Weak area shrinkage of glaciers by 2.5 ± 0.5 km2 (0.2 ± 0.1%) happened after 2000 and the period 2009–2014 even saw a slight expansion by 0.5 ± 0.1 km2 (0.1 ± 0.0%). The glaciers in this region have experienced an overall loss of –6.99 ± 0.80 km3 in ice volume or –0.15 ± 0.12 m water equivalent (w.e.) a–1 from 1971/76 to 2013/14. The mass budget of MAKT was –0.19 ± 0.19 m w.e. a−1 for the period ~1971/76–1999 and –0.14 ± 0.24 m w.e. a−1 during 1999–2013/2014. Similar to previous studies, there has been little mass change in the Pamir over recent decades despite such uncertainties. Glacier mass change showed spatial and temporal heterogeneity, with strong mass loss on debris-covered glaciers with an average of –0.32 ± 0.12 m w.e. a−1 from the 1970s to 2013/14.The assessment of glacier mass budget is crucial for assessing water reserves stored in glaciers. Derived glacier mass changes in the Muztag Ata and Kongur Tagh (MAKT) region in the eastern Pamir, northwestern China, is helpful in improving our knowledge of the dynamics of glaciers under a changing climate in High Mountain Asia. Here, glacier area and mass changes derived from remote sensing data are investigated for the period 1971/76–2013/14 for glaciers in MAKT. We have used ASTER images (2013/14), Cartosat-1 (2014) and Landsat, SRTM (Shuttle Radar Terrain Mission) digital elevation model (DEM) (2000), topographic maps (1971/76) and the first and second Chinese glacier inventories (CGIs). Our results indicated that the glacier area of MAKT decreased from 1018.3 ± 12.99 km Continued measurements based on additional observations above 5700 m a.s.l. showed less positive values of +0.05 m w.e. a−1 for 2010/2011 to 2013/2014 [−1 (1973–2009) to −0.01 ± 0.30 m w.e. a−1 (1973–2013), which revealed nearly balanced budgets at Muztag Ata in eastern Pamir for the last 40 years.Glaciers are sensitive to climate change and play an important role in the discharge of many Asian rivers . In past Moholdt  and a laBerthier  in the wBerthier  reported2 a–1 in Muztag Ata and Kongur Tagh (MAKT). Holzer, Vijay [Heterogeneous mass wastage has been found across the Pamirs, although the glacier area change was at almost the same level. In western Pamir (Tajikistan), a glacier area decrease of 18.4% was found in the Saukdara and Zulumart Ranges during 1978–2001 , increasr, Vijay  showed tr, Vijay , 18 indi2 in the eastern Pamir in Chinese territory [2 in the glacierized center of MAKT to understand the status of glaciers in the region with different additional data. The objectives of this study are summarized as follows:There are 1 265 glaciers with a total area of 2054.0 ± 75.9 kmerritory . Up to nerritory  mentioneTo update the glacier area change on decadal scale with new boundary data for glaciers in 2014, 2000, and 1971/76.To determine the mass change in different periods in MAKT using geodetic methods based on topographical maps (1971/76), SRTM DEM (2000), ASTER DEM (2013/14), and Cartosat-1 DEM (2014).To analyze the response of debris-covered glaciers in this region.Muztag Ata and Kongur Tagh (MAKT) are located in eastern Pamir, China , with altitudes ranging from 3000 m above sea level (a.s.l.) to maxima of 7546 and 7719 m above sea level (a.s.l.) at the two summits, respectively . Both maThirteen topographic maps with a 1:50,000 scale were obtained, which were constructed from aerial photogrammetry acquired in the 1970s by the Chinese Military Geodetic Service (CMGS) . The geohttp://earthexplorer.usgs.gov/). The SRTM 1 Arc-second global (approximately 30 meters) dataset was released in phases starting from September 24, 2014. The SRTM 1 data offer worldwide coverage of void-filled data at a resolution of 1 arc-second (30 meters). However, some tiles may still contain voids. The SRTM1 data in our study contains voids, especially at