Phytoscreening of Vochysiaceae species: Molecular identification by HPLC-ESI-MS/MS and evaluating of their antioxidant activity and inhibitory potential against human α-amylase and protein glycation

Scientific research based on medicinal plants has been highlighted as a complementary treatment to T2DM, stand out the Vochysiaceae family, which have been widely used in folk medicine by traditional South American communities to treat some diseases. Our study aimed to investigate the antioxidant and antiglycation activities of ethanol extracts of leaves (LF) and stem barks (SB) of Vochysiaceae species, evaluated their capacities to inhibit glycoside and lipid hydrolases related to T2DM and molecular identification by HPLC-ESI-MS/MS. Our main findings indicate that the ethanolic extract of four of eight analyzed plants such as LF and SB of Q. grandiflora, Q. parviflora, V. elliptica and Calisthene major exhibited, respectively, potential of α-amylase inhibition (IC₅₀ of LF: 5.7 ± 0.6, 4.1 ± 0.5, 5.8 ± 0.5, 3.2 ± 0.6 and IC₅₀ of SB: 3.3 ± 0.7, 6.2 ± 2.0, 121.0 ± 8.6 and 11.2 ± 2.8 μg/mL), capacities of antioxidant (ORAC of LF: 516.2 ± 0.1, 547.6 ± 4.9, 544.3 ± 6.1, 442.6 ± 2.4 and ORAC of SB: 593.6 ± 22.3, 497.7 ± 0.8, 578 ± 12.3, 593.6 ± 19.5 µmol trolox eq/g; FRAP of LF: 796.1 ± 0.9, 427.7 ± 22.0, 81.0 ± 1.9, 685 ± 37.9 and FRAP of SB: 947.4 ± 24.9, 738.6 ± 24.3, 98.8 ± 7.9, 970.8 ± 13.9 µmol trolox eq/g; DPPH IC₅₀ of LF: 14.2 ± 1.8, 36.3 ± 6.9, 11.8 ± 1.9, 13.3 ± 1.2 and DPPH IC₅₀ of SB: 16.0 ± 3.0, 15.5 ± 1.9, 126.1 ± 23. 6, 5.3 ± 0.3 μg/mL, respectively) and antiglycation (BSA/Frutose IC₅₀ of LF: 43.1 ± 3.4, 52.1 ± 6.0, 175.5 ± 32, 8, 111.8 ± 14.7 and BSA/Frutose IC₅₀ of SB:, 40.1 ± 11.9, 51.2 ± 16. 7, 46.6 ± 5.7, 53.5 ± 13.6 μg/mL) and presence of polyphenols, such as flavonoids and condensed tannins. The extracts presented low ability to inhibit α-glycosidase and lipase enzymes in the initial assays, with values below 40% of inhibition. In BSA/methylglyoxal, only Q. grandiflora SB, V. eliptica LF and V. tucanorum LF showed activity (IC₅₀: 655.5 ± 208.5, 401.9 ± 135.2 and 617.1 ± 80.6 μg/mL, respectively) and only C. major LF and SB, in Arg/methylglyoxal (IC₅₀: 485.1 ± 130.8 and 468.0 ± 150.5 μg/ml, respectively). This study presented new findings about the biological and pharmacological potential of some species of Vochysiaceae family, contributing to the understanding of the action and efficacy in use of these plants, in their management of postprandial hyperglycemia and in glycation and oxidative processes that contribute to managing diabetes mellitus.     

The megaepifauna of the Dogger Bank (North Sea): species composition and faunal characteristics 1991–2008

During a long-term study in the summer months of the years 1991–2008, 176 megaepifaunal species were recorded through a series of beam trawl surveys on a grid of fixed stations on the Dogger Bank (central North Sea). This paper gives a qualitative overview on species composition throughout the research period, determined from samples collected during 15 cruises. In recent years, a number of species with more oceanic distribution patterns (e.g. species from SW British coasts) has been collected. In spite of these newcomers, there was a slight decrease in total species numbers during the research period.     

Production of reactive oxygen species and reactive nitrogen species by angiosperm stigmas and pollen: potential signalling crosstalk?

