , 1336.3 , 1213.2 ; 1H-NMR:  δ ppm: 1.31 , 3.90 , 4.88 , 7.08 , 7.21 , 7.46 ; 13C-NMR:  δ ppm: 29.35 (C-1), 55.24 (C-6′), 104.89 (C-2), 110.77 (C-9), 120.56 (C-3), 116.81 (C-8), 116.23 (C-4), 108.91 (C-7), 144.63 (C-6), 147.63 (C-5).Compound 13C NMR spectra of 1 and 2 revealed C-30 compounds, while that of 3 showed 37 signals. The DEPT-135 experiment of 1 showed twelve methylene carbons, twelve methyl and methine carbons and six quaternary carbons. That of 2 differs from 1 by having eleven methylene carbons (one carbon less) attributed to the ketone substituent at δC 213.18 ppm. 1H NMR spectra of the three compounds showed seven methyl protons at δH 0.75–1.28 ppm, methylene at δH 1.50–2.32 ppm and methine at δH 2.50–3.50 ppm. The methyl signals resonated as five singlets and two doublets, which is typical of an α-amyrin (ursane-type) of triterpene.The C 213.18 ppm on both the spectra of 2 and 3 confirmed the presence of a ketone. While the C=O attachment at the C-3 position was based on the HMBC experiment. Upon consideration of the spectra of 1–3 and in comparison with spectra data on similar compounds reported in the literature, they were identified as 13,27-cycloursane (1), 13,27-cycloursan-3-one previously identified as phyllanthone (2) and hexadecahydro-8-hydroxy-9-(2-hydroxy-6-methylheptyl)-1,2,6a,6b,9,12a-hexamethyl-6bHcyclopropa[q]picen-10-one (3), named globraunone [The signal at δ11H,12bH,H-one (3)braunone ,17.1H NMR of 4 showed four signals, which were two aromatic methoxy at δH 3.85 and 3.91 ppm, hydroxy at δH 4.89 ppm and a singlet at δH 7.36 ppm indicating an aromatic proton. The 13C NMR showed nine signals, indicating a C9 compound. The signal at δC 168.05 ppm indicated a carbonyl ester, two pairs of signals, each at 106.80 for olefinic carbons (C-2/C-6) and 152.90 ppm for phenolic carbons (C-3/C-5), typical of an AABB para-substitution pattern [4 as a hydroxy benzoate with strong bands at 3367.6 cm−1 (OH) and 1586.0 cm−1 (R-O-C=O). Based on the available spectra data and in comparison, with literature data, 4 was identified as methyl-3,5-dihydroxy-4-methoxybenzoate, previously isolated from Sacoglottis gabonensis stem bark [The  pattern . The IR tem bark .1H NMR of 5 showed three aromatic protons at δH 6.83, 7.21 and 7.46 ppm: a phenolic OH at δH 4.88 ppm and aromatic methoxy at δH 3.90 ppm. A de-shielded methyl group at δH 2.01 ppm confirmed that it is directly attached to an aromatic ring. There were nine carbon signals on the 13C NMR spectrum. The most de-shielded signal resonated at δC 168.74. IR spectrum showed having a broad phenolic OH band at 3385.0 cm−1, a strong carbonyl C=O band at 1715.0 cm−1, C=C aromatic band at 1586.0 cm−1 and C–O stretching band at 1336.3–1123.8 cm−1. UV absorption at 267 nm showed the excitation of a benzoate skeleton (tabulated as 268 nm). The spectra data of 5 was compared with that of methyl-4-hydroxybenzoate, a bacterial inhibitor previously reported in the bark of Tsuga dumosa [The a dumosa ,21, and 13C NMR spectra of 6 showed seven signals. Based on the HSQC experiment, the key functional groups identified include aromatic methoxy at δC 55.34 and δH 3.91 ppm, four olefinic protons at δH 6.73, 6.98, 7.10 and 7.20 ppm, with carbon signals at δC 114.83, 108.17, 129.39 and 110.97 ppm, respectively. The NMR data of 6 agreed with that of guaiacol reported by Kitanovski et al. [The i et al. .1H NMR of 7 showed a singlet signal at δH 8.48 representing an aldehyde carbonyl, two olefinic protons at 7.21 and 7.74 ppm, typical of an A2B2 para-substitution pattern and a singlet at 1.82 ppm. The aldehydic (1716.4 cm−1) and olefinic (1459.3 cm−1) bands were prominent on the IR spectrum, while the UV-Vis spectrum showed λmax of 254.5 nm, indicative of a benzene nucleus. Based on the comparison of its spectra data with literature data, 7 was identified as 4-methyl-4-formaldehyde phenone [ phenone .8 was showed ten signals on the 13C NMR as a C-10, characterized as δC 55.24 (methoxy), one methylene carbon at δC 29.35, four methine carbons at δC 104.89, 108.91, 110.77 and 116.31 and four quaternary carbons at δC 116.81, 120.56, 144.63 and 147.63 ppm, based on the DEPT135 experiment. HSQC experiment showed that the carbon signals at δC 104.89 and 110.77 ppm were directly attached to the proton signal at δH 7.21, while the carbon signals at δC 116.81 and 108.91 ppm were directly attached to the protons at δH 7.46 and 7.08 ppm, respectively, thus, confirming an AABC ring system [Compound g system .1–3), three phenolics (4–6) and two phenolic derivatives (7 and 8) were isolated and identified in our study of G. braunii living on Leucena leucocephala (Fabaceae), its host, which marked the first report of these compounds in the plant and the family Loranthaceae. Previous phytochemical studies on the plant have shown the presence of tannins, phenolics, flavonoids, terpenoids and sterols [G. braunii living on Piliostigma thonningi (Fabaceae). Perhaps, this new additions to the repository of compounds in G. braunii might have occurred because of plant-host specificity, which was reported to play a critical role in the quality and quantity of constituents elicited by Mistletoes as well as its influence on their biological properties [Three ursane-type triterpenes  improved with further chromatographic separation.TLC-bioautography was used throughout the study. This is a reliable and cost-effective technique to isolate lead compounds by employing a suitable chromatographic process, followed by a biological detection system . The TLCp < 0.05) IC50 values of 8.58 and 154.87 µM in the DPPH and FIC assays, respectively.Quantitative assessments of the antioxidant activity (AOX) of the plant by DPPH, FRAP, TAC and FIC colorimetric assay methods are presented in p < 0.05) better than L-ascorbic acid (AA) with an IC50 value of 11.38 µM. The power (FRAP) and capacity (TAC) of the EtOAc fraction as an antioxidant were one-sixth and halved, respectively, when compared with AA. This implies that the fraction was able to perform its antioxidant role by hydrogen atom transfer (HAT) to the stable DPPH free radical, single electron transfer (SET) in the FRAP and TAC assays and by metal (Fe2+) chelation as in the case of FIC assay [The former was significantly  than AA in the DPPH assay, while, in the FRAP assay, it was 0.76-times as good as a single-electron-donor (SET) when compared with AA.As presented in nd Rajhi  on the spi electron-rich benzene ring and hydroxyl group, which can exhibit both HAT and SET mechanisms of antioxidant action. HAT is a one-step reaction by phenolic O-H to effect bond dissociation enthalpy (BDE). SET is a sequential two-step reaction, which entails proton-loss followed by electron-transfer (SPLET), thus, leading to proton affinity (PA) and electron transfer enthalpy (ETE). These mechanisms have been reported to be responsible for the high antioxidant activities of many simple phenolics including phenolic acids [This could be due to its ic acids .The micro-broth dilution method of microbial susceptibility testing was adopted in the study. It is an objective, high-throughput, cost-effective and quantitative method. It offers high reproducibility, fast generation of MICs, convenience of having pre-prepared panels and the economy of reagents and space that occurs due to the miniaturization of the test, which all make it suitable for the antimicrobial analysis of plant samples. It is also able to assist in generating computerized reports if an automated panel reader is used ,30.G. braunii as shown in 1–3) isolated. Globraunone (3) was only inhibitory against B. subtilis at 2.50 mg/mL and fairly against C. albicans at 5.00 mg/mL. This could be due to the ursanoid (α-amyrin), carbonyl and hydroxyl moieties, all playing key roles in the inhibitory action 3 against C. albicans. Similar natural compounds, such as ursolic acid and its derivatives, have been reported to exhibit inhibitory actions against B. subtilis, MRSA, P. aeruginosa and C. albicans at 0.10–0.25 mg/mL [The antimicrobial activity of the 25 mg/mL .4–6) and their derivatives (7 and 8) demonstrated remarkable antimicrobial properties. Guaiacol 6, a methoxyphenolexhibited the strongest inhibitory activities against B. subtilis (0.63 mg/mL) and C. albicans (1.25 mg/mL) among the compounds. This was closely followed by dihydroxy-4-methoxybenzoate (4) with MIC range of 2.50 and 5.00 mg/mL. However, ciprofloxacin and ketoconazole were 2.5- and 12.5- times better antimicrobial agents compared with guaiacol. Based on the observed antimicrobial activities of the isolated compounds, ranking can be done as follows: Ketoconazole, Ciprofloxacin > Guaiacol 6 > 4 > 8 > 5 > 7 > Globraunone 3 > Phyllanthone 2 > 1 > 50% Aq. MeOH.On the other hand, the phenolic compounds  have been shown to demonstrate remarkable bactericidal and fungicidal activities. These include catechol, caffeic acid, resveratrol and gallic acid amongst others. They all have the phenolic OH and/or other substituents, such as ester, carboxylic, amine, amide and thiol, which have significant antimicrobial properties . Methyl-nhibitor ,21, whilE coli and methicillin-sensitive Staphylococcus aureus (MSSA) at 0.20 mg/mL [50 range of 0.68–1.38 mg/mL against S. aureus, 1.21 mg/mL (capsaicin) against P. aeruginosa and 2.70 mg/mL (eugenol) against E. coli [According to a report, bulbiferates inhibited the growth of 20 mg/mL . In the  E. coli .O-methoxyphenol) (6) and methyl 3,5-dihydroxy-4-methoxybenzoate (4), which were the considerably active antioxidant and antimicrobial compounds identified in this study, could be candidate leads in this respect.These leave more to be desired on the biological potentials of natural methoxyphenols and hydroxybenzoates, especially as antioxidant and antimicrobial agents; thus, guaiacol (Globimetula braunii was collected during the wet season at the Obafemi Awolowo University (OAU), Ile-Ife Campus, Nigeria . It was found parasitizing Leucena leucocephala (Lam) De Wit. (Family Fabaceae), its host tree. It was authenticated at the Ife Herbarium, OAU, Ile-Ife, where a herbarium specimen was deposited with Voucher number IFE 17229.G. braunii were dried at room temperature (25–27 °C). They were powdered (5.0 kg) and macerated with 25 L of EtOH-H2O (4:1) at room temperature for 72 h with frequent agitation. This extraction method was a follow-up process to the previous study on the plant [n-Hexane (600 mL × 4), EtOAc (600 mL × 