Preparative isolation of paclitaxel and related taxanes from cell cultures of Taxus chinensis using reversed-phase flash chromatography

In this study, paclitaxel, baccatin III, taxuyunnanine C and sinenxane C were successfully separated by reversed-phase flash chromatography on a manually packed C₁₈ column from Taxus chinensis cell culture extract. The crude cell culture extract was first treated with Al₂O₃ column chromatography and then divided into two parts: fraction 1 and fraction 2. Ten milligrams of baccatin III and 19 mg of paclitaxel were obtained from 100 mg dried fraction 1. Fifty-two milligrams of taxuyunnanine C and 11 mg sinenxane C were obtained from 100 mg dried fraction 2. The purities of the four compounds were 98.02%, 98.53%, 98.93% and 98.76%, respectively. Their structures were characterised by using UV, MS and NMR. These results indicate that paclitaxel and related taxanes including baccatin III can be obtained from cell culture in a highly pure state using reversed-phase flash chromatography.     

Simultaneous determination of sulfoxaflor and its metabolites,X11719474 and X11721061, in brown rice and rice straw after field application using LC-MS/MS

The present study was carried out to develop an analytical method for simultaneously detecting and quantifying sulfoxaflor and its metabolites (X11721061, X11719474) in brown rice and rice straw using liquid chromatography–tandem mass spectrometry. The parent compound and its metabolites were extracted and purified using original ‘QuEChERS’ method with modification. The matrix-matched calibration curve of sulfoxaflor and its metabolites in both matrices achieved good linearity with determination coefficients (R²) ≥0.9944. The overall recoveries of sulfoxaflor at two fortification levels (rice: 0.2 and 1.0 mg/kg; rice straw: 0.4 and 2.0 mg/kg) ranged from 97.37% to 107.71% with relative standard deviations (RSDs) <5%. On the other hand, the recoveries of both metabolites (X11721061 and X11719474) at 0.1 and 0.5 mg/kg (rice) and 0.2 and 1.0 mg/kg (rice straw) were satisfactory with values ranging from 83.70 % to 112.60% with RSDs <8%. During storage at ?20°C, the analyte and its metabolites were stable for up to 87 days. The limits of quantification of 0.02 mg/kg were lower than the maximum residue limit (0.2 mg/kg) set by the Korean Ministry of Food and Drug Safety for brown rice. The method was successfully applied to paddy field treated with different programme schedules and a preharvest interval of 7 days was proposed based upon the current study. In sum, the developed method is accurate and reproducible for ensuring the reliable determination of sulfoxaflor (and its metabolites) in harvested rice grain and straw samples from the field. The residual level of parent compound does not seem to pose any hazardous effect and treated rice could be safely used for consumption.     

Anxiety-like behaviour and c-fos expression in rats that inhaled vetiver essential oil

Vetiver essential oil (VEO) has been used in aromatherapy for relaxation. This study aimed to investigate the effects of VEO on an anxiety-related behavioural model (the elevated plus-maze, EPM) and immediate-early gene c-fos in amygdala, known to be involved in anxiety. Male Wistar rats were administered diazepam (1 mg/kg i.p.) for 30 min or inhalated with VEO (1%, 2.5% or 5% w/w) for 7 min prior to exposure to the EPM. Then, the effects of 2.5% VEO, the anxiolytic dose, on c-fos expression in amygdala were investigated. The rats given either 2.5% VEO or diazepam exhibited an anxiolytic-like profile in the EPM. VEO and diazepam significantly increased c-fos expression in the lateral division of the central amygdaloid nucleus (CeL). Therefore, the anxiolytic properties of VEO might be associated with altering neuronal activation in CeL. However, future studies are needed to investigate the precise mechanism of action of VEO.     

Two New Iridoid Esters from the Root and Rhizome of Valeriana jatamansi Jones

Two new iridoid esters, named valerjatadoids A and B ( 1 and 2 , resp.), together with three known iridoid esters, jatamanin O ( 3 ), jatamanvaltrate P ( 4 ), and jatamanvaltrate Q ( 5 ), have been isolated from the root and rhizome of Valeriana jatamansi Jones . The structures of the two new compounds were elucidated by spectroscopic analyses, including 2D‐NMR techniques.     

