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In this paper, the effect of nitric oxide (NO) on the secondary metabolites of Taxus chinensis var. mairei under elevated UV-B radiation was studied. The 5-year-old seedlings were used as test materials. The sodium nitroprusside (SNP) was as the NO donor and Carboxy-PTIO potassium salt (cPTIO) as the NO scavenger. The results showed that the SNP, UV, UV-B+SNP treatments significantly increased the contents of photosynthetic pigments (pa, chlorophyll b, carotenoid and total chlorophyll exhibited CK < cPTIO < UV-B+cPTIO < UV-B < SNP < UV-B+SNP. SNP, cPTIO, UV-B, UV-B+SNP and UV-B+cPTIO significantly increased the concentrations of flavonoids, condensed tannins, total phenolics and taxol (p

To establish a method to simultaneously determine the main five alkaloids of Catharanthus roseus for trace samples, a high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (HPLC-ESI-MS/MS) analysis method was developed.The five Catharanthus alkaloids, vinblastine, vincristine, vinleurosine, vindoline, and catharanthine were chromatographically separated on a C18 HPLC column. The mobile phase was methanol-15 nmol·L(-1) ammonium acetate containing 0.02% formic acid (65 : 35, V/V). The quantification of these alkaloids was based on the Multiple Reaction Monitoring (MRM) mode.This method was validated, and the results achieved the aims of the study. The intra- and inter-day precision and accuracy of the five alkaloids were within 1.2%-11.5% (RSD%) and -10.9%-10.5% (RE%). The recovery rates of the five alkaloids of samples were from 79.9% to 91.5%. The five analytes were stable at room temperature for 2 h, at 4 °C for 12 h, and at -20 °C for two weeks. The developed method was applied successfully to determine the content of the five alkaloids in three plant parts of three batches of C. roseus with a minute amount collected from three regions of China.The HPLC-ESI-MS/MS method can be used for the simultaneous determination of five important alkaloids in trace C. roseus samples.

An ultrasound-assisted procedure for the extraction of pectin from heads ofHelianthus annuusL. (sunflower) was established. A Box–Behnken design (BBD) was employed to optimize the extraction temperature (X1: 30–50°C), extraction time (X2: 20–40 min) and pH (X3: 2.5–3.5) to obtain a high yield of pectin with high degree of esterification (DE) from sunflower heads. Analysis of variance showed that the contribution of a quadratic model was significant for the pectin extraction yield and DE. An optimization study using response surface methodology was performed and 3D response surfaces were plotted from the mathematical model. According to the RSM model, the highest pectin yield (23.11 ± 0.08%) and DE (39.85 ± 0.14%) can be achieved when the UAE process is carried out at 50°C for 40min using a hydrochloric acid solution of pH 3.0. These results suggest that ultrasound-assisted extraction could be a good option for the extraction of functional pectin from sunflower heads at industrial level.

Isoprene and monoterpenes have indirect effects on ozone concentration, methane lifetime and aerosol formation. At the same time, the atmospheric composition change (reflected in elevated CO2and O3concentrations) can affect the speciation of monoterpenes / isoprenoids and magnitude of isoprene and monoterpenes fluxes. Ambient concentrations of CO2and O3were 342 μmolmol-1and 40 nmolmol-1, respectively. The effects of elevated CO2(700 μmolmol-1) and O3(80 nmolmol-1) concentrations on isoprenoid emission rates and patterns fromPinustabulaeformiswere studied in open top chambers (OTCs). The released volatiles were quantified by thermal desorption and gas chromatography with flame ionization detector (GC-FID). The study showed that α-pinene was the main compound emitted from Ptabulaeformis, accounting for more 51.5 - 72.6% of the detected volatile compounds. The emissions of isoprene and monoterpenes were reduced in response to elevated CO2concentrations. Exposure to elevated O3for 3 years significantly promoted the isoprene and monoterpenes emissions (p3concentration increased isoprene emission from Ptabulaeformisby about 270.5 ± 13.5%. The response of monoterpene emission to elevated O3may vary during the course of exposure to the pollutant. The patterns of monoterpene and isoprene emissions under combined elevated CO2and O3concentrations were similar to those under elevated CO2concentration alone.