7) and n-BuOH (300 mL × 2).The leaves of he plant . The filThin-layer chromatography (TLC) bioautography method was used according to Wang et al. . This inAll the chemical reagents, solvents and standards used for antioxidant screening were purchased from Sigma Aldrich .L-ascorbic acid) and (plant fractions/isolated compounds)) at varying concentrations: 50.00, 25.00, 12.50, 6.25 and 3.13 µg/mL. The experiment was carried out in triplicate. The samples were incubated in the dark room for 30 min after which the absorbance was measured at 517 nm on a CamSpec M 107 Spectrophotometer , where methanol (negative control) was used as the blank. The percentage inhibition of DPPH by each test sample was calculated thus:control = Absorbance of negative control, Abssample = Absorbance of test sample. The result was expressed as % inhibition and/or IC50.The DPPH spectrophotometric assay was carried out according to Xiao et al. . A 1 mL 3+) 2,4,6-tri-(2-pyridyl)-1,3,5-triazine (TPTZ) to the bluish ferrous ion (Fe2+) by natural antioxidants at 593 nm absorbance measurement. The ferric reducing power of plant extracts were determined as ascorbic acid equivalent (AAE) from the calibration curve of the positive control (L-ascorbic acid) at concentrations 1000.00, 500.00, 250.00, 125.00, 62.50 and 31.25 µg/mL in methanol [This is based on the reduction of the greenish ferric ion (Femethanol .6+ to Mo5+ by the plant samples and subsequent formation of green phosphate/Mo (V) complex at acidic pH according to Prieto et al. [The TAC assay is based on the reduction of Moo et al. . A 0.3 m2·4H2O and 5 mM ferrozine were diluted 20 times. An aliquot (1 mL) of different concentrations of extract was mixed with 1mL FeCl2·4H2O. After 5 min incubation, the reaction was initiated by the addition of ferrozine (1 mL). The mixture was shaken vigorously, and, after a further 10 min incubation period, the absorbance of the solution was measured spectrophotometrically at 562 nm. The percentage inhibition of ferrozine–Fe2+ complex formation was calculated by using the formula:FIC assay was carried out according to the method of Singh and Rajini . Solutiopost-hoc test on a GraphPad Prism 9 .All quantitative antioxidant data were analyzed using a One-way Analysis of Variance (ANOVA), followed by the Bonferroni Escherichia coli ATCC 25923, Pseudomonas aeruginosa ATCC 10145, Bacillus subtilis NCTC 8236, methicillin-resistant Staphylococcus aureus ATCC 29213 and Candida albicans ATCC 24433.The assay was carried according to the Clinical and Laboratory Standard Institute ,41. Bact8 cfu/mL  were applied to the dilutions of the fraction/isolates, positive controls  and negative control 50% aqueous methanol employing a multipoint inoculator. Plates were incubated at 37 °C for 24 h. for bacteria strains and 25 °C for 72 h for fungal strains, after which all plates were observed for growth of the microorganisms. The minimum dilution of fractions completely inhibiting the growth and killing each organism was taken as the MIC and MBC/MFC. The sample with the lowest range of MIC and the widest spectrum of activity against bacteria and fungi was taken as the most active.Bacteria were maintained on nutrient broth and fungi on Sabouraud Dextrose broth at 4 °C and sub-cultured regularly. Bacteria were grown for 18 h in Nutrient broth and culture suspensions of 10n-Hex , EtOAc , EtOAc , EtOAc-MeOH  and MeOH . Eluates were collected in 20 mL test tubes (1–303). They were bulked into six sub-fractions G1–G6 based on their TLC profiles .The EtOAc fraction  was adsorbed unto 30 g silica gel , dry-packed on a 600 g silica gel stationary phase within a 300 cm × 5 cm glass column . Mobile phase comprising solvent systems of increasing polarity was introduced as thus: 1–4, respectively. The most active sub-fraction G3 (3.5 g), based on TLC-bioautography was further purified on a Silica gel column with mobile phase from DCM  to DCM-MeOH . The eluates (1–187) were collected in 10 mL test tubes and were subsequently bulked into four sub-fractions, G3a–G3d, based on their TLC profiles . A preparative TLC separation of G3c, using DCM-MeOH (96:4) afforded bands i–iv, labelled compounds 5–8.After a 24 h period, sub-fraction G1 (1.2 g) afforded solid deposits a, b and c, while sub-fraction G2 (3.1 g) gave a solid deposit d. Each deposit was washed with 100 mL of MeOH (100%), affording compounds 3 for phenolics and 10% H2SO4 for terpenes. Melting point ranges of the solid compounds were determined on Gallenkamp MPD350-BM 3.5 electrothermal instrument . The UV-Vis absorption was determined within 200–800 nm on Shimadzu UV-1800 UV/Visible Scanning Spectrophotometer: 115 VAC . Infrared spectroscopy was done within the 650–4000 cm−1 transmittance on Cary 630 FTIR Spectrometer . 1H, 13C and 2D  NMR spectra of compounds were recorded as solutions on Bruker AMX-300 Spectrometer , where tetramethylsilane was used as the internal standard.Thin-layer chromatography (TLC) of compounds was performed on aluminum-backed silica gel 60 F254 GF plates . Chemical detection of the class of compound isolated was done by spraying the developed TLC plates with chromogenic reagents, such as 5% FeClpart per million (ppm) relative to the indicated deuterated solvents , integral values (number of protons), multiplicity  and coupling constant (J) in hertz (Hz). Electrospray Ionization Mass Spectrometry (ESI-MS) was performed on a Varian 500-MS ion trap Mass Spectrometer  for molecular weight determination, expressed in mass-to-charge ratio (m/z). ESI-MS analysis was performed at 10 μL/min sample infusion flow rate; 2.56 kV capillary voltage; 3.0 V extraction cone; 475 L/h desolvation-gas flow rate; 80 and 100 °C for the source- and desolvation-gas temperatures, respectively; and 5.82 mm Vernier-probe-adjuster position. The spectrometer scan range was 99.5–1500.5 m/z in the positive mode.Signals were recorded in the order of chemical shifts (δ) in G. braunii led to isolation of eight compounds (1–8) from the most active EtOAc fraction. The compounds were identified based on their spectroscopic data and in comparison with literature reports. They were ursane-type triterpenes (1–3), phenolics (4–6) and phenolic derivatives (7 and 8), all reported for the first time in the plant and in the family Loranthaceae. Guaiacol (6) and methyl 3,5-dihydroxy-4-methoxybenzoate (4) were remarkably antioxidant with considerable antimicrobial potentials.Our activity-guided study on the leaves of


Penicillium solitum MCCC 3A00215 resulted in the isolation of one novel polyketide (1), two new alkaloids (2 and 3), and 22 known (4–25) compounds. The structures of the new compounds were established mainly on the basis of exhaustive analysis of 1D and 2D NMR data. Viridicatol (13) displayed moderate anti-tumor activities against PANC-1, Hela, and A549 cells with IC50 values of around 20 μM. Moreover, 13 displayed potent in vitro anti-food allergic activity with an IC50 value of 13 μM, compared to that of 92 μM for the positive control, loratadine, while indole-3-acetic acid methyl ester (9) and penicopeptide A (10) showed moderate effects .A systematic chemical investigation of the deep-sea-derived fungus  Penicillium solitum is a filamentous fungus associated with the decay of pomaceous fruits during storage +, requiring five degrees of unsaturation. The 1H and 13C NMR spectroscopic data ; δC 14.3 ], one methoxyl , six sp3 methylenes, nine methines including three aliphatic , three olefinic  and three oxygenated  ones, and two non-protonated carbons with one olefinic [δC 135.1 ] and one carbonyl [δC 173.9 ] group. Altogether, the 1H and 13C NMR spectra provided 19 carbons, categorized as one methyl, one methoxyl, six methylenes, nine methines, and two quaternary carbons.Compound pic data , Table 12-6/H2-7/H-8/H-8a/H-1/H2-9/H2-10/H-11/H2-12/H-13/H2-14 and H-1/H-2/H3-16/H-3/H-4, which constructed a long chain of C-5/C-6/C-7/C-8/C-8a/C-1/C-9/C-10/C-11/C-12/C-13/C-14 and C-1 via C-2 to C-16/C-3/C-4  and 5 (+73.3), and further by comparison of their electronic circular dichroism (ECD) spectrum , 7.60 , 8.04 , 8.54 ; δC 120.1 , 122.6 , 122.8 , 131.4 , 134.5 , 139.9 ], an isoprene , glutamic acid , β-aminopropanone , and one acylamide . These five fragments could be connected by the HMBC correlations of H3-11/H3-12 to C-2, H-1 to C-2/C-3a/C-7/C-7a, H2-4 to C-3, and H-14 to C-16 to construct the planar structure of 2  in the same concentration of MeOH (c 0.10), 2 was then deduced to be the enantiomer of 4. Accordingly, 2 was determined as (+)-solitumidine D.Compound ure of 2 , the sam3 presented its molecular formula asC21H28N2O4 by the positive HRESIMS at m/z 395.1947 [M + Na]+. The 1H and 13C NMR spectra consisted of signals almost the same as those of solitumidine B 20D +62.7 ; UV (MeOH) λmax (logε) 237 (3.08) nm; CD (MeOH) (Δε) 204 (−1.90), 233 (+0.53), 236 (+0.53), 245 (+0.40) nm; 1H and 13C NMR data, see m/z 361.1989 [M + Na]+ .15-2): white amorphous solid; [α]20D +6 ; UV (MeOH) λmax (logε) 231 (4.73) nm, 261 (4.25) nm, 325 (3.85) nm; CD (MeOH) (Δε) 203 (+2.62) nm; 1H and 13C NMR data, see m/z 388.1877 [M−H]− .(+)-solitumidine D (3): white amorphous power; [α]20D 0 ; UV (MeOH) λmax (logε) 222 (4.10) nm, 261 (3.70) nm, 291 (3.52) nm; CD (MeOH) (Δε) 203 (−0.48), 299 (−0.04) nm; 1H and 13C NMR data, see m/z 395.1945 [M + Na]+ .(±)-Solitumidine E  and two previously unreported alkaloids (2 and 3), together with 22 known compounds (4–25), were isolated from the deep-sea-derived Penicillium solitum MCCC 3A00215. Viridicatol (13) exhibited weak cytotoxic activities against PANC-1, Hela, and A549 cells with IC50 values of 18, 19, and 24, respectively, while it showed remarkable anti-food allergic activity, with an IC50 value of 13 μM.One new compactin analogue (