New Iridoid Triesters from Valeriana jatamansi

Two new iridoid triesters, valeriotriates A ( 1 ) and B ( 2 ), were isolated from the roots of Valeriana jatamansi Jones . Their structures were elucidated by HR‐ESI‐MS and 1D‐ and 2D‐NMR spectroscopy.     

Determination of 5-(3-Chlorophenyl)-4-hexyl-2,4-dihydro-3H-1,2,4-triazole-3-thione in Mouse Brain Tissue by Microwave-Assisted Extraction and High-Performance Liquid Chromatography with Fluorescence Detection

A sensitive and selective method was developed for the determination of 5-(3-chlorophenyl)-4-hexyl-2,4-dihydro-3H-1,2,4-triazole-3-thione (TP-315), an antiepileptic compound, in the tissue homogenate of mouse brains by microwave-assisted extraction and high performance liquid chromatography with fluorescence detection by use of an external standard. The analyses were performed on a C₁₈ column using a mobile phase consisting of 60% acetonitrile in water. Fluorescence detection was performed at an excitation wavelength of 263 nm and an emission wavelength of 411 nm. The method provided a high extraction yield with recoveries exceeding 90%. The precision values expressed by intra- and inter-day relative standard deviations were lower than 5%. The validated procedure was employed to develop a brain concentration profile after intraperitoneal administration of a single oral dose equal to 300 mg per kg of body weight. The time to the maximum anticonvulsant action of TP-315 in a mouse maximal electroshock-induced seizure test was 30 min after administration.     

Evaluation of in vivo analgesic activity of Scrophularia kotscyhana and isolation of bioactive compounds through activity-guided fractionation

The present study was undertaken to evaluate the in vivo analgesic activities of the extracts prepared from the aerial parts and roots of Scrophularia kotscyhana and to isolate the bioactive metabolites from the most active extract. Analgesic activities of all extracts and subextracts at the doses of 5, 10 and 30 mg/kg (i.p.) were examined using hot plate test in mice. Among the tested extracts, MeOH extract prepared from the aerial parts and the n-butanol subextract prepared thereof displayed the best analgesic activity at all doses. Phytochemical studies on n-butanol subextract led to the isolation of two new iridoid glycosides as an inseparable mixture, 8-O-acetyl-4′-O-(E)-(p-coumaroyl)-harpagide (1) and 8-O-acetyl-4′-O-(Z)-(p-coumaroyl)-harpagide (2) along with five known secondary metabolites, β-sitosterol 3-O-β-glucopyranoside (3), apigenin 7-O-β-glucopyranoside (4), apigenin 7-O-rutinoside (5), luteolin 7-O-β-glucopyranoside (6) and luteolin 7-O-rutinoside (7). The iridoid mixture (1 and 2), 3 and 4 elicited significant inhibition of pain at 5 mg/kg dose.     

Simultaneous determination and dissipation behaviour of thifluzamide and difenoconazole in grapes using a QuEChERS method with ultra high-performance liquid chromatography and tandem mass spectrometry

In this study, a modified quick, easy, cheap, effective, rugged, and safe (QuEChERS) method was used for the simultaneous determination of thifluzamide and difenoconazole in grapes by ultra-high-performance liquid chromatography coupled with tandem mass spectrometry (UHPLC–MS/MS) using a Waters Acquity UHPLC BEH C18 column. To compare the effects of different sorbents for thifluzamide and difenoconazole in grapes, samples were extracted with acetonitrile and cleaned up using five different types of sorbents at the same concentration level (0.1 mg/kg). The method was validated at three fortification concentrations (0.05, 0.1, 0.5 mg/kg) with five replicates in each matrix using 40 mg of primary secondary amine (PSA) + 10 mg of graphitised carbon black (GCB) as clean-up sorbents. The recoveries were between 76.3 and 109.3%, and the RSDr (intra-day precision, n = 5) and RSD_R (inter-day precision, n = 15) values ranged from 3.5 to 8.1% and 5.8 to 22.2%, respectively. The limits of quantification (LOQ) for thifluzamide and difenoconazole were 0.07 and 0.04 mg/kg, respectively. The method showed excellent linearity and reliability. The results demonstrated that the method was effective in detecting the two compounds. In this study, we also investigated the dissipation behaviours of thifluzamide and difenoconazole in grapes. The dissipations of thifluzamide and difenoconazole followed the first-order kinetics with the half-lives of 3.9–16.3 days.     