Equisetumone (1), an unprecedented sesquiterpenoid, which possesses a novel 4,5-olide tricyclic trans-caryophyllane skeleton, was isolated from the Equisetum palustre fern. The structure of 1 was established by extensive spectral data analysis. This usual metabolite was probably derived from the caryophyllane class of sesquiterpenes, and a biosynthetic pathway was proposed.

Proteomics responses and adaptations of Camptotheca acuminata seedlings to drought conditions stimulated by treatment with PEG8000 simulation were investigated. We determined the drought responses of seedlings after 30min, 3 h, and 5 h of treatment with15% PEG8000 . The following 2-DE and PMF identification analysis showed that there are many kinds of proteins involved in the regulation of plants responses to environmental drought. Heat shock protein (HSP) and Late embryo abundant protein (LEA) were discovered to take part in the response of C. acuminata to drought environment. Rubisco LSU was found to help C. acuminata to adapt this arid environment in the way of degradation.

Cultivated C. acuminata was distributed widely in arid area belonging to middle part of China where plant medicinal material industrials were highly developed due to limited wild resources. It was showed that drought situation could result in rapid decrease of leaf RWC lower than 50% after 6 hours of treatment and exerted adverse effects on growth and development of C. acuminata seedlings. In this paper, proteomics was used to investigate protein changes and their ecological adaptation to desiccation with the treatment of 15% PEG8000 solute to one-year C. acuminata seedlings. 2-DE was adopted to reflect total protein changes after 0 and 30 min, 3h and 5h of treatment and the different expressing blots was identified using PMF attempting to demonstrate damage mechanism of desiccation to plants and ecologically adapting. The results showed that the degradation of Rubisco large subunit (Rubisco LSU) occurred and HSP70 was implicated in the modulation process within 5 h treatment.

The allozyme and RAPD technologies in this paper were used to investigate ecologically genetic characteristics of natural population of Leynus chinensis, Ambrosia artemisiifolia and Rhodiola sachalinensis in northeast of China. The populations of them separately displayed characteristics of steady, expansive and endangered. According to the theory of plant life cycle forms established by Zu, they belong to Vegetative growth form (V), Sexual reproduction form (S) and Clone reproduction form (C). The result of allozyme and RAPD analyses indicated that genetic diversity and genetic differentiation in population corresponded with classification by this theory. This will provide an alternative for us to understand and assess the ecologically genetic characteristics of different populations.

Micronized Coenzyme Q10 (CoQ10) is prepared with a rapid expansion of supercritical solution (RESS) process, which does not employ toxic organic solvent. Four factors, namely extraction pressure, extraction temperature, nozzle diameter and precipitation temperature are optimized by a four-level orthogonal array design. The optimum micronization conditions are determined as follows: extraction pressure 25 Mpa, extraction temperature 40°C, nozzle diameter 200 μm, and precipitation temperature 25°C. Under the optimum conditions, micronized CoQ10 with a MPS of 147.9±27.3 nm is obtained. The micronized CoQ10 obtained was characterized by High performance liquid chromatography-mass spectrometry (LC–MS) and High performance liquid chromatography (HPLC) analyses. The results showed that RESS micronization process does not induce degradation of CoQ10 and the obtained CoQ10 particles have higher dissolution rate and solubility compared with unprocessed CoQ10 powder.