Araliopsis soyauxii (Rutaceae) afforded an unreported indolopyridoquinazoline (compound 1) along with nine previously known alkaloids 2–10. In addition, six semi-synthetic derivatives 3a–c, 4b, 5a and 6a were prepared by allylation and acetonidation of soyauxinium nitrate (5), edulinine (3), ribalinine (4) and arborinine (6). The structures and spectroscopic data of five of them are reported herein for the first time. The suggested mechanism for the formation of the new N-allylindolopyridoquinazoline 5a is presented. The structures of natural and derived compounds were determined employing extensive NMR and MS techniques. The absolute configuration of stereogenic centers in compounds 2–4 were determined using NOESY technique and confirmed by the single-crystal X-ray diffraction (SC-XRD) technique. The use of SC-XRD further enabled us to carry out a structural revision of soyauxinium chloride recently isolated from the same plant to soyauxinium nitrate (5). The TAE, fractions, compounds 1–7 and 9, and semi-synthetic derivatives 3a–c, 4b, 5a and 6a were evaluated for their cytotoxic activity towards the cervix carcinoma cell line KB-3-1. No significant activity was recorded for most of the compounds except for 9, which showed moderate activity against the tested cancer cell lines.The chemical investigation of the total alkaloid extract (TAE) of the stem bark of  Araliopsis soyauxii Engl. [syn.Vepris soyauxii (Engl.) Mziray] (Rutaceae) is a rich source of new bioactive substances. Our previous reports provided valuable insights on the biology and chemistry of the studied plant, which contains different types of alkaloids, including quinazoline, furoquinoline, indolopyridoquinazoline, and oxazole  shpurposes .5), edulinine (3), ribalinine (4) and aborinine (6) which led to six semi-synthetic derivatives, among which five  are reported here for the first time   (3) and allyl bromide with sodium hydride as base. They were obtained as yellowish oils isolated by preparative TLC of the dried mixture eluting with petroleum ether/EtOAc (70:30).2′,3′-Diallyledulinine   and N-memg, 13%)  were pre3c can be explained by the following mechanism of a nucleophilic substitution on an unsaturated carbon favoured by the presence of a carbonyl group, as shown in The formation of 3b, 10.2 mg, 34%) (3) and p-toluene sulfonic acid and obtained as white needles.Edulinine-2′,3′-acetonide   was prep4) and sodium hydride with allyl bromide as reagent gave 2-allylribalinine  as a yellow oil (6) and potassium carbonate afforded semisynthetic 1-allylarborinine  also obtained as a yellow oil  afforded N-allylsoyauxinine   . The mecA. soyauxii (TAE), fractions obtained from TAE (FTAE1-FTAE3), natural compounds 1–7 and 9, and semi-synthetic derivatives 3a–c, 4b, 5a, 6a were evaluated for their anticancer properties against the cervix carcinoma cell line (KB-3-1) as described in previous reports ) at least 50 times. Some culture medium was added to the wells to adjust the volume of the wells to the required dilution factor. The dilution prepared from stock solution was added to the wells, and each concentration was tested in six replicates. The control contained the same concentration of DMSO as the first dilution. After incubation at 37 °C and 5.3% CO2-humidified air for 72 h, 30 μL of an aqueous resazurin solution (175 μM) was added to each well. The cells were incubated for 6 h at the same conditions. Thereafter, the fluorescence was measured. The excitation was affected at a wavelength of 530 nm, whereas the emission was recorded at 588 nm. The IC50 were recorded as the values equal to the drug concentrations, at which vitality was 50%, and was calculated as a sigmoidal dose-response curve using GraphPad Prism 4.03.The cytotoxicity assay of some isolated compounds was performed on human cervix carcinoma (KB-3-1) cancer cell lines using the previously reported method . The KB-3, 15.0 mg, 0.051 mmol) was dissolved in dimethylformamide (300 µL), and sodium hydride  was added to the medium. After complete dissolution, allyl bromide  was added and stirred at room temperature for 16 hrs. The reaction progress was monitored with LC-MS, and after the complete conversion of starting material, the reaction was quenched with water. Then, the organic layer was washed with brine water and reduced to dryness using a rotary evaporator. 2′, 3′-diallyledulinine  and N-methylflindersine  were obtainedEdulinine : Yellowish oil; 1H-NMR : δH  7.88 , 7.63 , 7.59 , 7.34 , 5.86 , 5.54 , 5.22 , 5.01 , 4.94 , 4.81 , 4.06 , 4.00 , 3.92 , 3.74 , 3.35 , 3.03 , 2.82 , 1.30 , 1.30 ; 13C-NMR : δC 165.8 (C-1), 163.4 (C-3), 140.2 (C-7a), 137.4(C-2′), 136.4(C-2′), 131.6 (C-6), 124.5 (C-5), 123.5 (C-4), 121.8 (C-2), 119.0 (C-3a), 116.3 (C-3′), 115.8(C-7), 115.4, 83.3 (C-3′’), 79.4 (C-2′’), 64.1 (C-1′), 62.8 (C-1′), 30.3 (NCH3), 21.8 (C-4′’), 23.4 (C-5′’), 21.8 (C-5′’); HRESIMS [M + Na]+ at m/z 394.1978 .N-Methylflindersine (3c): Yellowish oil; 1H-NMR ; δ 7.96 , 7.32 , 7.54 , 7.23 , 6.75 , 5.53 , 3.69 , 1.51 ; 13C-NMR : δC 161.1 (C-1), 118.0 (C-2), 155.3 (C-3), 114.1 (C-3a), 123.2 (C-4), 126.4 (C-5), 130.9 (C-6), 116.2 (C-7), 139.4 (C-7a), 78.6 (C-1′), 118.0 (C-2′), 126.4 (C-3′), 115.4, 28.3 (C-4′), 29.3 (NCH3); HRESIMS [M + Na]+ at m/z 264.0999 .3, 26.2. mg, 0.090 mmol) was dissolved in acetone (3.0 mL), and a catalytic amount of p-toluene sulfonic acid  was added to the reaction mixture and was allowed to stir for four hours at room temperature. The reaction progress was monitored with LC-MS, and after the complete conversion of starting the material, the reaction was quenched with water. The reaction mixture was dissolved in water and extracted with chloroform. The organic layer was washed with brine water and reduced to dryness using a rotary evaporator. Edulinine-2′,3′-acetonide  was thus obtained. Yellow oil; 1H-NMR : δH  7.85 , 7.55 , 7.38 , 7.27 , 4.27 , 3.03 , 2.70 , 4.04 , 3.73 , 1.36 , 1.40 , 1.25 1.26 ; 13C-NMR : δC 163.9 (C-1), 161.6 (C-3), 139.4 (C-7a), 130.4 (C-6), 123.8 (C-5), 122.0 (C-4), 119.9 (C-3a), 117.9 (C-2′’), 114.3 (C-7), 106.6 (C-2), 80.6 (C-3′), 80.4 (C-2′), 62.6 (3-OCH3), 29.7 (NCH3), 28.7 (1′’-CH3), 27.0 (3′’-CH3), 26.0 (5′-CH3), 25.9 (4′-CH3), 23.2 (1′-CH2); HRESIMS [M + Na]+ at m/z 354.1681 .Edulinine  was dissolved in dimethylformamide (300 µL), and sodium hydride  was added to the medium. After complete dissolution, allyl bromide  was added and stirred at room temperature for 16 h. The reaction progress was monitored with LC-MS, and after the complete conversion of the starting material, the reaction was quenched with water and reduced to dryness on a lyophilisator, and 2-allylribalinine  was thus obtained. Yellow oil; 1H-NMR : δH  7.86 , 7.23 , 7.12 , 6.91 , 5.44 , 4.82 , 4.72 , 3.58 , 3.17 , 2.46 , 2.33 , 1.02 ; 1.02 ; 13C-NMR : δC 178.0 (C-8), 156.2 (C-5a), 139.8 (C-4a),135.3 (C-3),132.8 (C-2′),126.3 (C-8a), 124.1 (C-1), 123.8 (C-2), 117.7 (C-3′), 115.9 (C-4), 97.7 (C-7a), 83.2 (C-5), 75.6 (C-6), 71.3 (C-1′), 31.0 (NCH3), 25.6 (C-7), 22.6 (C-2′’), 22.4 (C-1′’); HRESIMS [M + Na]+ at m/z 322.1407 Ribalinine  was dissolved in dimethylformamide (300 µL) and sodium hydride  was added to the medium. After complete dissolution, allyl bromide  was added and stirred at room temperature for 16 hrs. The reaction progress was monitored with LC-MS, and after the complete conversion of the starting material, the reaction was quenched with water and reduced to dryness on a lyophilisator. N-allylsoyauxinine  was obtained. Yellow oil; 1H-NMR ; δH  7.78 , 7.66 , 7.42 , 7.38 , 7.19 , 6.12 , 6.12 , 5.94 , 5.22 , 5.06 , 4.92 . 4.11 , 3.20 , 2.92 , 3.84 ; 13C-NMR : δC 94.7(C-2), 102.4 (C-4), 108.9 (C-5a), 111.0 (C-12), 116.2 (C-3′), 120.7 (C-9), 120.9 (C-10), 123.1 (C-8a), 124.1(C-2′), 125.2 (C-9a), 126.1 (C-11), 134.0(C-13a), 135.3 (C-5), 139.6 (C-12a), 153.9 (C-1), 162.2 (C-13), 165.2 (C-3), 175.6 (C-6), 55.2 (3-OCH3), 47.4 (C-1′), 46.8 (C-7), 29.7 (1-NCH3), 21.4 (C-8); HRESIMS [M + Na]+ at m/z 412.1629 .Soyauxinium nitrate  was dissolved in dimethylformamide (300 µL), and potassium carbonate  was added to the medium. After complete dissolution, allyl bromide  was added and stirred at room temperature for 16 h. The reaction progress was monitored with LC-MS, and after the complete conversion of the starting material, the reaction was quenched with water and reduced to dryness on a lyophilisator. The reaction afforded 1-allylarborinine . Yellow oil; 1H-NMR : δH  8.49 , 7.65 , 7.44 , 7.24 , 6.62 , 6.29 , 5.41 , 5.22 , 4.69 , 4.02 , 3.90 , 3.36 ; 13C-NMR : δC 177.6 (C-9), 158.5(C-2), 153.1 (C-1), 142.5 (C-4a), 142.3(C-5), 138.4(C-3), 133.7(C-6), 127.4(C-8), 123.7(C-8a), 121.8(C-7), 118.1(C-3′),115.1(C-5a), 112.2(C-9a), 93.5(C-4), 76.0(C-1′), 61.6(2-OCH3), 56.3(3-OCH3), 35.1(NCH3); HRESIMS ([M + Na]+) at m/z 348.1200 .Arborinine (Araliopsis soyauxii was evidenced after the chemical analysis of the total alkaloid extract of its stem bark compared to previous reports on the same plant, with the isolation of ten alkaloids, including a new indolopyridoquinazoline alkaloid. This will contribute to the enrichment of the databases of organic compounds from Nature. SC-XRD was used as a powerful tool to determine the absolute and conformational stereochemistry of alkaloids and their counterions. The technique allowed us to carry out a structural revision of the previously described soyauxinium chloride to soyauxinium nitrate. The evaluation of the cytotoxic activity of A. soyauxii alkaloids revealed the isolated compounds to be almost inactive, except for maculine, which showed moderate activity against the KB-3-1 cancer cell line. The inactivity of the semi-synthetic derivatives after allylation and acetonidation reactions allowed us to conclude that increasing the lipophilicity of these compounds alone is insufficient to enhance their cytotoxic activity and the moderate activity of arborinine and soyauxinium nitrate might be related to their free hydroxyl and amine groups, respectively.A completely different chemical profile of


In the original article, there was a mistake in Figure 7 as published. The incorrect y-axis header was miR-27a-3p, miR-27b-3p, miR-20a-5p and miR-20b-5p in Figure 7A. The correct y-axis header in Figure 7A is miR-20a-3p, miR-106a-5p, miR-381-3p and miR-206-3p. Moreover, the incorrect miR-20b-5p graph in Figure 7B has been substituted with the correct miR-20b-5p graph. The corrected In the original article, there was a mistake in the legend for Figure 9 as published. The incorrect legend caption was “miR-27a-3p, miR-27b-3p, miR-20a-5p, and miR-20b-5p expression levels”. The correct legend caption is “miR-20a-3p, miR-106a-5p, miR-381-3p, and miR-206-3p expression levels”.In the original article, there was a mistake in Figure 9 as published. The name of each graph was incorrectly reported as miR-27a-3p, miR-27b-3p, miR-20a-5p, and miR-20b-5p. The correct graph names are miR-20a-3p, miR-106a-5p, miR-381-3p, and miR-206-3p. The corrected The authors apologize for these errors and state that this does not change the scientific conclusions of the article in any way. The original article has been updated.