Angiosperm stigmas exhibit high levels of peroxidase activity when receptive to pollen. To explore possible function(s) of this peroxidase activity we investigated amounts of reactive oxygen species (ROS), particularly hydrogen peroxide, in stigmas and pollen. Because nitric oxide (NO) was recently implicated in pollen tube growth, we also investigated amounts of NO in pollen and stigmas. Reactive oxygen species accumulation was assessed with confocal microscopy and light microscopy using ROS probes DCFH2-DA and TMB, respectively. NO was assayed using the NO probe DAF-2DA and confocal microscopy. Stigmas from various different angiosperms were found to accumulate ROS, predominantly H2O2, constitutively. In Senecio squalidus and Arabidopsis thaliana high amounts of ROS/H2O2 were localized to stigmatic papillae. ROS/H2O2 amounts appeared reduced in stigmatic papillae to which pollen grains had adhered. S. squalidus and A. thaliana pollen produced relatively high amounts of NO compared with stigmas; treating stigmas with NO resulted in reduced amounts of stigmatic ROS/H2O2. Constitutive accumulation of ROS/H2O2 appears to be a feature of angiosperm stigmas. This novel finding is discussed in terms of a possible role for stigmatic ROS/H2O2 and pollen-derived NO in pollen-stigma interactions and defence.     

Vietnamese species of Taumacera viridis–group with a description of new species (Coleoptera: Chrysomelidae: Galerucinae)

The species of Taumacera viridis species–group from Vietnam are revised. Four species are recognized. Taumacera indica (Jacoby, 1889), T. occipitalis (Laboissière, 1933) and T. variceps (Laboissière, 1933), are redescribed. Taumacera phuquoca sp. nov. from Phu Quoc National Park (southern Vietnam) is described. An identification key is provided for all Vietnamese species.www.zoobank.org/urn:lsid:zoobank.org:pub:396862F9-8A47-4101-A27C-CD0625E68FA1.     

Simultaneous determination of a novel antifibrotic agent and three metabolites in human urine by LC–MS/MS

A robust and validated high performance liquid chromatography tandem mass spectrometry (LC–MS/MS) method has been developed for simultaneous determination of F351 (5-methyl-1-(4-hydroxylphenyl)-2-(1H)-pyridone) and three major metabolites in human urine sample. This assay method has also been validated in terms of selectivity, linearity, lower limit of quantification (LLOQ), accuracy, precision, stability, matrix effect and recovery. Chromatography was carried out on an XTerra RP 18 column and mass spectrometric analysis was performed using an API 4000 mass spectrometer coupled with electro-spray ionization (ESI) source in the positive ion mode. The MRM transitions of m/z 202 → 109, 232 → 93, 282 → 202 and 378 → 202 were used to quantify F351 and three metabolites, respectively. Retention times for F351 and three metabolites were 2.54, 1.38, 1.53 and 1.34 min, respectively. The assay was validated from 20 to 4000 ng/mL for F351 and M1, from 80 to16,000 ng/mL for M2 and M3. Intra- and inter-day precision for all analytes was <6.3%, method accuracy was between −11.2 and 0.3%. This assay was used to support a clinical study where multiple oral doses were administered to healthy subjects to investigate the pharmacokinetics, safety, and tolerability of F351.     

Treatment by acidification followed by solid–liquid separation affects slurry and slurry fractions composition and their potential of N mineralization

The aim of the present work was to assess the effect of treatments by acidification, solid–liquid separation or acidification followed by solid–liquid separation on the physical and chemical composition of pig slurry (S) and pig slurry fractions (non acidified and acidified solid (SF and ASF) and liquid (LF and ALF) fractions), as well as on the potential of N mineralization of these pig slurry derived materials. Acidification strongly decrease the inorganic carbon content of S, SF and LF and it also affects the distribution of P, Ca and Mg between the solid and liquid fraction leading to an ALF more equilibrated than LF in terms of nutrients. Acidification increases the potential of organic N mineralization in SF and decreases the potential of N immobilization in S and LF. It can be concluded that the proposed treatment generates valuable slurry fractions with distinct characteristics and potential of N mineralization that may be incorporated to soil at different periods after sowing to comply with plant nutrient requirements.     

Chemical compositions by using LC–MS/MS and GC–MS and antioxidant activities of methanolic extracts from leaf and flower parts of Scabiosa columbaria subsp. columbaria var. columbaria L.