Resveratrol (RES) is a naturally occurring triphenolic phytoalexin compound exerting numerous beneficial effects in the organism. It has a wide range of biological activities in vitro as well as in vivo, such as anti-cancer, antioxidant, anti-inflammatory and beneficial cardiovascular effects. But, its low solubility in water led to its poor absorption in vivo and low bioavailability. Bovine serum album (BSA) nanoparticles have emerged as versatile desired carrier systems due to its ready availability, biodegradability, lack of toxicity and immunogenicity with fast development of nano technology. In this study, RES-BSANPS were prepared by a desolvation method and chemical cross-linking with glutaraldehyde successfully. Results controlled conditions (concentration of BSA, 10 mg/ml; pH = 9.0; volume of ethanol, 6 ml; volume of 0.25 % glutaraldehyde, 100 µl; amount of RES, 6.7 mg; cross-linking time, 24 h at room temperature (1 ml/min)) for entrapment efficiency, loading efficiency, mean particle size and zeta potential, were found to be 88.7 %, 39.4 %, 175.4 ± 0.5 nm, -35.93 ± 0.79 mV, respectively.

Epigallocatechin-3-gallate (EGCG), a principal polyphenolic, which is most abundant and active component in tea. It is considered key to these healthful qualities. However, EGCG used in clinical application which is still shortcomings of short half-life and low bioavailability. Chitosan (CS) has been widely used in pharmaceutical and medical areas, particularly for its potential in the development of controlled release drug delivery systems due to its well properties. In this study, we prepared EGCG-loaded chitosan nanoparticles by ionic polymeric method using sodium tripolyphosphate(TPP) as ionic polymeric agent successfully. Results controlled conditions (concentration of CS, 2 mg/mL; pH = 5.4; volume of TPP(0.5 mg/mL), 6.6 mL; amount of EGCG, 15 mg; ionic polymeric time, 24 h at room temperature (0.5 mL/min))volume of TPP(0.5 mg/mL), 6.6 mL; amount of EGCG, 15 mg; ionic polymeric time, 24 h at room temperature (0.5 mL/min)) for entrapment efficiency, loading efficiency, mean particle size and Zeta potential, were found to be 62.3 %, 33.8 %, 141.5 ± 0.4 nm and -31.21 ± 0.54 mV, respectively, and CS-EGCG-NPS have well property of sustained release.

Glycyrrhizic acid (GA), the main component from Radix glycyrrihizae, has a variety of pharmacological actions. In present study, GA nanoparticles suspension was prepared by supercritical anti-solvent (SAS) process and high-press homogenization process, and comparative studies were done on oral bioavailability, hepatic function and liver antioxidative activities in carbon tetrachloride–induced liver damage rats after treated with GA nanosuspension and unprocessed GA solution. Micronized CA under the optimum SAS conditions and CA nanosuspension prepared by high-pressure homogenization with the average particle size of about 80nm were obtained. GA nanoparticles suspension, compared with the unprocessed GA can significantly enhance the oral bioavailability. For carbon tetrachloride–induced liver damage rats, GA nanoparticles suspension can significantly reduce the release of ALTand AST. The plasma ALT and AST levels of GA supplementation were effective in decreasing up to 55 and 78.6 percent of model control group, respectively. For the GA solution group, the plasma ALT and AST levels were in decreasing up to 62.8 and 84.5 percent of model control group, respectively. GA nanosuspension could also significantly reduce plasma MDA and enhance SOD, GSH-Px and CAT activity. The CAT, SOD and GSH-Px activities increased by GA nanosuspension were about 1.31-fold, 1.43-fold and 1.28-fold of the unprocessed GA solution group, respectively. These results indicated that the GA nanoparticles suspension can be absorbed more easily than unprocessed GA after oral administration.

The aim of this study was to obtain nanoparticles and improve solubility of ursolic acid (UA), using supercritical antisolvent process (SAS). The four effects of process variables on drug particle formation during SAS process were investigated. Particles with mean particle size ranging from 139.4 ± 19.4 to 1039.8 ± 65.2 nm were obtained. The UA was characterized by scanning electron microscopy, X-ray diffraction (XRD), Fourier transform infrared spectroscopy, specific surface area and dissolution test. The XRD analyses showed that the processed UA was amorphous. The processed UA had solubility 4.4 times higher than the unprocessed UA.