Larrea tridentata and reported triterpene glycosides and lignan derivatives. In continuation of the preceding studies, 17 lignans and lignan glycosides (1–17) were isolated, including seven new compounds (1–7). Herein, the structure of the new compounds was determined based on spectroscopic analysis and enzymatic hydrolysis. The cytotoxicity of 1–17 against HL-60 human promyelocytic leukemia cells was examined. Compounds 4–11 and 14–16 were cytotoxic to HL-60 cells, with IC50 values in the range of 2.7–17 μM. Compound 6, which was the most cytotoxic among the unprecedented compounds, was shown to induce apoptotic cell death in HL-60 cells.Previously, the authors conducted phytochemical investigations of the aerial parts of  Larrea tridentata is an evergreen small shrub belonging to the Zygophyllaceae family. It grows in the desert areas of the southwestern United States and Mexico, and is commonly called ‘Creosote bush’. It has been used as a medicinal plant for the treatment of a variety of illnesses—including infertility, rheumatism, arthritis, diabetes, gallbladder and kidney stones, pain, and inflammation , a 1,4-disubstituted aromatic ring , two methyl groups , a methylene group , and three methine groups . These spectral features are analogous to those of 8. The glycosidic nature of 1 was corroborated by the strong absorption band of the hydroxy groups (3365 cm−1) in the infrared (IR) spectrum, and the signals of an anomeric proton and carbon  in the 1H- and 13C-NMR spectra. Enzymatic hydrolysis of 1 with β-D-glucosidase in AcOH/AcONa buffer (pH 5.0) yielded 8 and D-glucose. D-Glucose was identified by HPLC analysis using a combination of refractive index and optical rotation detectors. In the heteronuclear multiple bond correlation (HMBC) spectrum of 1, a long-range correlation was observed between H-1″ of β-D-glucopyranosyl (Glc) and C-6 of the aglycone moiety (δC 143.3). The β-anomeric configuration was ascertained based on the large coupling constant of the anomeric proton (J = 8.0 Hz). Therefore, 1 was characterized as 3′-demethoxy-6-demethylisoguaiacin 6-O-β-D-glucopyranoside.Compound 1H- and 13C-NMR spectra of 2 (C30H40O13) were closely related to those of 1; however, signals of an additional anomeric proton and carbon were observed. Furthermore, the molecular formula of 2 had an additional C6H10O5 compared to that of 1. Thus, 2 was suggested to have one more hexopyranosyl group than 1. Compound 2 was enzymatically hydrolyzed under the same conditions as those used for 1 to afford 8 and D-glucose. Analysis of the 1H-1H correlation spectroscopy (COSY), heteronuclear single quantum coherence (HSQC), 1-dimensional selective total correlation spectroscopy (1D-TOCSY), and HSQC-TOCSY data for the sugar moieties indicated the presence of two terminal β-D-glucopyranosyl units  and . The HMBC spectrum of 2 exhibited long-range correlations from H-1″ of Glc (I) to C-6 of the aglycone moiety (δC 143.4), and from H-1′″ of Glc (II) to C-4′ of the aglycone moiety (δC 155.4). Accordingly, 2 was identified as 3′-demethoxy-6-demethylisoguaiacin 6,4′-di-O-β-D-glucopyranoside.The 3 (C36H50O19) was obtained as an amorphous powder. The 1H- and 13C-NMR spectra of 3 were similar to those of 2, except for the signals assignable to the aromatic ring attached to C-1 of the aliphatic moiety. Signals arising from a 1,2,4-trisubstituted aromatic ring  were observed in the 1H- and 13C-NMR spectra of 3. Enzymatic hydrolysis of 3 with β-D-glucosidase yielded 9 and D-glucose. 1H-1H COSY, HSQC, 1D-TOCSY, and HSQC-TOCSY analyses of the sugar moieties revealed the following three terminal β-D-glucopyranosyl units: Glc (I) , Glc (II) , and Glc (III) . The HMBC spectrum of 3 showed correlations between H-1″ of Glc (I) and C-6 of the aglycone moiety (δC 145.4), H-1′″ of Glc (II) and C-3′ of the aglycone moiety (δC 148.3), and between H-1″″ of Glc (III) and C-4′ of the aglycone moiety (δC 147.2). Thus, 3 was determined to be 6,3′-didemethylisoguaiacin 6,3′,4′-tri-O-β-D-glucopyranoside.Compound 4 (C18H20O6) was obtained as an amorphous powder. The 1H- and 13C-NMR spectra of 4 were suggestive of a 1-aryl tetralin-type lignan closely related to 9, and 4 and 9 shared the same 1,2,4-trisubstituted aromatic ring and 1,2,4,5-tetrasubstituted aromatic ring. Compound 4 differed from 9 in the aliphatic ring moiety; that of 4 was composed of two methyl groups , two methine groups , an oxygenated methine group , and an oxygenated quaternary carbon [δC 72.3 (C-3)]. The 1H-1H COSY spectrum of 4 indicated spin-coupling correlations of the H-2 methine proton with the H-1 methine proton and Me-11 methyl protons. The HMBC spectrum of 4 indicated long-range correlations from H-2 to C-3 and C-4, Me-12 to C-2, C-3, and C-4, and from H-4 to C-12. These correlations indicated the introduction of hydroxy groups at C-3 and C-4 -2,3-dimethyl-3,4,6,7-naphthalenetetrol. In the nuclear Overhauser effect spectroscopy (NOESY) spectrum of 4, the NOE correlations between H-1 and Me-11, H-2 and H-4/Me-12/H-2′, and between H-4 and Me-12 are indicative of the 1β, 2α, 3α-hydroxy, and 4α relative configurations -1,2,4-trihydro-1--2,3-dimethyl-3,4,6,7-naphthalenetetrol.Compound  and C-4 . Thus, turations . The cir4 C18HO6 was ob5 (C18H18O6) was collected as an amorphous powder. The IR spectrum of 5 shows absorption bands of the hydroxy groups (3271 cm−1) and carbonyl groups (1715 and 1695 cm−1). The 1H- and 13C-NMR spectra of 5 showed signals of a 1,2,4,5-tetrasubstituted aromatic ring , a 1,2,4-trisubstituted aromatic ring , two methyl groups , two methine groups , an aldehyde group , and a carbonyl carbon [δC 213.2 (C-8)]. The spin-coupling correlation H-7′/H-8′/Me-9′ in the 1H-1H COSY spectrum of 5 indicates the existence of a structural fragment of -C(7′)H-C(8′)H(Me(9′))- ((7′)H-C(8′)H(Me(9′))-C(8)(=O)-Me(9), which was shown to be linked to C-1′ of the 1,2,4-trisubstituted aromatic ring and C-2 of the 1,2,4,5-tetrasubstituted aromatic ring based on the observed HMBC correlations between H-7′ and C-1′/C-2′/C-6′/C-1/C-2/C-3. Furthermore, the aldehyde group was confirmed to be attached to C-1 of the 1,2,4,5-tetrasubstituted aromatic ring based on the long-range correlation from the H-7 aldehyde proton to C-1/C-2/C-6 of the aromatic carbons -2-methyl-3-oxobutyl]-4,5-dihydroxybenzaldehyde.Compound Me(9′))- . The HMB carbons . The con6 (C18H18O3) was obtained as an amorphous powder and is suggested to be a benzofuran-type neolignan, the 1H- and 13C-NMR spectra of which resembled those of 12, except for the signals arising from the aromatic ring bonded to C-2 of the benzofuran moiety. In the 1H-NMR spectrum of 6, the aromatic proton showed a typical 1,2,4-trisubstituted spin-coupling system at δH 6.87 , 6.80 , and 6.76 . Furthermore, the molecular formula of 6 was larger than that of 12 by an oxygen atom. The above data revealed that 6 is the C-3″ hydroxy derivative of 12. Because the proton spin-coupling constant between H-2 and H-3 was 8.6 Hz, and the CD spectrum of 6 was in good agreement with that of 12  was verified based on 1H- and 13C-NMR spectral analysis. Comparison of the 13C-NMR spectrum of 7 with that of 6 showed that the signal assignable to C-4″ was moved downfield by 2.5 ppm, whereas that attributable to C-5″ was shifted upfield by 3.5 ppm. Additionally, an HMBC correlation from δH 3.88 (OMe) to δC 147.2 (C-4″) was detected. The above data revealed that 7 is a C-4″-O-methyl derivative of 6. Accordingly, 7 was deduced to be -2,3-dihydro-2-(3-hydroxy-4-methoxyphenyl)-3-methyl-5-(E)-propenylbenzofuran.The 1–17 to HL-60 cells was evaluated using a modified 3--2,5-diphenyl-2H-tetrazolium bromide (MTT) assay  of the aromatic group(s) diminished the cytotoxicity. In benzofuran-type neolignans , the catechol group  was necessary to induce cytotoxicity (6), and C-4″-O-methylation reduced the cytotoxicity (7). For the tetrahydrofuran-type lignans (13–17), the presence of the catechol group was also essential for exerting cytotoxicity .The cytotoxicity of T) assay . Compoun6 was evaluated. Among the unprecedented compounds, 6 exerted the highest cytotoxicity in HL-60 cells. HL-60 cells were treated with 40 μM of 6 for 24 h, stained with Annexin V and propidium iodide (PI), and analyzed by flow cytometry. The cell populations of early (Q4 area) and late (Q2 area) apoptotic cells, for which the vehicle control was 1.9 ± 0.033% and 4.5 ± 0.38%, increased to 6.5 ± 0.033% and 24 ± 0.87%, respectively , and their morphology was observed under a fluorescence microscope. As shown in 6 for 24 h, the sub-G1 population increased to 25 ± 0.58%, compared to 5.5 ± 0.088% in the vehicle control populations  increased , whereas those in the S phase declined (P4 area)  compared with the vehicle control.The apoptosis-inducing activity of ectively . HL-60 culations . Notably1H-NMR: 400 MHz; 13C-NMR: 100 MHz), Bruker DRX-500 , Bruker AV-500 , or Bruker AV-600  spectrometer using a standard Bruker pulse programs at 300 K . Chemical shifts are rendered as δ values with reference to tetramethylsilane (TMS) as an internal standard. HR-ESI-TOF mass were measured using a Waters Micromass LCT mass spectrometer . Diaion HP-20 porous polymer polystyrene resin , silica gel Chromatorex BW-300 , and ODS silica gel COSMOSIL 75C18-OPN  were employed for CC. Thin layer chromatography (TLC) analysis carried out by precoated silica gel 60F254 or RP18 F254S plates , and the sample spots were detected by spraying the TLC plates with H2SO4/H2O (1:9), followed by heating. The HPLC system was constituted with the following instrument; a Tosoh CCPM pump , a Shodex OR-2 detector , a RheodyneTM injection port , and a Capcell Pak NH2 UG80 . Enzymatic hydrolysis was conducted using β-D-glucosidase . The following materials and biochemical-grade reagents were used for the cell culture assays: a 96-well flat-bottom plate ; RPMI-1640 medium, cisplatin, and MTT (Sigma); penicillin