high and steep elevation regions due to radar shadows and layover effects, similar to the unfilled finished SRTM 3 data (90 m) [The SRTM data were obtained during the single-pass interferometric synthetic aperture radar SRTM mission during an 11-day mission in February 2000, in which DEMs can be seen as representative of the glacier surface at the end of the 1999 ablation period with slight seasonal variances , 33–37. a (90 m) ; the cooa (90 m) . Becausea (90 m) , 40. Thea (90 m) . A compaHowever, the penetration of the C-band radar waves needs to be taken into account for volume change as it varies from 0 to 10 m depending on the local climatic conditions , 39, 42.The ASTER sensor mounted on the Terra satellite platform provides multispectral imagery between 82° N and 82° S . There aCartosat-1 (IRS-P5) was launched by the Indian Space Research Organisation (ISRO) in 2005 and is equipped with two panchromatic cameras tilted at –5° (AFT) and +26° (FWD) from the pitch axis , 52. The™. The hillshade based on the SRTM3 DEM, TOPO DEMs and the calculated ASTER and Cartosat-1 DEMs provided additional information for detection of the glacier boundary. We estimated the uncertainty using a buffer of 6 m for topographic maps with a scale of 1:50 000 (cf. [EAC) was calculated using Eq  datasets , 59 prov000 cf. , 61) and and™. Th000  DEMs , 66. TheMED) of the nonglacier area between the final adjusted DEMs was in the range of –0.99 to 0.34 m (STDVno glac) would probably overestimate the uncertainty due to averaging over larger regions, which reduces the errors [SE) used for estimation of the uncertainty is given byN is the number of included pixels. To minimize the influence of autocorrelation, it is recommended that the de-correlation length be chosen based on the spatial resolution. For DEMs with a spatial resolution of 5 m, a de-correlation length of 100 m is suitable [σ) was determined using the SE and MED over the ice-free terrain [After the adjustments, we calculated statistical parameters for the differences of the nonglacier area between the final adjusted DEMs . The meao 0.34 m . The stae errors , 69. Thusuitable . Berthiesuitable  used val terrain :σ=MED2+The accuracies for all subregions were calculated for the investigated periods. An accuracy ranging from 0.65 to 1.26 m  indicateε) is defined as:n is the number of years for a period.Considering the law of error propagation from one year to the next, the annual mean uncertainty .Mass change was calculated based on the glacier surface elevation change or volume change using a density assumption or model. Kääb, Berthier  proposed3. Huss  suggeste2 in 1971/76; however, heterogeneous variations were observed for different periods and different glaciers  and even increased during 2009–2014   were relatively stable (only decreased by 0.4 ± 0.1%) in the past 40 years. The area change was highly variable between different glaciers, even for those adjacent to each other. Most glaciers shrank slightly or were stable after 2000; however, Kuokuosele Glacier (No. 1) and Kuosikulake Glacier (No. 3) decreased by ~0.3%–1.5% (~0.01% a–1–0.06% a–1) from 1976 to 2000 and then subsequently advanced (~1% or ~0.04% a–1). Glacier G075087E38696N (No. 18) kept advancing throughout the period of investigation and showed rapid change trends, i.e., +26.5 ± 0.6 m a–1 from 1976–2000, followed by +43.1 ± 1.6 m a–1 from 2000 to 2009, and +79.4 ± 3.0 m a–1 from 2009 to 2014 (enlargement of +5.3 ± 1.1%). The three largest glaciers  are heavily covered by debris and were relatively stable from 1976 to 2014  . Fifty-e to 2014 .3 in ice volume or –0.15 ± 0.12 m w.e. a–1 from 1971/76 to 2013/14. The overall mass budgets of glaciers in Muztag Ata and Kongur Tagh were –0.15 ± 0.18 and –0.21 ± 0.19 m w.e. a−1, respectively, for ~1971/76–1999; –0.13 ± 0.23 and –0.16 ± 0.25 m w.e. a−1 during 1999–2013/2014  during 1976–2014. The mass changes varied from glacier to glacier in different periods. Glacier G075247E38305N (No. 10) showed a negative mass change (–0.47 ± 0.18 m w.e. a–1) before 1999 and a nearly balanced state (0.01 ± 0.23 m w.e. a–1) after 1999. Karayaylak Glacier (G075254E38623N), the largest glacier of Muztag Ata and Kongur Tagh, exhibited a mass loss (–0.32 ± 0.19 m w.e. a−1) during 1976–1999; however, the mass budget of Karayaylak Glacier was positive (0.37 ± 0.25 m w.e. a−1) after 1999.Glaciers in MAKT exhibited heterogeneous mass changes during 1970s–2014. Some glaciers showed stable or little change in their area, but down-wasting at the same time. For example, Koksay Glacier (No. 5), the largest glacier of Muztag Ata massif, thinned by 9.96 ± 0.65 m  exhibited a decrease of only ~0.1% (0.3 km2) from 1976 to 2014, and the total glacier area in Muztag Ata and Kongur Tagh increased by ~0.02% (0.18 km2) from 2009. Overall, large glaciers retreat slower than small ones [Muztag Ata and Kongur Tagh showed the least glacier shrinkage (~–1.9% during 1971/76–2014) compared with the other regions in the eastern Pamir [all ones , which iall ones .2 in our study from 54.5 ± 0.11 km2 in Holzer, Vijay [A consensus of glacier retreat measured in their area was commonly found in previous studies for glaciers in MAKT, but with various rates from –0.6% to –7.9% in different periods , 15, 16.–1) during the 1970s–1990 and 0.2 ± 2.5% (0.01 ± 0.32% a–1) during 1990–2010s. No significant area changes were reported for Karakoram [–1) (1975–2008) for central Tian Shan [–1 in the Ili River basin [Glaciers in the region might be weak retreating ones if compared with those in other mountain ranges in High Mountain Asia , 73. Thearakoram –76. The ian Shan  is also ian Shan  and westian Shan , and coner basin .−1 during 1973–2013 or –0.03 ± 0.33 m w.e. a−1 during 1973–2009) despite such uncertainties. Most results for glaciers in Muztag Ata might vary from Holzer and others. For example, the Koksay Glacier mass loss value (–0.22 ± 0.10 m w.e. a−1 during 1976–2014) might be higher than Holzer and others (–0.08 ± 0.30 m w.e. a−1 during 1973–2013 or –0.11 ± 0.33 m w.e. a−1 during 1973–2009). We consider that accuracies of different DEMs and GCPs may be the main reason for these deviations. The accumulation regions showed high noise from optical data with a high uncertainty for elevation, which was also subject to such eventual biases. However, the trends in glacier mass change in the past 40 years were consistent between us and Holzer and others. The trend biases after 1999 may contribute to uncertain penetration of SRTMs C-band radar into ice and snow, which also may explain why there are discrepancies with Gardner, Moholdt [−1), Kääb, Treichler [−1), and Gardelle, Berthier [−1) for glaciers’ mass change in Pamir after 1999 despite different study extents.By integration of our result with that from Holzer, Vijay , we conc−1 [−1) [Early glacier surveys in the Muztag Ata verify that glacier ablation was intensified by atmospheric warming from 1960/87 to 2001 on Kematulejia Glacier about 4460–4600 m a.s.l. in the summer season . Hydrolo−1 , which i−1 [−1) . Thus, u−1 [−1) . However−1 [−1) .