The members of the Scabiosa genus are one of the traditional medicinal plants used in the treatment of many diseases, in particular the treatment of scabies. In this study, it was aimed to determine antioxidant activities and chemical composition of methanolic extracts of leaves and flowers of Scabiosa columbaria subsp. columbaria var. columbaria. The phenolic contents of both parts of the plant were analyzed by LC–MS/MS. A total of 6 phenolic compounds were determined and chlorogenic acid was the major compound in both flower and leaf parts of the plants, with 5936.052 µg/g and 8021.666 µg/g, respectively. 6 different methods were used to determine the antioxidant activity of the plant parts. Both leaf and flower parts of the plant showed high antioxidant activity in all tested methods and the antioxidant activity values of the leaf part were measured higher than those of the flower part for four tests. The methanol extracts of the plant parts was analyzed with GC–MS and number of the essential oil compounds in the leaf and flower parts were determined as 17 and 13, respectively. Linalool compound was also found to be common in both parts of the plant. The major compounds of the essential oils were identified as 4-Octadecenal (30.01%) in the flower and carvone (35.44%) in the leaf. In addition, terpene derivatives was determined as 90.32% of the highest essential oil group in the leaf, while this value was determined as 1.42% in the flower. For the flower, aromatics were determined as the main component group with 21.31%.     

Metabolomic and elemental analysis of camel and bovine urine by GC–MS and ICP–MS

Recent studies from the author’s laboratory indicated that camel urine possesses antiplatelet activity and anti-cancer activity which is not present in bovine urine. The objective of this study is to compare the volatile and elemental components of bovine and camel urine using GC–MS and ICP–MS analysis. We are interested to know the component that performs these biological activities. The freeze dried urine was dissolved in dichloromethane and then derivatization process followed by using BSTFA for GC–MS analysis. Thirty different compounds were analyzed by the derivatization process in full scan mode. For ICP–MS analysis twenty eight important elements were analyzed in both bovine and camel urine. The results of GC–MS and ICP–MS analysis showed marked difference in the urinary metabolites. GC–MS evaluation of camel urine finds a lot of products of metabolism like benzene propanoic acid derivatives, fatty acid derivatives, amino acid derivatives, sugars, prostaglandins and canavanine. Several research reports reveal the metabolomics studies on camel urine but none of them completely reported the pharmacology related metabolomics. The present data of GC–MS suggest and support the previous studies and activities related to camel urine.     

Determination of mildronate by LC–MS/MS and its application to a pharmacokinetic study in healthy Chinese volunteers

A simple, rapid and accurate liquid chromatography–tandem mass spectrometry (LC–MS/MS) method has been developed and validated for the determination of mildronate in human plasma. Following a simple protein precipitation with methanol, the analyte was separated on a C₁₈ column by isocratic elution with methanol and 10 mM ammonium acetate (55:45; v/v), and then analyzed by mass spectrometry in the positive ion MRM mode. Good linearity was achieved over a wide range of 0.01–20 μg/mL. The intra- and inter-batch precisions (as RSD, %) were less than 7.1%. The average extraction recovery was 87.5%. The method described above has been used, for the first time, to reveal the pharmacokinetics of mildronate injection in healthy subjects. After single intravenously administration of 250, 500 and 1000 mg mildronate, the elimination half-life (t_1/2) were (5.56 ± 1.55), (6.46 ± 1.07) and (6.55 ± 1.17) h, respectively. The Student–Newman–Keuls test results showed that peak plasma concentration (C_max) and the area under the plasma concentration versus time curve from time 0 to 24 h (AUC_0–24) were both linearly related to dose. The pharmacokinetics of mildronate fitted the linear dynamic feature over the dose range studied. The essential pharmacokinetic parameters of multidoses administration intravenously (500 mg, b.i.d) were as follows: t_1/2 was (15.34 ± 3.14) h; C_max was (25.50 ± 3.63) μg/mL; AUC_0–24 was (58.56 ± 5.57) mg h/L. The t_1/2 and AUC of multidoses administration intravenously were different from those of single-dose administration significantly. These findings suggested that accumulation of mildronate in plasma occurred.     