G sodium salt and streptomycin sulfate ; paraformaldehyde and phosphate-buffered saline (PBS) ; fetal bovine serum (FBS) ; a Spectra Classic microplate reader .Optical rotations were measured on a JASCO P-1030 automatic digital polarimeter . IR, UV, and CD spectral data were collected using a FT-IR 620 spectrometer (JASCO), V-630 spectrometer (JASCO), and J-720 spectrometer (JASCO), respectively. NMR spectral data were obtained by a Bruker DPX-400  in 2007. A voucher specimen was kept at the herbarium of the Tokyo University of Pharmacy and Life Sciences.The aerial parts of L. tridentata  were extracted with hot MeOH (15 L × 4 times), and the extract was evaporated under reduced pressure. The MeOH extract (940 g) was loaded on a Diaion HP-20 column and successively eluted with MeOH/H2O (3:7), MeOH/H2O (1:1), MeOH, EtOH, and EtOAc. The MeOH eluted portion  was subjected to silica gel CC and eluted with CHCl3/MeOH/H2O  to obtain 7 fractions (Frs. C-1–C-7). Fr. C-3 was separated by silica gel CC eluted with CHCl3/MeOH  and ODS silica gel CC eluted with MeOH/H2O  to obtain 8 (79 mg) and 13 (5.2 mg). Fr. C-4 was applied to silica gel CC eluted with CHCl3/MeOH/H2O , and ODS silica gel CC eluted with MeOH/H2O  and MeCN/H2O (2:3) to collect 4 (4.6 mg) and 5 (5.0 mg). Fr. C-5 was divided by silica gel CC eluted with CHCl3/MeOH/H2O , and ODS silica gel CC eluted with MeOH/H2O  and MeCN/H2O  to get 1 (11 mg). Fr. C-6 was subjected to silica gel CC eluted with CHCl3/MeOH/H2O , and ODS silica gel CC eluted with MeOH/H2O  and MeCN/H2O  to afford 2 (12 mg) and 3 (7.0 mg). The EtOH eluted portion  was chromatographed on silica gel CC eluted with CHCl3/MeOH  to get 8 fractions (Frs. D-1–D-8). Fr. D-2 was purified using silica gel CC eluted with hexane/EtOAc (5:1) and ODS silica gel CC eluted with MeOH/H2O  to yield 7 (5.3 mg). Fr. D-5 was separated by silica gel CC eluted with CHCl3/MeOH  and hexane/EtOAc  and ODS silica gel CC eluted with MeCN/H2O (1:1) to obtained 6 (12 mg) and 12 (4.7 mg). Fr. D-7 was loaded on silica gel CC eluted with hexane/EtOAc (2:1), hexane/CHCl3/MeOH , and CHCl3/MeOH (19:1), and ODS silica gel CC eluted with MeOH/H2O (3:2) and MeCN/H2O (1:2) to afford 10 (7.8 mg), 11 (2.6 mg), 16 (10 mg), and 17 (12 mg). Fr. D-8 was divided by silica gel CC eluted with CHCl3/MeOH  to furnish 9 (17 mg), 14 (12 mg), and 15 (21 mg).The aerial parts of 1: Amorphous powder; [α]D25 −71.6 ; IR (film) νmax: 3365 (OH), 1613, 1513, 1453 (aromatic ring) cm−1; HR-ESI-TOF-MS m/z: 469.1843 [M + Na]+ ; UV λmax (MeOH): 281 (log ε = 3.74) nm; CD λmax (MeOH) (Δε): 232 (−9.41), 273 (−1.32), 289 (+1.97). 1H-NMR spectral data for the aglycone moiety : δH 6.97 , 6.85 , 6.71 , 6.31 , 3.60 , 2.90 , 2.49 , 2.04 , 1.94 , 0.93 , 0.91 . For 1H-NMR spectral data of the sugar moiety, see 13C-NMR spectral data, see Compound 2: Amorphous powder; [α]D25 −99.8 ; IR (film) νmax: 3363 (OH), 1643, 1507, 1453 (aromatic ring) cm−1; HR-ESI-TOF-MS m/z: 631.2376 [M + Na]+ ; UV λmax (MeOH): 282 (log ε = 3.78) nm; CD λmax (MeOH) (Δε): 237 (−10.2), 270 (−2.66), 286 (+1.92). 1H-NMR spectral data for the aglycone moiety : δH 7.03 , 6.98 , 6.96 , 6.28 , 3.67 , 2.91 , 2.51 , 2.04 , 1.95 , 0.93 , 0.92 . For 1H-NMR spectral data of the sugar moieties, see 13C-NMR spectral data, see Compound 3: Amorphous powder; [α]D25 −104.5 ; IR (film) νmax: 3287 (OH), 1639, 1506, 1453 (aromatic ring) cm−1; HR-ESI-TOF-MS m/z: 809.2808 [M + Na]+ ; UV λmax (MeOH): 281 (log ε = 3.73) nm; CD λmax (MeOH) (Δε): 233 (−11.2), 270 (−3.23), 286 (+1.68). 1H-NMR spectral data for the aglycone moiety : δH 7.54 , 7.51 , 7.38 , 6.85 , 6.68 , 3.76 , 2.70 , 2.31 , 1.92 , 1.91 , 0.79 , 0.69 . For 1H-NMR spectral data of the sugar moieties, see 13C-NMR spectral data, see Compound 4: Amorphous powder; [α]D25 36.8 ; IR (film) νmax: 3348 (OH), 1607, 1521, 1448 (aromatic ring) cm−1; HR-ESI-TOF-MS m/z: 355.1118 [M + Na]+ ; UV λmax (MeOH): 285 (log ε = 3.75) nm; CD λmax (MeOH) (Δε): 244 (+1.13), 275 (+4.64), 294 (−5.53). 1H-NMR spectral data : δH 7.04 , 6.73 , 6.54 , 6.51 , 6.15 , 4.48 , 3.56 , 1.88 , 1.41 , 0.92 . For 13C-NMR spectral data, see Compound 5: Amorphous powder; [α]D25 −87.0 ; IR (film) νmax: 3271 (OH), 1715 and 1695 (C=O), 1667, 1519, 1453 (aromatic ring) cm−1; HR-ESI-TOF-MS m/z: 353.1021 [M + Na]+ ; UV λmax (MeOH): 282 (log ε = 3.97) nm; CD λmax (MeOH) (Δε): 238 (+11.6), 319 (−3.72). 1H-NMR spectral data : δH 10.2 , 7.22 , 7.03 , 6.78 , 6.73 , 6.70 , 5.05 , 3.50 , 2.09 , 1.08 . For 13C-NMR spectral data, see Compound 6: Amorphous powder; [α]D25 95.6 ; IR (film) νmax: 3232 (OH), 1609, 1524, 1486 (aromatic ring) cm−1; HR-ESI-TOF-MS m/z: 283.1386 [M + H]+ ; UV λmax (MeOH): 264 (log ε = 4.37) nm; CD λmax (MeOH) (Δε): 230 (−3.44), 262 (+12.0), 269 (+10.2). 1H-NMR spectral data : δH 7.20 , 7.13 , 6.87 , 6.80 , 6.76 , 6.71 , 6.39 , 6.13 , 5.01 , 3.35 , 1.88 , 1.40 . For 13C-NMR spectral data, see Compound 7: Amorphous powder; [α]D25 113.1 ; IR (film) νmax: 3366 (OH), 1650, 1512, 1484 (aromatic ring) cm−1; HR-ESI-TOF-MS m/z: 297.1489 [M + H]+ ; UV λmax (MeOH): 265 (log ε = 4.62) nm; CD λmax (MeOH) (Δε): 221 (−14.5), 261 (+25.0), 271 (+21.4). 1H-NMR spectral data : δH 7.20 , 7.14 , 6.94 , 6.89 , 6.87 , 6.72 , 6.39 , 6.13 , 5.04 , 3.88 , 3.35 , 1.87 , 1.40 . For 13C-NMR spectral data, see Compound 1–3: Compounds 1 (4.0 mg), 2 (5.1 mg), and 3 (2.7 mg) were independently dealt with β-D-glucosidase (each 30 mg) in AcOH/AcONa  at 28 °C for 42 h, 16 h, and 28 h, respectively. Each reactant was divided by silica gel CC eluted with CHCl3/MeOH/H2O (7:4:1) to afford 8 (1.4 mg) from 1, 8 (1.8 mg) from 2, 9 (0.1 mg) from 3, and sugar fractions , respectively. HPLC analysis of D-glucose were carried out following conditions: solvent, MeCN/H2O (17:3); flow rate, 1.0 mL/min. D-Glucose was specified by comparing the retention time and optical rotation with those of an authentic sample: D-glucose .Enzymatic hydrolysis of 2. HL-60 cells (4 × 104 cells/mL) seeded in a 96-well plate. After 24 h, 4 μL of an EtOH/H2O (1:1) solution containing the test samples was added and 4 μL of EtOH/H2O (1:1) was added into control wells. HL-60 cells treated with each compound for 72 h. Then, cell viability was measured with a modified MTT reduction assay method [4–11 and 14–16, all of which resulted in less than 50% cell growth at a concentration of 20 μM, and the exact concentration at which 50% inhibition (IC50) of cell growth occurred was calculated.HL-60 cells  were maintained in RPMI-1640 medium, which contained heat-inactivated 10% FBS supplemented with L-glutamine, penicillin G sodium salt, and 100 μg/mL streptomycin sulfate. The cells kept in a humidified incubator at 37 °C with 5% COy method . Briefly6 cells/well) were pre-incubated for 4 h in a 6 well plate, and then treated with either EtOH/H2O (1:1) (control), 20 μM of cisplatin, or 40 μM of 6. After 24 h treatment, HL-60 cells were collected and washed with PBS, and incubated for 15 min at 28 °C in 1 × Annexin V binding buffer containing Annexin V-FITC and propidium iodide (PI) as provided by the manufacturer . Apoptotic cells were analyzed by a BD FACSCelestaTM flow cytometer .HL-60 cells (1 × 105 cells/mL) were pre-incubated in a 96 well plate. After 24 h, HL-60 cells were delt with either EtOH/H2O (1:1), 20 μM of cisplatin, or 40 μM of 6 for 24 h. After treatment, the cells fixed with 1% glutaraldehyde for 30 min at 28 °C. Then stained with DAPI (0.5 μg/mL in PBS) for 10 min at 28 °C. Finally, HL-60 cells were observed by a CKX41 fluorescence microscopy  [HL-60 cells  .6 cells/well) were seeded for 4 h in a 6 well plate, and then treated with either EtOH/H2O (1:1), 20 μM of cisplatin, or 40 μM of 6. HL-60 cells were collected using PBS and fixed EtOH/H2O (7:3) overnight at −20 °C. Following procedures were carried out same as previously described [TM flow cytometer (BD Biosicences).HL-60 cells  value of less than 0.001 or 0.05 was considered to represent a statistically significant difference.Statistical analysis was carried out one-way analysis of variance (ANOVA) followed by Dunnett′s test. A probability (L. tridentata was performed to obtain 17 lignans and lignan glycosides (1–17), including seven novel compounds (1–7). Compound 3 has a unique structure with the D-glucosyl moiety on three different aromatic hydroxy groups. Compounds 4–11 and 14–16 are cytotoxic to HL-60 cells, with IC50 values in the range of 2.7–17 μM. Additionally, 6 induced apoptotic cell death in HL-60 cells. The apoptosis-inducing mechanism of 6 is currently under investigation.A phytochemical investigation of the aerial parts of