Nanoscale lignin was successfully prepared with a supercritical antisolvent (SAS) apparatus using acetone as a solvent and superciritical carbon dioxide as an antisolvent. Four factors were studied and optimized by a four-level orthogonal array design (OAD). According to analysis of variance, precipitation pressure had a significant effect on mean particle size. The optimal conditions are as follows: precipitation temperature 35 °C, precipitation pressure 30 MPa, temperature difference +10 °C and concentration of lignin solution 0.5 mg/mL. The micronized lignin under the optimal conditions was characterized by Scanning Electron Microscopy (SEM), Fourier-transform Infrared Spectroscopy (FTIR), Dynamic Light Scattering (DLS) and X-Ray Diffraction (XRD) analyses. The results showed the mean particle size of micronized lignin was 0.144 ± 0.03 μm and had no degradation. The solubility of micronized lignin was improved significantly in distilled water.

Taxus chinensis var. mairei was used as test material, exogenous nitric oxide (NO) fumigation treatment with different concentration (0, 0.01, 0.1, 0.5 or 1 mM) of sodium nitrop russide (SNP), a donor for exogenous NO, was used to study the effects of NO on the active oxygen metabolism index of superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), peroxidase (POD), Hydrogen peroxide (H2O2) and malondialdehyde (MDA) in Taxus chinensis var. mairei. The results showed that low concentration of exogenous NO treatment increased the activities of the protective enzyme, such as SOD, CAT, POD and APX of Taxus chinensis var. mairei leaves, reduced the content of H2O2, and inhibit the increase of MDA in the first three days. NO fumigation treatment enhanced the antioxidant ability of Taxus chinensis var. mairei, prevented lipid membrane peroxidation.

In the study, the gas exchange parameters and chlorophyll contents in the needles of Pinus koraiensis Sieb. et Zucc. exposed to exogenous nitric oxide (NO) were determined. The Pinus koraiensis Sieb. et Zucc. seedlings were treated with sodium nitroprusside (SNP), a NO donor, at 5 different concentrations ranging from 0 mM to 1 mM. The result showed that at different concentrations exogenous NO had different effects on the photosynthetic parameters and photosynthetic pigments. The content of chlorophyll was maximal in the treatment with SNP at 0.01 mM. At low concentrations (0.1 mM) SNP significantly increased the photosynthetic rate (PN) and decreased the Respiration rate (RD) and Compensation irradiance (IC) of the needles (p

In this study, batch adsorption experiments were carried out for the biosorption of Cd2+ and Pb2+ from aqueous solutions by the invasive alien plant Iva Xanthifolia Nutt (IXN) biomass as adsorbent. Parameters effects such as contact time, pH, initial metal ions concentration, and biomass concentration on the biosorption process were studied. The Langmuir biosorption isotherm was applied to correlate the equilibrium data and fitted quite well. The maximum biosorption capacities for two metal ions were 30.35 mg Cd2+/g and 39.68 mg Pb2+/g, respectively. XRD studies combined with FTIR spectroscopy have also been investigated to represent the biosorption mechanism. Thermodynamic parameters such as enthalpy change (H0), entropy change (S0) and free energy change (G0) were calculated for predicting the nature of biosorption. This study proved that the invasive alien plant IXN can be used as an effective, low-cost and abundant absorbent for the treatment of Cd2+ and Pb2+ containing wastewater.

In this study, a new method (negative pressure cavitation method, NPC) for metal ions removal from industry waste water has been studied. Under the identical adsorption conditions，we have seen some different effects among static adsorption, shaking adsorption and NPC adsorption. The results are as following: four kind metal ions showed the accordant phenomenon that the sequence of adsorption capacity is NPC adsorption>shaking adsorption>static adsorption. The different metal ions have different improvement of removal ratio. The sequence is Cd>Hg>Pb>Cu. The intercellular pH of absorbed yeast cells increased with the adsorption process from acidic to alkaline. These may suggest that the Candida utilis has certain resistance against heavy metals. The increased pH is also an environment stress response of Candida utilis. So NPC adsorption may have potential values to removal of metal ions.