Ayurvedic preparation of Zingiber officinale Roscoe: effects on cardiac and on smooth muscle parameters

The rhizome of the Zingiber officinale Roscoe, a biennial herb growing in South Asia, is commonly known as ginger. Ginger is used in clinical disorders, such as constipation, dyspepsia, diarrhoea, nausea and vomiting and its use is also recommended by the traditional medicine for cardiopathy, high blood pressure, palpitations and as a vasodilator to improve the circulation. The decoction of ginger rhizome is widely used in Ayurvedic medicine. In this papery by high-performance liquid chromatography, we have seen that its main phytomarkers were 6-gingerol, 8-gingerol and 6-shogaol and we report the effects of the decoction of ginger rhizome on cardiovascular parameters and on vascular and intestinal smooth muscle. In our experimental models, the decoction of ginger shows weak negative inotropic and chronotropic intrinsic activities but a significant intrinsic activity on smooth muscle with a potency on ileum is greater than on aorta: EC₅₀ = 0.66 mg/mL versus EC₅₀ = 1.45 mg/mL.     

Evaluation of hepatoprotective activity of aqueous extracts of leaves of Basella alba in albino rats

This study was done to evaluate possible hepatoprotective effects of aqueous leaf extracts of Basella alba in comparison with silymarin in paracetamol-induced hepatotoxicity in albino rats. Six groups of six albino rats each received orally for 6 weeks, vehicle, paracetamol (2 g/kg/day), paracetamol (2 g/kg/day) plus silymarin (50 mg/kg/day), paracetamol (2 g/kg/day) plus B. alba extract (60 mg/kg/day), paracetamol (2 g/kg/day) plus B. alba extract (80 mg/kg/day) and paracetamol (2 g/kg/day) plus B. alba extract (100 mg/kg/day). Hepatoprotective effect was evaluated by comparing serum bilirubin, serum glutamic oxaloacetic transaminase, serum glutamic pyruvic transaminase, proteins, alkaline phosphatase and liver histopathology. Results were represented as mean ± SEM. One-way ANOVA was done followed by post hoc Tukey's test with a highly significance level of P < 0.001. Aqueous leaf extracts of B. alba 100 mg/kg/day orally had significant hepatoprotective effect in paracetamol-induced hepatotoxicity in albino rats. The results were well comparable and even in some respects superior to standard drug silymarin.     

Hilic MS/MS determination of amino acids in herbs of Fumaria schleicheri L., Ocimum basilicum L., and leaves of Corylus avellana L.

The aim of research was to study the content of amino acids using in extracts of Fumaria schleicheri L., Ocimum basilicum L., and Corylus avellana L. by HILIC MS/MS method. Separation of amino acids in the samples was carried out with Acquity H-class UPLC system (Waters, Milford, USA) equipped with SeQuant ZIC-Hilic collumn (2.1 × 150 mm, 3.5 μm) (Merck Millipore, Darmstadt, Germany). The MS/MS fragment ion chromatograms of the test solutions established the presence of 19 amino acids. The obtained results have shown that O. basilicum L. characterized the highest concentrations of different neurogenic amino acids (128.1 mg/kg), comparing with F. schleicheri L. and C. avellana L. (57.72 and 52.91 mg/kg, respectively).     

GC–MS characterisation and biological activity of essential oils from different vegetative organs of Plectranthus barbatus and Plectranthus caninus cultivated in north Italy

Essential oils (EOs) from the roots, stems and leaves of Plectranthus barbatus (A) and Plectranthuscaninus (B), cultivated in north Italy, were obtained by steam distillation and chemically characterised by gas chromatography–mass spectrometry. The highest yields were obtained from roots (268.15 and 673.60 mg/kg from A and B), followed by leaves (64.34 and 26.65 mg/kg) and stems (19.76 and 18.63 mg/kg). A total of 128 structures were identified in A and 121 in B. Fe⁺⁺ chelating and antiradical activities (DPPH and ABTS) were evaluated: root and stem EOs showed the strongest activities, while EOs from leaves did not show relevant activities. All EOs were tested for their in vitro antimicrobial activity, showing optimal growth-inhibition in antibiogram (?>35 mm) and MIC tests (32–64 μg/mL) against Candida albicans, while EOs from leaves of both species showed a good activity (25 < ? < 34 mm, MIC 64–128 μg/mL) against Escherichia coli.     