We performed transformations using 10 strains of fungi, five of which efficiently produced 3-butyl-3-hydroxyphthalide. The product yield, determined by high-performance liquid chromatography, reached 97.6% when Aspergillus candidus AM 386 was used as the biocatalyst. Increasing the scale of the process resulted in isolation yields of 29–45% after purification via reversed-phase thin layer chromatography, depending on the strain of the microorganism used. We proposed different mechanisms for product formation; however, hydration of 3-n-butylidenephthalide seems to be the most probable. Additionally, all phthalides were tested against clinical strains of Candida albicans using the microdilution method. Two phthalides showed a minimum inhibitory concentration, required to inhibit the growth of 50% of organisms, below 50 µg/mL. The 3-n-butylidenephthalide metabolite was generally inactive, and this feature in combination with its low lipophilicity suggests its involvement in the detoxification pathway. The log P value of tested compounds was in the range of 2.09–3.38.Phthalides are bioactive compounds that naturally occur in the family Apiaceae. Considering their potentially versatile applications, it is desirable to determine their physical properties, activity and metabolic pathways. This study aimed to examine the utility of whole-cell biocatalysts for obtaining 3-butyl-3-hydroxyphthalide, which is the metabolite formulated during mammalian metabolism of 3- Angelica acutiloba, Angelica sinensis, Apium graveolens, Cnidium officinale, Levisticum officinale and Ligusticum porteri 2) isolated from particular strains were as follows: −2.4  for A. cylindrospora AM 336; −0.5  for A. candidus AM 386; and −3.6  for C. indicum AM 158.The optical rotation of 3-butyl-3-hydroxyphthalide  of compounds (1),  were recorded on a Bruker Avance DRX-500 spectrometer  in CDCl3. 3-butyl-3-hydroxyphthalide (2)  was measured on JNM-ECZS 400 MHz NMR spectrometer . The spectral data are presented below as well as in the attached NMR spectra .3-1H NMR (500 MHz), δ (ppm): 0.98 , 1.54 , 2.45 , 5.63 , 7.49 , 7.64 , 7.66 , 7.88 .13C NMR (151 MHz), δ (ppm): 13.9 (C-11), 22.6 (C-10), 28.0 (C-9), 109.6 (C-8), 119.8 (C-4), 124.6 (C-12), 125.4 (C-7), 129.5 (C-6), 134.3 (C-5), 139.7 (C-3), 145.9 (C-13), 167.3 (C-1).GC-EIMS 189 (M + 1).2).3-butyl-3-hydroxyphthalide (1H NMR: (400 MHz), δ (ppm): 0.85 , 1.14 , 1.31 , 1.38 , 2.07 , 2.20  7.56 , 7.70 , 7.81 .13C NMR (101 MHz), δ (ppm): 13.9 (C-11), 22.6 (C-10), 25.5 (C-9), 38.7 (C-8), 107.9 (C-3), 122.4 (C-4), 125.6 (C-7), 126.9 (C-12), 130.7 (C-6), 134.8 (C-5), 149.0 (C-13), 168.9 (C-1).3-butyl-3-hydroxyphthalide (2) due to the low volatility was assessed at the GC-EIMS after esterification as 3-butyl-3-methoxyphthalide (3)-GC-EIMS 221 (M + 1).3).3-butyl-3-methoxyphthalide (1H NMR: (500 MHz), δ (ppm): 0.83 , 1.12 , 1.28 , 1.39 , 2.01 , 2.16  3.04 , 7.47 , 7.59 , 7.71 , 7.88 .13C NMR (151 MHz), δ (ppm): 13.9 (C-11), 22.7 (C-10), 25.3 (C-9), 38.3 (C-8), 51.2 (C-3a), 111.1 (C-3), 122.6 (C-4), 125.6 (C-7), 128.2 (C-12), 130.7 (C-6), 134.6 (C-5), 146.8 (C-13), 168.5 (C-1).GC-EIMS 221 (M + 1).n-butylphthalide (4).3-1H NMR (500 MHz), δ (ppm): 0.91 , 1.39 , 1.48 , 1.76 , 2.04 , 5.47 , 7.44 , 7.52 , 7.66 , 7.89 .13C NMR (151 MHz), δ (ppm): 14.0 (C-11), 22.6 (C-10), 27.0 (C-9), 34.6 (C-8), 81.6 (C-3), 121.8 (C-4), 125.8 (C-7), 126.3 (C-12) 129.1 (C-6), 134.1 (C-5), 150.3 (C–13), 170.8 (C-1).GC-EIMS 191 (M + 1).1–4) was 0.1 mg/mL. Dead time (t0) was measured by injecting 1% aqueous NaNO3 solution. The injection volume was 10 μL, the temperature was 35 °C and the flow rate was 1.5 mL/min. The retention time (tR) of the samples was also assessed by RP-HPLC in triplicate using the abovementioned column and isocratic elution. The eluents consisted of methanol and water, both of which were acidified by 1% formic acid v/v at 60–80% of the organic phase. The retention factor (k) was calculated according to the following equation: k = (tR − t0)/t0. A graph of log k as a function of the volume fraction of organic solvent was plotted. The value of the chromatographic lipophilicity index, log kw , was determined by extrapolating this correlation. The chromatographic hydrophobicity index φ0 was also assessed as the volume fraction of methanol that yields a log k value of 0 [P (partition coefficient) values were calculated with the use of ALOGPS 2.1. Pearson’s correlation coefficients between the log P and log kw were determined using GraphPad Prism, and differences with p < 0.05 were considered statistically significant.The concentration of tested compounds (lue of 0 . Theoret1–4) were tested against four Candida albicans strains  using the broth microdilution technique. The medium used for the tests was YPD, consisting of 20 g glucose , 20 g bactopeptone , 10 g yeast extract  and 1 L distilled water; the pH was adjusted to 6.5. The solutions of the compounds were prepared in dimethyl sulfoxide (DMSO) and diluted in YPD to obtain final concentrations in the range of 50–250 µg/mL. Fluconazole was tested at the range of 0.064–64 µg/mL with the exception of the resistant strain 636/20, which was additionally tested in the range of 50–250 µg/mL. Next, 100 µL of each solution was pipetted into wells of a 96-well microtiter plate. The inoculum was standardized to 0.5 McFarland standard and then diluted to obtain the final suspension with a cell density of 0.5 × 103 to 2.5 × 103 CFU/mL. The inoculum size used was 100 µL. The positive control comprised DMSO added in the same concentration as the tested compounds in the inoculum, while the negative control was DMSO diluted in the broth without the addition of inoculum. All samples were tested at least in triplicate. The microtiter plates were incubated at 35 °C for 24 h in a Biosan PST-60 HL ThermoShaker  at 1000 rpm. The fungistatic activity of the compounds was assessed by measuring the absorbance at a wavelength of 595 nm  to determine the MIC50 value .Compounds (2) of bioactive 3-n-butylidenephthalide (1). We show that microbiological transformations can efficiently produce the target compound and can be used to better understand phthalide lactone metabolism. Overall, 10 fungal strains were tested, five of which efficiently produced hydroxy-3-butylphthalide (2). We proposed three mechanisms by which 3-n-butylidenephthalide (1) conversion may occur. Even though hydroxylation remains popular in xenobiotic metabolism, we excluded this pathway based on 13C NMR spectra and suggested the hydration pathway as the most probable mechanism. Valuable contributions of this research include the analysis of the lipophilicity and fungistatic activity of both phthalides (1) and (4) and the metabolite (2). Antifungal assays revealed the potential of both 3-n-butylidenephthalide (1) and 3-n-butylphthalide (4) against clinical isolates of C. albicans, with an MIC50 value below 50 µg/mL. We did not observe the influence of a double bond-containing side chain on the activity of compounds (1) and (4). We noticed weak or even a lack of inhibitory properties for metabolite (2), which has lower lipophilicity and presumably lower permeability than compounds (1) and (4). We did not observe a straightforward correlation between lipophilicity and fungistatic activity. Apart from the lipophilicity, the spatial structure of compounds is the factor that influences their biological activity. The presence of –OH groups in compound (2) or –OCH3 groups in compound (3) causes the decrease of antifungal activity compared to that of 3-n-butylphthalide (4). Compound (1), which has a double bond in its structure, shows similar polarity to 3-butyl-3-methoxyphthalide (3); however, this compound had high biological activity. Considering the possible applications for phthalides, there is a necessity for further research, particularly regarding the biological activity of their metabolites.In this study, we confirmed the utility of microbially catalyzed biotransformation for obtaining the mammalian metabolite (


N1-, N2-, and N3-alkylation products, regardless of the conditions and reagent nature . This reaction offers broad opportunities for expanding the variability of substituents on the nitrotriazole ring in the series of primary and secondary aliphatic, alicyclic, and aromatic substituents, which is undoubtedly crucial for solving the problems related to both high-energy materials development and medicinal chemistry when searching for new efficient bioactive compounds. An efficient methodology for the separation of regioisomeric N-alkyl(aryl)nitrotriazoles has been devised and relies on the difference in their basicity and reactivity during quaternization and complexation reactions. Based on the inaccessible N3-substitution products that exhibit a combination of properties of practical importance, a series of energy-rich ionic systems and coordination compounds were synthesized that are gaining ever-increasing interest for the chemistry of energy-efficient materials, coordination chemistry, and chemistry of ionic liquids.The regularities and synthetic potentialities of the alkylation of 4(5)-nitro-1,2,3-triazole in basic media were explored, and new energetic ionic and nitrotriazole-based coordination compounds were synthesized in this study. The reaction had a general nature and ended with the formation of  The 1,2,3-triazole ring is a major pharmacophore system among nitrogen-containing heterocycles  anti-HI.The favorable properties of the enhanced biological activities of the triazole ring include hydrogen bonding capability under in vivo conditions, a strong dipole moment, high chemical stability , and rigidity .2 groups―are commonly used in the development of efficient high-energy compounds, including ionic ones, that exhibit enhanced technological and operational safety for various applications [1,2,3-Triazole nitro derivatives―five-membered heteroaromatic systems bearing three endocyclic nitrogen atoms and exocyclic explosophoric NOications ,12,13,142 groups promotes enhanced density and increases the number of oxidizing elements  required to oxidize components and maximize the energetic potential of the overall system [The aromatic nature provides the triazole heterocyclic molecule with high thermal and chemical stabilities and a low sensitivity to mechanical stimuli ,10. The l system . The capl system .The high potential for practical application of 1,2,3-triazole derivatives provides for the highly relevant problem of finding directed synthesis methods for various functionalized triazoles-bearing systems.N-substituted 4-nitro-1,2,3-triazoles. The first approach refers to the heterocyclization of nitrogen derivatives that contain activated C–C, N–N, and C–N bonds. A good deal of original papers and review articles report the findings of the first approach [N3-substituted nitro derivatives of 1,2,3-triazoles.There are two well-known, conceptually distinct approaches for the synthesis of approach ,19. HoweN1, N2, and N3 isomers is by alkylating unsubstituted 4-nitro-1,2,3-triazoles. Varying the nature of substituents and their location within the structure of N-substituted nitrotriazoles imparts a specified set of characteristics to the compounds and allows the control