−1 in our study area from the 1970s to 2013/14, although debris-covered area change was not significant. Early glacier surveys on Kematulejia Glacier about 4460–4600 m a.s.l. verify that glacier ablation on debris-covered ice was greater than on clean ice [One of the notable results is that the debris-covered regions obviously exhibit higher thinning rates with an average of –0.32 ± 0.12 m w.e. alean ice . The maslean ice . The higlean ice .2 from 1963 to 2014, which accounts for 3.8 ± 0.6% of its area in 1963. Muztag Ata and Kongur Tagh were reduced by an area of –30.0 ± 4.7 km2 (–2.7 ± 0.5%) between 1971/76 and 2014. The glacier shrank slightly after 2000  and increased during the period 2009–2014 . The mass budget of MAKT was –0.19 ± 0.19 m w.e. a−1 for the period ~1971/76–1999 and –0.14 ± 0.24 m w.e. a−1 during 1999–2013/2014. Overall, Muztag Ata and Kongur Tagh lost –7.07 ± 0.80 km3 in ice volume  from 1971/76 to 2013/14. These losses are heterogeneous and differ spatially as well as temporally. Although debris-covered area change was not significant, the debris-covered regions obviously exhibit higher thinning rates, with an average of –0.32 ± 0.12 w.e. a−1 from the 1970s to 2013/14.We used remote sensing datasets of ASTER images (2013/14), Cartosat-1 (2014), and Landsat in conjunction with SRTM data (2000), topographic maps (1971/76), and the first and second Chinese glacier inventories to investigate glacier area and mass changes of Muztag Ata and Kongur Tagh. Our results show that the overall area loss of Muztag Ata and Kongur Tagh was –39.4 ± 5.8 km


C. gracilis were studied for the first time under submerged culture conditions. Effect of different factors on polysaccharides production was studied by orthogonal experiments using one-factor-at-a-time method. Incubation of culture in the medium with capacity 200 mL (675.12 ± 5.01 and 385.20 ± 5.01 mg/L), rotation speed 150 rpm (324.62 ± 3.32 and 254.62 ± 4.62 mg/L), 6-day culture incubation time , pH 6.0 (374.81 ± 2.52 and 219.45 ± 2.59 mg/L), and temperature 23°C (405.24 ± 1.11 and 215.60 ± 1.71 mg/L) produced higher EPS and IPS, respectively. Maximum EPS and IPS production was observed in the medium supplemented with glucose as a carbon source (464.82 ± 2.12 and 264.42 ± 2.62 mg/L) and yeast extract as a nitrogen source (465.21 ± 3.11 and 245.17 ± 3.24 mg/L), respectively. Carbon to nitrogen ratio for maximum EPS and IPS production was observed as 10 : 1 (395.29 ± 2.15 and 235.62 ± 1.40 mg/L), respectively. Glucose was found to be the major monosaccharide (62.15 ± 7.33%). Both EPS and IPS of C. gracilis showed significant DPPH radical scavenging activity, ABTS radical scavenging activity, reducing power, and iron chelating activity.Under present experiments, EPS and IPS production, monosaccharide composition, and antioxidant activities of Cordyceps Fr. is a widespread genus with 400 species worldwide , cyclo-[Leu-Pro], cyclo-[Val-Pro], cyclo-[Ala-Leu], cyclo-[Ala-Val], and cyclo-[Thr-Leu] and small amounts of polyamines, such as 1,3-diamino propane, cadaverine, spermidine, spermine, and putrescine  × 100, where Ab is the absorbance of the blank and As is the absorbance in the presence of the extract [x-axis and chelating activity on the y-axis. EC50 values were calculated using the following equation:x0log is the center of the curve.For this, 1 mL of the sample (2–10 mg/mL) was mixed with 3.7 mL of ultrapure water, following which the mixture was reacted with ferrous chloride  and ferrozine  for 20 min. And the absorbance was read at 562 nm. EDTA was used as positive control. The chelating activity on the ferrous ion was calculated using the following formula: chelating activity (%) = [( extract . Chelati9 (34) was used for optimal culture conditions in submerged cultures.Different factors considered for the orthogonal design were carbon sources, nitrogen sources, and ratio of carbon to nitrogen sources, temperature, pH value, medium capacities, rotation speed, and culture time. According to the results of the single factor experiment, the orthogonal LP < 0.05 was considered significant, and SPSS software  was used to calculate differences. All experiments were performed at least 3 times and with 3 replicates. The results were analyzed using one-way analysis of variance (ANOVA). Tests of significant differences were determined by Tukey-HSD at P < 0.05.All results are expressed as mean ± standard error.  