Chemical composition,protoscolicidal effects and acute toxicity of Pistacia atlantica Desf. fruit extract

The present study was designed to evaluate the chemical composition and scolicidal effects of Pistacia atlantica Desf. extract against protoscoleces of hydatid cysts and its acute toxicity in mice model. Various concentrations of the methanolic extract (5–50 mg/mL) were used for 10–60 min. Viability of protoscoleces was confirmed using eosin exclusion test (0.1%). Acute toxicity was also determined in mice model. The main components were β-myrcene (41.4%), α-pinene (32.48%) and limonene (4.66%). Findings demonstrated that P. atlantica extract at the concentrations of 25 and 50 mg/mL after 20 and 10 min of exposure killed 100% protoscoleces. The LD₅₀ of the intraperitoneal injection of the P. atlantica methanolic extract was 2.43 g/kg and the maximum non-fatal dose was 1.66 g/kg. Obtained results showed the potential of P. atlantica extract as a natural source with no significant toxicity for the production of new scolicidal agent to use in hydatid cyst surgery.     

High-Speed Counter-Current Chromatography Combined with Column Chromatography for Isolation of Methyllycaconitine from Delphinium pseudocyanthum

Preparative high-speed counter-current chromatography (HSCCC) combined with conventional column chromatography (CC) has been used for isolation and purification of methyllycaconitine from Delphinium pseudocyanthum. n-Hexane-ethyl acetate-methanol-water, 1:1:1:2 (v/v), was used as the solvent system for HSCCC. Separation of methyllycaconitine from an HSCCC fraction was successfully achieved by CC on silica gel using chloroform-methanol, 7:1 (v/v), as mobile phase. A total of 113.45 mg methyllycaconitine of purity >95% was obtained from 1.044 g extract of D. pseudocyanthum. Its structure was identified by MS and NMR.

     

Determination of Imidazole, 4-Methylimidazole,and 2-Methylimidazole in Cigarette Additives by Ultra-High Performance Liquid Chromatography

Ultra-high performance liquid chromatography was employed for the determination of imidazole, 4-methylimidazole, and 2-methylimidazole in cigarette additives. Following solid phase extraction and filtration, the analytes were separated using isocratic elution with 5 mmol/L acetonitrile-ammonium formate (80:20, v/v) at 0.5 mL/min. The quantification of these analytes was achieved with the external standard method on a diode-array detector at 215 nm. Baseline separation was achieved within 1.3 min. The linear dynamic ranges for imidazole, 4-methylimidazole, and 2-methylimidazole were between 0.0375 and 18.0300 mg/kg. The limits of detection for the analytes were 0.0094 mg/kg. The recoveries and the relative standard deviations at fortification levels of 0.1322 mg/kg to 1.6220 mg/kg were 95.20% to 101.93% and 0.55% to 2.54%, respectively. The method offers easy operation, rapid analysis, and accurate results, and is suitable for the determination of imidazole, 4-methylimidazole, and 2-methylimidazole in cigarette additives.     

A syringic acid derivative and two iridoid glycosides from the roots of Stachys geobombycis and their antioxidant properties

A new derivative of syringic acid, stageobester A (1) and two iridoid glycosides which including a new one, stageoboside A (2), were isolated from the roots of Stachys geobombycis. Their structures were elucidated on the basis of spectroscopic methods, including extensive NMR and MS spectra. In addition, all the isolates were tested for their antioxidant capacity. Compounds 1 and 2 showed moderate antioxidant activities against DPPH radical, with IC₅₀ values of 113.33 ± 1.53 and 40.33 ± 2.08 mg/L, respectively.     