of their biological activity, energy performance, complexing, and other useful properties.The rational method for the functionalization of 1,2,3-triazole nitro derivatives for the potential synthesis of 1) are limited, have no systematic nature, and tend to be controversial. Most of the examples described previously regarding the alkylation of triazole 1 in basic media highlight the formation of a mixture of only two isomers [1 and different electrophilic agents in basic media ends with the resulting mixed reaction products related to N1- and N2-isomeric 4-nitro-1,2,3-triazoles. The reaction [1 with propargyl bromide [N3-isomer  has been ascribed by mistake.The alkylation reaction of azoles is usually carried out in the presence of bases. In these reactions, nitrotriazoles manifest properties of ambident nucleophiles. The data available in the literature on the alkylation of 4-nitro-1,2,3-triazole . From tnization  and compnization ,33 reactN3 isomers allow for new quaternization and complexation processes that afforded a series of ionic and coordination compounds that have a set of characteristics combining enhanced energy performance and safety.Thus, the present study was focused on the synthesis and transformations of a wide array of N-alkyl(aryl)-nitrotriazoles differing in substituent types. The present study reports the results of the alkylation of 4-nitro-1,2,3-triazole in alkali whereby the triazole heterocycle is functionalized over all the three endocyclic nitrogen atoms. The unique properties of the 4·7H2O (99.5%), sodium hydroxide (99.6%), potassium hydroxide (85.7%), perchloric acid (72.8%), CuCl2·2H2O (99.0%), diethyl ether (99.2%), and tert-butanol (99.4%) were obtained from Vekton, Saint Petersburg, Russian Federation. Diethyl sulfate (98.0%), iodomethane (99.0%), bromoethane (99.0%), 1-bromopropane (99.0%), 2-bromopropane (99.0%), 1-bromo-3-methylbutane (99.0%), benzyl chloride (99.0%), and 2-ethylhexyl nitrate (97.0%) were procured from Aldrich, St. Louis, MO, USA. 1-Bromobutane (99.0%) was obtained from Sigma-Aldrich, St. Louis, MO, USA. Triazole 1 was prepared by the common procedure [All the reagents and solvents were used as received. Ethanol (EtOH) (96.2%) was obtained from Kirov BioChemPlant, Kirov, Russian Federation. Dimethyl sulfoxide-d6 (DMSO-d6) (99.9%), dimethyl sulfate (99.8%), and sodium carbonate (99.8%) were acquired from Chemical Line Co. Ltd., Saint Petersburg, Russian Federation. Dichloromethane (99.8%), MgSOrocedure .1H and 13C NMR spectra were recorded on a Bruker Avance III spectrometer . 1H NMR spectra were acquired at 400.13 MHz, while 13C NMR spectra were taken at 100.61 MHz. The measurements were conducted at 298 K unless otherwise stated. The spectra were calibrated using residual solvent signals . All NMR spectra of the new compounds are shown in the 1 was synthesized by the procedure reported [The reagents were procured from commercial sources and used as received unless otherwise stated. The commercially available compounds were used without additional purification unless otherwise stated. Triazole reported .Synthesis of triazoles 2–4a, 2–4b, 2–4c, 2–4d, 2–4e, 2–4f, 2–4g, 2–4h . A suspension of triazoles 1  in ethanol (15 mL)  and the corresponding alkali metal hydroxide (25 mmol) was heated to 40 °C with stirring until a solution was prepared. Then, dialkyl sulfate (benzyl chloride)  or alkyl halide (50 mmol) was added and stirred. Low-boiling alkyl halides  were added dropwise with constant stirring. The reaction temperature and time for each case are summarized in 2Cl2 (3 × 25 mL). The reaction mixture in water was cooled, and extraction with CH2Cl2 (3 × 25 mL) was performed. The combined organic layers were washed successively with 3% aqueous sodium carbonate (7.5 mL), water (7.5 mL), dried over MgSO4, and then concentrated in a rotary evaporator. The overall yield and the composition of mixed alkylation products are listed in Synthesis of triazoles 2–4j. To a suspension of triazoles 1  in ethanol (15 mL) was added an equimolar quantity of sodium hydroxide and cyclohexyl nitrate (22.5 mmol) and stirred at 78–80 °C for 13 h. 1H NMR spectroscopy identified the formation of three isomeric N-cyclohexylnitrotriazoles 2–4j (conversion degree of the starting cyclohexyl nitrate did not exceed 1%) in the reaction mixture. The reaction mixture was cooled to room temperature and ethanol was removed in vacuo. To the residue was added water (7.5 mL), the whole mixture was heated to 90–95 °C and held for 25 h with stirring. The product was isolated in a manner similar to the previous procedure. The overall yield and the ration of isomers 2–4j are specified in 4 General procedure for quaternization of mixed 2–4a, 2–4b, 2–4c, 2–4d, 2–4e, 2–4f, 2–4g, 2–4h or 2–4j in t-BuOH-HClO.2–4a, 2–4b, 2–4c, 2–4d, 2–4e, 2–4f, 2–4g, 2–4h or 2–4j (20 mmol) and tert-butanol (20 mmol) in conc. HClO4  was stirred for 12 h, and then, water was added (4 mL). The precipitated product was combined by filtration, washed with water, and dried to yield 1-tert-butyl-3-alkyl-4-nitro-1,2,3-triazolium salts 5a–d, 5g, 5j.A solution of the corresponding mixed triazoles 5e,f,h were isolated as follows: after being diluted, the reaction mixture was extracted with CH2Cl2 (20 mL) to recover products 5e,f,h together with unreacted triazoles 2e,f,h and 3e,f,h. The organic layer was concentrated in a rotary evaporator and the residue was treated with Et2O. The precipitated products were combined by filtration and washed with Et2O to furnish salts 5e,f,h.Salts 5a-j were calculated equivalent to the proportions of N3 isomers 4a–j in mixtures 2–4a, 2–4b, 2–4c, 2–4d, 2–4e, 2–4f, 2–4g, 2–4h, or 2–4j 2O). 1H NMR : δ 0.95 , 1.37–1.46 , 1.75 ), 1.95–2.02 , 4.93 , 10.40  ppm. 13C NMR : δ 13.3 (CH3), 18.8 (CH2), 28.2 ((CH3)3), 29.5 (CH2), 55.0 (N-CH2), 68.7 (N-C), 128.6 (=C-H), 145.1 (C-NO2) ppm. FTIR (KBr): 624, 747, 844, 1030, 1092, 1191, 1334, 1366, 1381, 1469, 1546, 1582, 2944, 2977, 3096 cm−1. Elemental analysis: calcd. C 36.76, H 5.86, N 17.15, O 29.38; found C 36.68, H 5.74, N 17.08, O 29.12.White solid. Yield: 84%, 0.33 g. Mp: 164–165 °C 2O). 1H NMR : δ 0.97 2), 1.67–1.75 , 1.75 ), 1.88–1.93 , 4.95 , 10.40  ppm. 13C NMR : δ 22.4 ((CH3)2), 25.4 (CH), 28.6 ((CH3)3), 31.2 (CH2), 54.2 (N-CH2), 69.2 (N-C), 129.1 (=C-H), 145.5 (C-NO2) ppm. FTIR (KBr): 624, 747, 845, 1093, 1191, 1162, 1380, 1467, 1547, 1584, 2937, 2970, 3103 cm–1. Elemental analysis: calcd. C 38.77, H 6.21, N 16.44, O 28.17; found C 38.70, H 6.14, N 16.38, O 28.14.White solid. Yield: 82%, 0.34 g. Mp: 152–153 °C 2O). 1H NMR : δ 1.71 ), 6.17 , 7.43–7.49 , 10.43  ppm. 13CNMR : δ 28.4 ((CH3)3), 58.3 (N-CH2), 69.3 (N-C), 129.2–129.5 (CHarom), 129.6 (=C-H), 131.0 (Carom), 145.0 (C-NO2) ppm. FTIR (KBr): 626, 693, 717, 830, 855, 1028, 1087, 1190, 1259, 1297, 1342, 1379, 1457, 1544, 1583, 2995, 3100 cm−1. Elemental analysis: calcd. C 43.28, H 4.75, N 15.53, O 26.61; found C 43.12, H 4.68, N 15.48, O 26.57.White solid. Yield: 87%, 0.38 g. Mp: 137–139 °C 2O). 1H NMR : δ 0.87–0.93 , 1.24–1.41 CH2CH2CH2),1.75 ), 2.09–2.15 CH2CH2CH2), 4.84 , 10.42  ppm. 13C NMR : δ 10.4 (CH3), 14.4 (CH3), 22.8 (CH2), 23.1(CH2), 27.9 (CH2), 28.6 ((CH3)3), 29.6 (CH2), 38.1 (CH), 58.3 (N-CH2), 69.2 (N-C), 129.4 (=C-H), 145.6 (C-NO2) ppm. FTIR (KBr): 625, 745, 845, 1095, 1164, 1190, 1379, 1466, 1548, 1587, 2860, 2934, 2961, 3133 cm−1. Elemental analysis: calcd. C 43.92, H 7.11, N 14.63, O 25.07; found C 43.88, H 7.07, N 14.72, O 25.12.White solid. Yield: 79%. Mp: 149–150 °C  and N2-substituted derivatives (3a–j) were separated as follows: readily volatile isomers 3 were removed from mixed liquid compounds 2,3b–f, 2,3h, and 2,3j by vacuum distillation and from mixed crystalline compounds 2,3a and 2,3g by extraction with nonpolar solvents. The residue was crystallized to yield individual N1 isomers 2a–j. The compounds N-methyl-(2–4a), N-ethyl-(2–4b), N-n-propyl-(2–4c), N-isopropyl- (2–4d), N-cyclohexylnitrotriazoles (2–4j), 1-n-butyl-4-nitro- (2e), 1-n-butyl-5-nitro-1,2,3-triazole (4e), 1-i-amyl-4-nitrotriazole (2f), and 1-benzyl-4-nitrotriazole (2g) corresponded by spectral characteristics to the compounds synthesized and characterized in the previous studies [After  studies ,38,39,402-n-Butyl-4-nitro-1,2,3-triazole (3e)1H NMR : δ 0.84 , 1.22–1.31 , 1.87–1.95 , 4.52 , 8.33  ppm. 13C NMR : δ 12.7 (CH3), 19.1 (CH2), 30.8 (CH2), 56.0 (N-CH2), 130.5 (=C-H), 152.9 (C-NO2) ppm. FTIR (KBr): 674, 758, 827, 862, 1025, 1103, 1296, 1340, 1444, 1537, 2876, 2962, 3145 cm−1. Elemental analysis: calcd. C 42.35, H 5.92, N 32.92, O 18.80; found C 42.40, H 5.90, N 32.97, O 18.74. Colorless liquid. Yield: 39%, 1.7 g. Bp: 98–99 °C/1–2 mmHg). 2-i-Amyl-5-nitro-1,2,3-triazole (3f)1HNMR : δ 0.88 2), 1.48–1.56 , 1.81–1.87 , 4.54 , 8.28  ppm. 13C-NMR : δ 21.4 ((CH3)2), 25.0 (CH), 37.5 (CH2), 54.6 (N-CH2), 130.3 (=C-H), 152.9 (C-NO2) ppm. FTIR (KBr): 675, 758, 826, 859, 1024, 1110, 1296, 1342, 1446, 1538, 2872, 2960, 3150 cm−1. Elemental analysis: calcd. C 45.64, H 6.57, N 30.42, O 17.37; found C 45.57, H 6.45, N 30.38, O 17.42.Colorless liquid. Yield 39%, 1.8 g. Bp 95–96 °C/1–2 mmHg). 2-Benzyl-4-nitro-1,2,3-triazole (3g)1H-NMR : δ 5.81 , 7.35–7.40 , 8.74  ppm. 13C-NMR : δ 59.4 (N-CH2), 128.5–128.8 (CHarom), 131.9 (=C-H), 134.1 (Carom), 153.3 (C-NO2) ppm. FTIR (KBr): 3139, 1952, 1774, 1525, 1442, 1383, 1354, 1325, 1309, 1171, 1074, 1028, 930, 891, 829, 760, 710, 691, 673 cm−1. Elemental analysis: calcd. C 52.94, H 3.95, N 27.44, O 15.67; found C 53.01, H 3.89, N 27.40, O 15.72.White solid. Yield: 25%, 1.1 g. Mp: 68–70 °C (EtOH). 2-Ethylhexyl-4-nitro-1,2,3-triazole (3h)1H NMR : δ 0.81–0.87 , 1.22–1.28 CH2CH2CH2), 1.99 CH2CH2CH2), 4.46 , 8.53  ppm. 13C NMR : δ 9.8 (CH3), 13.3 (CH3), 22.4 (CH3), 23.2 (CH2), 27.9 (CH2), 29.8 (CH2), 39.6 (CH), 59.1 (N-CH2), 130.4 (=C-H), 152.9 (C-NO2) ppm. FTIR (KBr): 677, 758, 827, 1026, 1180, 1295, 1341, 1385, 1446, 1541, 2931, 2960 cm−1. Elemental analysis: calcd. C 53.08, H 8.02, N 24.76, O 14.14; found C 53.11, H 8.07, N 24.81, O 14.23.Colorless liquid. Yield: 33%, 1.8 g. Bp: 119–120 °C/−2 mmHg). 