Cordyceps gracilis was grown in flasks (500 mL) containing liquid media with different capacities. The maximum EPS (675.12 ± 4.01 mg/L) and IPS (385.20 ± 3.00 mg/L) production was observed in 200 mL of the medium, while the least values for EPS (302.17 ± 4.46 mg/L) and IPS (122.13 ± 5.46 mg/L) production were obtained in 50 mL of the medium. However, no significant difference P < 0.05 was observed in EPS and IPS production in the medium with 100–150 mL capacities. Variation in rotation speed showed a direct relation with EPS and IPS production. Results obtained for the effect of the rotation speed on polysaccharide production showed maximum EPS (324.62 ± 3.32 mg/L) and IPS (254.62 ± 4.62 mg/L) production in culture medium with rotation speed 150 rpm. Similar results were obtained for medicinal fungus C. ophioglossoides (Ehrh.) Link and other ascomycetes. The reason for this is low oxygen requirement by the culture at 150 rpm rotation [ C. ophioglossoides [ C. gracilis incubated for 6 days and pH 6.0 showed maximum EPS and IPS production  and IPS (215.60 ± 1.71 mg/L) production at 23°C. This temperature for C. sinensis was observed as 20°C and, for C. ophioglossoides, it was found to be 25°C [To find the suitable medium capacity for maximum EPS and IPS production,rotation , 37. Theoduction . In the  be 25°C . Cordyceps gracilis. Although all the tested carbon sources yielded EPS and IPS, maximum EPS (464.82 ± 2.12 mg/L) and IPS (264.42 ± 2.62 mg/L) production took place in the medium supplemented with glucose as carbon source. Glucose was also found to be the most favorable carbon source for polysaccharide production in many of the Cordyceps species [ C. gracilis  production [Six different carbon sources were studied to find the suitable medium source for the production of EPS and IPS in species , 48, 49. species . C/N ratgracilis . Presentoduction . C. militaris (L.) Link and other medicinal basidiomycetes, in which glucose was found to be the major monosaccharide along with sucrose and galactose [Monosaccharide composition of polysaccharides showed glucose in major percentage (62.15 ± 7.33%) followed by rhamnose (36.81 ± 4.57%) and galactose in minor quantities (0.10 ± 0.0%) . Similaralactose , 50.Results obtained for influence of different factors on yield of EPS and IPS showed a significant positive effect. Results revealed the effect on EPS and IPS production in the order temperature > incubation time > pH > rotation speed > medium capacity . C. gracilis showed positive and direct correlation with the concentration of the sample  and IPS (58%). However, at higher sample concentrations, EPS showed higher iron chelating ability than IPS (50 values ( C. gracilis have shown significant antioxidant activities as reported in C. sinensis [ C. militaris [ C. gracilis [The results obtained for reducing power abilities of EPS and IPS in submerged culture ofncreased . Howeverthan IPS . Both EP0 values . EPS andsinensis  and C. militaris . Since agracilis , 56. Cordyceps gracilis is a medicinal fungus and it contained both EPS and IPS in appreciable amount. In vitro evaluation of antioxidant activities of EPS and IPS showed significant higher antioxidant activities. However, submerged cultivation of this fungus required several factors for the production of EPS and IPS. Factors such as temperature, rotation speed, pH, incubation time, carbon, nitrogen, and carbon to nitrogen ratio showed significant effect on the production of EPS and IPS. Due to significantly higher DPPH radical scavenging activity, ABTS radical scavenging activity, reducing power, and iron chelating abilities, this fungus is useful for its pharmaceutical applications. Present findings will open the scope for its large-scale industrial fermentations for commercial uses like other commercially explored Cordyceps species.Results obtained under present investigations showed that