Lipid rafts are involved in the life cycle of many viruses. In this study, we investigated the role of lipids in the life cycle of vesicular stomatitis virus (VSV). Cholesterol depletion by pretreatment of BHK cells or VSV particles with methyl-beta-cyclodextrin (MbetaCD), a cholesterol-sequestering drug, inhibited the production of VSV dramatically. This effect was reversible, and virus production was restored by the addition of cholesterol, indicating that the reduction was caused by the loss of cholesterol in the cell membrane and virus, respectively. Cholesterol depletion at the adsorption stage also reduced the production of VSV significantly, but in contrast, only had a limited effect on virus production at the post-entry stage. Inhibition of sphingomyelin by myriocin treatment only showed a minor effect on VSV production. However, reduction of cholesterol and sphingomyelin at the same time dramatically reduced VSV production, showed a significant synergistic effect. These results suggest that lipid rafts play an important role in the life cycle of VSV.

The Dahurian larch forest in northeast China is important due to its vastness and location within a transitional zone from boreal to temperate and at the southern distribution edge of the vast Siberian larch forest. The continuous carbon fluxes were measured from May 2004 to April 2005 in the Dahurian larch forest in Northeast China using an eddy covariance method. The results showed that the ecosystem released carbon in the dormant season from mid-October 2004 to April 2005, while it assimilated CO2 from the atmosphere in the growing season from May to September 2004. The net carbon sequestration reached its peak of 112 g·m−2·month−1 in June 2004 (simplified expression of g (carbon)·m−2·month−1) and then gradually decreased. Annually, the larch forest was a carbon sink that sequestered carbon of 146 g·m−2·a−1 (simplified expression of g (carbon)·m−2·a−1) during the measurements. The photosynthetic process of the larch forest ecosystem was largely affected by the vapor pressure deficit (VPD) and temperature. Under humid conditions (VPD 1.0 kPa), the GEP decreased with the increasing VPD at a rate of 3.0 μmol·m−2·s−·kPa−1 and the ecosystem respiration was also enhanced simultaneously due to the increase of air temperature, which was linearly correlated with the VPD. As a result, the net ecosystem carbon sequestration rapidly decreased with the increasing VPD at a rate of 5.2 μmol·m−2·s−1·kPa−1. Under humid conditions (VPD < 1.0 kPa), both the GEP and NEP were obviously restricted by the low air temperature but were insensitive to the high temperature because the observed high temperature value comes within the category of the optimum range.

Vitexin, a naturally occurring flavone glycoside in plants, has many pharmacological effects, which is widely distributed in nature. This paper reviewed the research progress of the distribution of vitexin in the plant resources and its pharmacological effects, and summarized its application prospects, aiming to provide a useful reference for the development of vitexin-enriched plant resources.

Plantlet regeneration through shoot formation from young leaf explant-derived callus of Camptotheca acuminata is described. Calli were obtained by placing leaf explants on Woody plant medium (WPM) supplemented with various concentrations of 6-benzyladenine (BA) and naphthaleneacetic acid (NAA) or 2,4-dichlorophenoxyacetic acid (2,4-D). Callus induction was observed in all media evaluated. On the shoot induction medium, the callus induced on the WPM medium containing 19.8 μM BA and 5.8 μM NAA was the most effective, providing high shoot regeneration frequency (70.3 %) as well as the highest number of shoots (11.2 shoots explant−1). The good rooting percentage and root quality (98 %, 5.9 roots shoot−1) were achieved on WPM medium supplemented with 9.6 μM indole-3-butyric acid (IBA). 96 % of the in vitro rooted plantlets with well developed shoots and roots survived transfer to soil.