2-Cyclohexyl-4-nitro-1,2,3-triazole (3j)1H-NMR : δ 1.20–1.31 , 1.40–1.50 , 1.63–1.67 , 1.75–1.85 , 2.14–2.17 , 4.62–4.70 , 8.71  ppm. 13C-NMR : δ 24.0 (2CH2), 24.5 (CH2), 31.8 (2CH2), 65.0 (N-CH), 131.0 (=C-H), 152.6 (C-NO2) ppm. FTIR (KBr): 696, 760, 833, 882, 986, 1034, 1351, 1405, 1446, 1482, 1536, 1766, 2854, 2928, 3140 cm−1. Elementary analysis: calcd. C 48.97, H 6.16, N 28.56, O 16.31; found C 48.86, H 6.22, N 28.61, O 16.32.White solid. Yield 3.5%, 0.15 g. Mp 43–45 °C (EtOH). Complex 6. To a solution  of mixed nitrotriazoles 2–4c (ratio of 2c:3c:4c = 35:56:9) in 96% EtOH (1 mL) was added a solution  of CuCl2·2H2O in EtOH (2 mL). The whole was stirred at room temperature and held for about 1 month to furnish crystals of complex Cu4OCl6L4, where L = 1-n-propyl-5-nitro-1,2,3-triazole 4c.1H NMR : δ 0.85 , 1.85 , 4.60 , 8.63  ppm. 13C NMR : δ9.4 (N-CH3), 20.8 (N-CH2), 51.4 (N-CH2), 132.1 (C-H), 142.9 (C-NO2) ppm. FTIR (KBr): 697, 745, 802, 835, 877, 997,1098, 1162, 1222, 1294, 1368, 1431, 1464, 1521, 1563, 2879, 2973, 3110, 3156 cm−1.Green crystals. Yield: 0.09 g (4.4%). X-ray CrystallographySingle crystal X-ray diffraction intensity data were collected at 296(2) K using a Bruker APEX-II CCD diffractometer equipped with graphite monochromated MoKα radiation (λ = 0.71073 Å). Data reduction was carried out using the program Bruker SAINT, and an empirical absorption correction was applied with the Bruker SADABS program based on the multi-scan method. The structure of the complex was solved by the direct method (SHELXT-18) and refined by the full-matrix least-square technique (SHELXL-18) with anisotropic thermal parameters. All hydrogen atoms were refined isotropically in riding positions. CCDC 2119451 contains the supplementary crystallographic data of 6. These data can be obtained free of charge from the Cambridge Crystallographic Data Centre.More information can be found in the 1 in basic media. All the three available reaction sites―N1, N2, and N3 heteroatoms―were found to undergo an electrophilic attack, irrespective of the alkylating agent nature , solvent and base types, and reaction temperature and time. All the cases resulted in high yields of three regioisomeric N-alkylation products 2–4 among which N2-substituted derivatives 3 were mostly prevailing . The latter underwent an attack by different electrophilic agents to furnish N-substitution products.At the first stage, the equimolar quantity of alkali produced easily a highly nucleophilic anion from 1 was completed within 5–10 min (Entries 1–4). Prolongation of the reaction involving N-substituted azoles, including N-alkyl-4-nitro-1,2,3-triazoles [1 and diethyl sulfate from 5 min to 10–15 h did not considerably alter the composition and ratio of isomers 2–4b (Entries 4–6).The structure of the alkylating reagent determined significantly the reaction conditions and the ratio of alkylation products. The reaction with dialkyl sulfates (DAS) at 78 °C in ethanol or at 84–90 °C in water in excess of triazoles riazoles ,38, in a1 is reacted with DAS, the alkylating reagent was used in deficiency (0.9 equiv). Since alkyl halides do not engage in the quaternization reaction with N-substituted 4-nitro-1,2,3-triazole, excess alkyl halides (2–3 equiv) were utilized in the alkylation of triazole 1, which is especially important when using lower alkyl halides because of possible reaction losses associated with their low boiling points. When alkyl halides or DAS were employed in ethanol, isomer 3 was prevailing in mixed alkylation products 2–4 in all cases. The proportion of 3 in the mixture was 53–57% . The alkylation conditions using water as the polar solvent increased naturally a proportion of the most polar N1-isomer 2, in which case one of regioisomers 2 or 3  was observed to dominate slightly .To exclude possible quaternization processes when triazole N3-isomer 4 in the mixed alkylation products of triazole 1 and dialkyl sulfates was 11–12% (Entries 1–4) and did not exceed 9% when alkyl halides were used (Entries 7–16). A minimum proportion of isomer 4  was documented in the reaction with ethylhexyl nitrate, which is likely due to the steric effects of the ethylhexyl substituent. Unexpectedly, when cyclohexyl nitrate was used, the composition of the isomers changed dramatically, and the proportion of 4 came up to 18% (Entry 16). Moreover, the alkylation reaction between triazoles 1 and cyclohexyl nitrate in basic media considerably diminished the yield of products 2–4j. The yield of the target N-cyclohexylnitrotriazoles 2–4j in water was not above 8%, and when reacted in alcohol, the products were documented only in the 1H NMR spectra. This is attributed to the tendency of cyclohexyl nitrate to undergo a side elimination reaction to form cyclohexene. The latter did not probably participate in the primary reaction; instead, it was consumed during polymerization. As opposed to the conditions considered, unsaturated alicycles have successfully been used to alkylate azoles in acidic media [The content of minor ic media ,42.N1-substitution product 2h. The proportion of 2h in the mixture reached 64% (Entry 14).Benzyl chloride used as the activated electrophilic reagent afforded mixed regioisomers containing chiefly N1-, N2- and N3-alkyl-4-nitro-1,2,3-triazoles 2–7a–c were estimated from the integral intensities of characteristic proton signals of the cyclic carbon atom of the H-5 heterocycle in the 1H NMR spectra. For the signal assignment in the 1H NMR spectra, the known regularity for alkyl nitrotriazoles [N1-isomer) > δ (H-5 N3-isomer) > δ (H-5 N2-isomer). For example, such signals for compounds 2g (N1-isomer), 3g (N2-isomer), and 4g (N3-isomer) are observed at 9.45, 8.74, and 8.81 ppm, respectively. In addition, the spectrum showed pronounced proton signals from the benzyl substituents: singlets of methyls associated with endocyclic nitrogen atoms were recorded at 5.73, 5.81, and 5.96 ppm for isomers 2g, 3g, and 4g, respectively. The proton signals from the benzyl rings for isomers 2–4g were close and observed near 7.27–7.43 ppm -4-nitro-1,2,3-triazoles 2a–j, 3a–j, 4a–j (N3-derivatives in the series of regioisomeric N-substituted 4(5)-nitro-1,2,3-triazoles have the highest density and enthalpy of formation, making them a promising scaffold for the construction of high-energy compounds.Regioisomerism exerts a significant impact on the physicochemical and energetic characteristics of –j, 4a–j . It was N-alkyl-4-nitro-1,2,3-triazoles as per quantum–chemical predictions is also dependent on the location of the substituent on endocyclic nitrogen atoms in the nitrotriazole ring and increases in the row: N2- <N1- <N3-isomer. The calculated pKBH+ value of 1-methyl-5-nitro-1,2,3-triazole (N3-isomer) is more than 4 log units higher than those of the N1-substituted derivatives [The basicity of ivatives .N3-isomers makes them attractive from the synthetic perspective. The dramatic difference of the N3 isomers from other regioisomers is that they are involved in complexation and quaternization reactions in acidic media. For instance, the use of efficient t-BuOH-HClO4 allows for the selective quaternization of N3-isomers 4a–j. The quaternization of isomers 4a–j proceeded over the N1 atom to furnish 1-(tert-butyl)-3-alkyl-4-nitro-1,2,3-triazolium perchlorates 5a–j in a high yield of 78–87%  precipitated as crystals from the reaction mixture. As the length of the substituent  increased due to the higher solubility, salts 5e,f,h required other isolation conditions: products 5e, f, and h were extracted with dichloromethane from the water-diluted reaction mixture; the solvent was removed in vacuo; they were treated with diethyl ether, and the precipitated products were combined by filtration to yield salts 5e,f,h individually.Nitrotriazolium salts 2–4c with copper(II) chloride afforded coordination compound 6 in which N3-isomer 4c acted as the monodentant ligand (L) due to the N1 atom being involved in the coordination  and N2-substituted derivatives (3a–j) were separated as below. Highly volatile isomers 3 were removed from mixed liquid compounds 2, 3b–f, 2, 3h, 2, and 3j by vacuum distillation, and from mixed crystalline compounds 2, 3a, 2, and 3g by extraction with nonpolar solvents. The residue was crystallized to give individual N1-isomers 2a–j.After 6 (CCDC-2119451) [6 exist in a distorted trigonal-bipyramidal environment, with the N triazole atoms and the central oxygen atom in axial positions. Three chlorine atoms lie in the equatorial plane with the Cu atom deviations by 0.230, 0.186, 0.225, and 0.211 Å directed to the N atom accordingly for Cu1, Cu2, Cu3, and Cu4. The Cu–O, Cu–Cl, and Cu–N lengths range from 1.892(1), 2.3384(6), and 1.972(2) Å to 1.904(1), 2.5233(7), and 1.979(2) Å, respectively. For example, these ranges are close to the analogous ranges for hexakis(µ-chlorido)-(µ-oxido)-tetrakis(1-vinyl-1H-imidazole)-tetra-copper(II) [The structure of complex 2119451)  was unampper(II)  and (µ4-pper(II) . InteresN-alkyl(aryl)-4(5)-nitro-1,2,3-triazoles , including inaccessible N3-substitution products, have been devised herein. Due to the unique physicochemical characteristics , the inaccessible N3-substitutuion products appeared to be quite attractive as cages for the construction of energetic ionic and coordination compounds. The synthesis involved the N-monoalkylation in basic media, quaternization using the highly efficient t-BuOH–HClO4 system, and complexation with transition metal salts. The methodology for the separation of regioisomeric N-alkyl(aryl)nitrotriazoles by quaternization and complexation reactions warranted the synthesis of a range of new energy-efficient nitrotriazole salts and coordination compounds.Synthetic methods for a wide array of