Redox activation of Fe(III)–thiosemicarbazones and Fe(III)–bleomycin by thioredoxin reductase: specificity of enzymatic redox centers and analysis of reactive species formation by ESR spin trapping

Thiosemicarbazones such as Triapine (Tp) and Dp44mT are tridentate iron (Fe) chelators that have well-documented antineoplastic activity. Although Fe–thiosemicarbazones can undergo redox cycling to generate reactive species that may have important roles in their cytotoxicity, there is only limited insight into specific cellular agents that can rapidly reduce Fe(III)–thiosemicarbazones and thereby promote their redox activity. Here we report that thioredoxin reductase-1 (TrxR1) and glutathione reductase (GR) have this activity and that there is considerable specificity to the interactions between specific redox centers in these enzymes and various Fe(III) complexes. Site-directed variants of TrxR1 demonstrate that the selenocysteine (Sec) of the enzyme is not required, whereas the C59 residue and the flavin have important roles. Although TrxR1 and GR have analogous C59/flavin motifs, TrxR is considerably faster than GR. For both enzymes, Fe(III)(Tp)₂ is reduced faster than Fe(III)(Dp44mT)₂. This reduction promotes redox cycling and the generation of hydroxyl radical (HO) in a peroxide-dependent manner, even with low-micromolar levels of Fe(Tp)₂. TrxR also reduces Fe(III)–bleomycin and this activity is Sec-dependent. TrxR cannot reduce Fe(III)–EDTA at significant rates. Our findings are the first to demonstrate pro-oxidant reductive activation of Fe(III)-based antitumor thiosemicarbazones by interactions with specific enzyme species. The marked elevation of TrxR1 in many tumors could contribute to the selective tumor toxicity of these drugs by enhancing the redox activation of Fe(III)–thiosemicarbazones and the generation of reactive oxygen species such as HO.     

Sensitive quantification of sirolimus and everolimus by LC–MS/MS with online sample cleanup

Sirolimus and its derivative everolimus are widely used today as immunosuppressive agents for example in the transplantation medicine. The problematic pharmacokinetic behavior of those substances makes therapeutic drug monitoring mandatory. Therefore, a fast, simple and sensitive high-throughput procedure using online extraction with turbulent flow chromatography for the concurrent measurement of sirolimus and everolimus has been developed. 200 μl of whole blood was mixed with internal standard (23-desmethoxyrapamycin) and the precipitation solution and centrifuged. An aliquot of the supernatant was transferred into autosampler vials. 50 μl of the supernatant was injected into the LC system, where the analytes were extracted using turbulent flow chromatography and thereafter analyzed using reversed phase chromatography. Detection was done by atmospherical pressure chemical ionization (APCI) mass spectrometry in the negative ionization mode. The method has been fully validated and compared to a previously used method. The method was shown to be linear over the entire calibration range (2.2–43.7 μg/l for everolimus and 2.9–51.2 μg/l for sirolimus). The lower limit of quantification was 0.5 μg/l for both compounds. For within-day and between-day analysis, the CV's were <7.6% for everolimus and <8.7% for sirolimus, respectively. The accuracy was between 92.1% and 105% for everolimus and 96.1% and 106% for sirolimus. Recovery ranged between 46.3% and 50.6% for everolimus and 51.2% and 57.2% for sirolimus. The method was demonstrated to be free of matrix effects and comparable to the previously used method. The presented LC–MS/MS method, using turbulent flow chromatography online extraction, allows a fast, simple and reliable determination of everolimus and sirolimus.     

Qualitative and quantitative determination of major saponins in Paris and Trillium by HPLC-ELSD and HPLC–MS/MS

High-performance liquid chromatographic (HPLC) with evaporative light scattering detection (ELSD) and HPLC with electrospray ionization multistage tandem mass spectrometry (HPLC–ESI-MSⁿ) were used to identify and quantify steroid saponins in Paris and Trillium plants. The content of the known saponins such as Paris I, II, III, V, VI, VII, H, gracillin and protodioscin in Paris and Trillium plants was determined simultaneously using the developed HPLC-ELSD method. Furthermore, other 12 steroid saponins were identified by HPLC–ESI(+/−)-MSⁿ detection. In the end, a developed analytical procedure was proved to be a reliable and rapid method for the quality control of Paris and Trillium plants. In addition, the alternative resources for Paris yunnanensis used as a traditional Chinese medicine were discovered according to the hierarchical clustering analysis of the saponin fraction of these plants.