Rh. sachalinensis is a perennial,medicinal herb. The species is rare with apatchy distribution. Due to heavy disturbanceby human activities, it is now endangered.Allozyme analysis was employed to estimate itsgenetic diversity. Nineteen loci encoding 12enzyme systems were used. Rh.sachalinensis has a lower genetic variationwithin patches and higher differentiationbetween patches than other endangered, endemicspecies with limited distributions. The geneticdifferentiation between patches withingeographic regions is lower than that amonggeographic regions. Genetic drift is likely adominant factor contributing to the observedpattern of genetic structure. Because a higherdegree of genetic differentiation existsbetween patches in Rh. sachalinensis,more populations should be protected insitu, and samples from more populations shouldbe collected for ex situ conservation.

Starch-gel electrophoresis was used to examine the levels and distribution of genetic diversity in two Adenophora species: the narrow endangered Adenophora lobophylla and its widespread congener, A. potaninii. Based on allozyme variation at 18 putative loci, we measured high levels of genetic variability both in the endangered and the widespread species, with 83.3% of the loci being polymorphic. The mean expected heterozygosity within populations ( Hep ) and within species ( Hes ) were 0.234 and 0.244 for A. potaninii and were as high as 0.210 and 0.211 for A. lobophylla. There was higher differentiation among populations in A. potaninii ( FST = 0.155) than in A. lobophylla ( FST = 0.071). The high levels of genetic diversity in the present allozyme survey are consistent with the morphological variation observed in these species and may be attributed to high outcrossing rates in the Adenophora species. In addition, A. lobophylla was identified as a distinct species on the basis of Nei’s genetic distances and thus should be given a high priority for protection. It is noteworthy that the endangered A. lobophylla maintains much higher genetic diversity than most endemic or narrowly distributed plant species in spite of its restricted distribution. We hypothesize that A. lobophylla has become endangered for ecological and stochastic reasons, including habitat destruction or environmental changes, mud slides, and human disturbance such as grazing and mowing. Consequently, habitat protection is of particular importance for conserving this endangered species. Resumen: Se utilizo electroforesis con geles de almidon para examinar los niveles y distribucion de la diversidad genetica de dos especies de Adenophora: la amenazada Adenophora lobophylla y su congenere de amplia distribucion A. Potaninii. En base a la variacion de alozimas de 18 loci supuestos, medimos los altos niveles de variabilidad genetica tanto para la especie amenazada como para la especie ampliamente distribuida, con 83.3% de los loci siendo polimorficos. La heterocigocidad media esperada dentro de las poblaciones ( Hep ) y entre especies ( Hes ) fue de 0.234 y 0.244 para A. potaninii y fue tan alta como 0.210 y 0.211 para A. lobophylla. Hubo una diferenciacion mas alta entre poblaciones de A. potaninii ( FST = 0.155) que en A. lobophylla ( FST = 0.071). Los niveles altos de diversidad genetica en el presente estudio de alozimas son consistenes con la variacion morfologica observada para estas especies y podria atribuirse a altas tasas de cruza en las especies de Adenophora. Ademas, A. lobophylla fue identificada como una especie diferente en base a las distancias geneticas de Nei y esto deberia ser considerado como alta prioridad para su proteccion. Es notable que la especie amenazada A. lobophylla mantiene una diversidad genetica mucho mayor que la mayoria de las especies de plantas endemicas o con distribucion estrecha a pesar de su distribucion restringida. Hipotetizamos que A. lobophylla se ha convertido en amenazada por razones ecologicas y estocasticas incluyendo la destruccion del habitat o los cambios ambientales, los deslaves de tierra y las perturbaciones humanas como son el pastoreo y el corte. Consecuentemente, la proteccion del habitat es de particular importancia para la conservacion de esta especie amenazada.