LC‐MS/MS determination of pogostone in rat plasma and its application in pharmacokinetic studies

Pogostone is an important constituent of Pogostemon cablin (Blanco) Benth., and possesses various known bioactivities. A rapid, simple and sensitive liquid chromatography tandem mass spectrometry (LC‐MS/MS) method was developed for the analysis of pogostone in rat plasma using chrysophanol as internal standard (IS). The analytes were extracted with methanol and separated using a reversed‐phase YMC‐UltraHT Pro C₁₈ column. Elution was achieved with a mobile phase consisting of methanol–water (75:25, v/v) for 5 min at a flow rate of 400 μL/min. The precursor/product transitions (m/z) under MS/MS detection with negative electrospray ionization (ESI) were 223.0 → 139.0 and 253.1 → 224.9 for pogostone and IS, respectively. The calibration curve was linear over the concentration range 0.05–160 µg/mL (r = 0.9996). The intra‐ and inter‐day accuracy and precision were within ±10%. The validated method was successfully applied to the preclinical pharmacokinetic investigation of pogostone in rats after intravenous (5, 10 and 20 mg/kg) and oral administration (5, 10 and 20 mg/kg). Finally, the oral absolute bioavailability of pogostone in rats was calculated to be 70.39, 78.18 and 83.99% for 5, 10 and 20 mg/kg, respectively. Copyright © 2013 John Wiley & Sons, Ltd.     

Iridoid-glycoside isolation and purification from Premna fulva leaves

Premna fulva Craib, rich in iridoid glycosides, is widely used to treat periarthritis, osteoproliferation, pain, and other diseases. However, no studies have reported effective purification methods for obtaining iridoid glycosides as active materials. This paper describes an efficient strategy for separating iridoid glycosides from Premna fulva leaves using high-speed counter-current chromatography and preparative high-performance liquid chromatography. A two-phase solvent system, ethyl acetate/n-butanol/water (7.5:2.5:10, v/v), was selected for high-speed counter-current chromatography separation. The proposed method effectively separated and purified four iridoid glycosides and four lignans, including three new iridoid glycosides ( 4–6 ) and five known compounds ( 1–3, 7, 8 ), from Premna fulva leaves, indicating that high-speed counter-current chromatography combined with prep-HPLC can efficiently isolate catalpol derivatives from the genus Premna. Additionally, the in vitro anti-inflammatory activities of all isolated compounds were analyzed using lipopolysaccharide-stimulated RAW 264.7 cells, and the results indicated that six compounds ( 1 and 3–7 ) exhibited potential anti-inflammatory activities.     

Antidiabetic,cytotoxic and antioxidant activities of oil extracted from Acrocomia aculeata pulp

This study evaluated the hypoglycemic effect of the oil extracted from the Acrocomia aculeata pulp (OPAC) in normoglycemic rats and streptozotocin (STZ), fructose-induced diabetic rat models and its in vitro antioxidant and cytotoxic potential. OPAC (3, 30 or 300 mg/kg, v.o.) significantly decreased (p < 0.05) the high glucose levels induced by a high fructose-diet in rats. Persistent treatment with OPAC for 24 days also reduced the high plasmatic glucose induced by STZ. In normoglycemic animals, OPAC significantly decreased glucose levels. While A. aculeata oil exhibited good in vitro antioxidant activity, no sign of cytotoxicity was observed in LLC-PK1 cells (5–500 μg/mL). OPAC has antidiabetic and antioxidant activities without causing in vitro cytotoxicity.     

Chemical constituents of Oldenlandia pinifolia and their antiproliferative activities

This study describes the chemical constituents of Oldenlandia pinifolia (Wall. Ex G. Don) Kuntze (synonym Hedyotis pinifolia Wall. Ex G. Don) and discusses their anti-proliferative activities. Thirteen compounds were isolated from the n-hexane, ethyl acetate and n-butanol extracts of whole plants O. pinifolia by chromatography method. Their structures were elucidated using MS and NMR analysis and compared with reported data. They are three anthraquinones, a carotenoid, two triterpenes, four iridoid glycosides and three flavonoid glycosides. Among them, 2-methyl-1,4,6-trihydroxy-anthraquinone is a new one, and three compounds were found for the first time in this genus. MTT assay resulted that the n-butanol extract and four isolated compounds inhibited the proliferation of chronic myelogenous leukaemia cells. The results from Hoechst 33343 staining and caspase 3-inducing exhibited that those four tested compounds induced apoptosis and activated caspase 3 (p < 0.05). One of them, isorhamnetin-3-O-β-rutinoside showed the most activity with IC₅₀ value of 394.68 ± 25.12 μM.