Aims The aim was to isolate, identify and characterize endophytes from pigeon pea (Cajanus cajan [L.] Millsp.), as novel producer of cajanol and its in vitro cytotoxicity assay. Methods and Results Isolation, identification and characterization of novel endophytes producing cajanol from the roots of pigeon pea were investigated. The endophytes were identified as Hypocrea lixii by morphological and molecular methods. Cajanol produced by endophytes were quantified by liquid chromatography–tandem mass spectrometry (LC-MS/MS). R-18 produced the highest levels of cajanol (322·4 ± 10·6 μg l−1 or 102·8 ± 6·9 μg g−1 dry weight of mycelium) after incubation for 7 days. The cytotoxicity towards human lung carcinoma cells (A549) of fungal cajanol was investigated in vitro. Conclusions First, a novel endophyte Hypocrea lixii, producing anticancer agent cajanol, was isolated from the host pigeon pea (Cajanus cajan [L.] Millsp.). Fungal cajanol possessed stronger cytotoxicity activity towards A549 cells in time- and dose-dependent manners. Significance and Impact of the Study This endophyte is a potential handle for scientific and commercial exploitation, and it could provide a promising alterative approach for large-scale production of cajanol to satisfy new anticancer drug development.

The contents of 6 metal elements (Ca, Mg, K, Na, Cu and Fe ) were determined in serum, urine, feces and different tissue and organs (heart, kidney, stomach, liver, spleen, intestine and lung) of Wistar rats and mice by flame atomic absorption spectrometry (FAAS) with wet digestion. The samples were digested by the mixture of HNO3 and HClO4 (4 : 1, V/V) at 120 degrees C, the correlation coefficient for the standard curves was 0.999 4-0.999 8, the relative standard deviation (RSD) was from 0.33% to 4.52%, and the recovery for the samples was from 97.7% to 104.2%, meeting the demand of elements content determination in biological samples. The assay method for the determination of the 6 metal elements in animal samples established in this study has the advantages of easy operation, high sensitivity, less sample requiring and accurate results, and the method can be widely used in the determination of various metal elements in biological samples.

An ionic liquid-based surfactant combined with microwave-assisted extraction method, followed by RP-HPLC-diode array detection (DAD) with a core shell column, was successfully applied in extracting and quantifying four major phloroglucinols from Dryopteris fragrans. Eight ionic liquids with different cation and anion were investigated, and 1-octyl-3-methylimidazolium bromide presented the best relative extraction efficiency for four phloroglucinols. The optimum conditions of this method were as follows: ionic liquid concentration 0.75 M, liquid/solid ratio 12:1 mL/g, extraction time 7 min, extraction temperature 50°C, and irradiation power 600 W. The quality analytical parameters of the method were obtained based on the linearity, precision, accuracy, detection, and quantification limits. The recoveries were between 96.90 and 103.5% with standard deviations not higher than 4.7%. Compared with ionic liquid-based heat reflux extraction, ultrasonic-assisted extraction, negative-pressure cavitation extraction, and conventional microwave-assisted extraction, the relative extraction efficiencies of the proposed method for four phloroglucinols increased 1.5-40.4%. The method was successfully applied for the quantification of four major phloroglucinols from D. fragrans. All these results suggest that the developed method represents an excellent alternative for the extraction and quantification of phloroglucinols in other plant materials.

Corilagin is a member of polyphenolic tannins. Its antimicrobial activity and action mechanism against Escherichia coli, Staphylococcus aureus and Candida albicans were investigated through membrane permeability. Crystal violet staining determination, outer membrane (OM) and inner membrane (IM) permeability, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and atomic force microscopy (AFM) were used as methods for our investigation. The minimum inhibitory concentrations were 62.5, 31.25 and 62.5 µg/mL for E. coli, S. aureus and C. albicans, respectively. Crystal violet results and SDS-PAGE of supernatant proteins showed that corilagin dose-dependently affected membrane permeability of E. coli and C. albicans but not of S. aureus. OM and IM permeability assays revealed comparable results for E. coli. By using AFM, we demonstrated extensive cell surface alterations of corilagin-treated E. coli and C. albicans. SDS-PAGE of precipitated proteins revealed possible targets of corilagin, i.e. Fib, Sae R, Sar S in S. aureus and Tye 7p in C. albicans. In conclusion, corilagin inhibited the growth of E. coli and C. albicans by disrupting their membrane permeability and that of S. aureus by acting on Fib, Sae R and Sar S but not on membrane integrity.