Such performance is comparable to those of the best-performing nickel–iron-layered double hydroxides for water oxidation in alkaline media. Mechanistic studies indicate that the nickel–vanadium-layered double hydroxides can provide high intrinsic catalytic activity, mainly due to enhanced conductivity, facile electron transfer and abundant active sites. This work may expand the scope of cost-effective electrocatalysts for water splitting.Highly active and low-cost electrocatalysts for water oxidation are required due to the demands on sustainable solar fuels; however, developing highly efficient catalysts to meet industrial requirements remains a challenge. Herein, we report a monolayer of nickel–vanadium-layered double hydroxide that shows a current density of 27 mA cm Water oxidation is a key component of water splitting, but more effective catalysts for the process are still required. Here the authors synthesize a monolayer nickel-vanadium layered double hydroxide material showing high activity for water oxidation owing to enhanced conductivity and abundant active sites. Although some first-row transition metal oxides  had been developed as low-cost electrocatalysts for water oxidation, most of them still cannot compete with IrO2 and RuO21345668102 in alkaline media68Water splitting is considered one of the most promising strategies to produce chemical fuels such as hydrogen. The half reaction of the water splitting process, water oxidation, remains the bottleneck of the whole process at present. Therefore, developing highly efficient water oxidation catalysts is crucial. Some precious metal-based electrocatalysts, such as IrO2 to explore novel LDHs for water oxidation, for example, recently Koper and co-workersIt is already known that Fe(III) incorporated in Ni(II)-based LDH is the key aspect for the high catalytic performance, although the role of Fe in LDH is still ambiguous561314152: vanadium, and succeed in forming NiV-LDH as an efficient catalyst for the water oxidation reaction. A simple one-step hydrothermal method is employed to synthesize NiV-LDH. Without need for exfoliation or hybridization with other materials, the resulting monolayer NiV-LDH catalyst exhibits comparable activity to the best-performing NiFe-LDH for water oxidation in alkaline electrolyte.In this work, we incorporate another earth-abundant element into Ni(OH)2 and Ni0.75V0.25-LDH. As can be seen, pure Ni(OH)2 was successfully synthesized in the simple hydrothermal system without addition of V sources, and the X-ray diffraction pattern supports the formation of pure hexagonal α-Ni(OH)2 (JCPDS 380715), which exhibits a layered structure constructed from [NiO6] coordinated octahedra connected by sharing their edges. After incorporation of V into the structure of α-Ni(OH)2, no obvious change can be observed in the X-ray diffraction spectrum, indicating Ni0.75V0.25-LDH is the isomorphous compound as α-Ni(OH)2 with the layered structure. The X-ray diffraction pattern of prepared bare V-based hydroxide without Ni source  core-level spectra of the Ni0.75V0.25-LDH powder. The Ni 2p spectrum presents two main structures, resulting from the spin–orbit splitting of the p orbital that are assigned as Ni 2p3/2 (850−870 eV region) and Ni 2p1/2 (870−890 eV region). The binding energy difference between the 3/2 and 1/2 components is ∼17.4 eV. Ni 2p3/2 presents a main peak at ∼855.4 eV with an intense satellite structure at ∼861.2 eV, , this signature is characteristic of Ni2+  glass substrate used with an O1s-binding energy ∼530 eV. The V 2p core-level spectrum is also decomposed into V 2p3/2 and V 2p1/2 due to the spin–orbit splitting that are separated by ∼7.5 eV. A closer view on the V 2p3/2 region is shown in ∼515.4 eV (in blue), ∼516.3 eV (in light grey) and ∼517.5 eV (in grey) and are in good agreement with respectively V3+, V4+ and V5+ 2 and Ni0.75V0.25-LDH are shown in α-Ni(OH)2 shows a narrow nanosheet morphology with the size of few tens of nanometre. Aggregated by these nanosheet crystals, a porous sphere structure with the diameter of few micrometres was obtained, as observed by scanning electron microscopy  and height profile of Ni0.75V0.25-LDH in 0.75Fe0.25-LDH was also prepared by a similar protocol in the literature0.75Fe0.25-LDH can be observed, showing that the size of aggregations by nanosheets in Ni0.75Fe0.25-LDH is bigger than Ni0.75V0.25-LDH. However, the TEM, AFM images and height profile of Ni0.75Fe0.25-LDH in 0.75Fe0.25-LDH is also ultrathin with thickness of ∼1.2 nm. Meanwhile, differing from the above nanosheet-based structure, the bare V-based and Fe-based hydroxides (β-FeOOH) without Ni content show mainly ‘nanostick' morphologies after the hydrothermal processes  images of opy SEM; . After arocesses .−2. As shown in 2, bare V-based and Fe-based hydroxides are very low, so they are not shown here for clarity). When the molar ratio of Ni/X (X=V or Fe) is 3:1, that is, Ni0.75V0.25-LDH and Ni0.75Fe0.25-LDH, the best water oxidation performances are achieved with the highest catalytic current density  curves, but also reflected in its lower Tafel slope . Tafel slopes can be influenced by mass transport and electron transport26−1 for LSV have been done, which resulted in negligible change of current densities for Ni0.75Fe0.25-LDH and Ni0.75V0.25-LDH catalysts, indicating sufficiently fast mass transport for both LDHs260.75V0.25-LDH is likely ascribed to the facile electron transport through the layers of the catalyst.It is known that the composition can affect the water oxidation performance of the catalysts significantly. First, we investigated the electrocatalytic activities of NiV-LDHs and NiFe-LDHs on glassy carbon (GC) electrodes with different Ni content to optimize the composition (the measurements were carried out without ohmic-drop correction unless noted otherwise). All the amount of catalyst loadings on GC electrodes were 0.143 mg cm density  and turn density , and the density  in NiV-L density . Either opy EDS; , showingoducible  and compel slope . Ni0.75Vη of 350 mV, Ni0.75V0.25-LDH can achieve current density of 27.0±1.6 mA cm−2, which is more than twice higher than the one of Ni0.75Fe0.25-LDH (11.7±1.5 mA cm−2). TOF was also calculated at an overpotential of 350 mV in 1 M KOH, assuming all the metal sites were electrochemically active in the water oxidation reaction. Ni0.75V0.25-LDH also shows the better TOF of 0.054±0.003 s−1 compare with 0.021±0.003 s−1 for Ni0.75Fe0.25-LDH. It is worth noting that the above TOFs in this study are obviously underestimated, since some of the metal sites are electrochemically non-accessible. Meanwhile, the overpotential η required to achieve 10 mA cm−2 current density, which is approximately the current expected at the anode in a 10% efficient solar water splitting device under 1-sun illumination0.75Fe0.25-LDH and Ni0.75V0.25-LDH show required overpotential 0.347±0.002 and 0.318±0.003 V, respectively, exhibiting the lower required overpotential of Ni0.75V0.25-LDH.At 0.75Fe0.25-LDH and Ni0.75V0.25-LDH in 1 M KOH  of Ni0.75V0.25-LDH and Ni0.75Fe0.25-LDH were obtained from cyclic voltammetry (CV) curves in 1 M KOH and compared. 0.75V0.25-LDH with different scan rates. By plotting the ΔJ=(Ja–Jc) at 0.25 V versus Ag/AgCl against the scan rate, the linear slope that is twice the double layer capacitance (Cdl) can be obtained, and is normally used to represent the corresponding ECSA1911To understand the reason behind the high catalytic performance of Ni0.75Fe0.25-LDH is 0.199±0.018 mF cm−2, while Ni0.75V0.25-LDH has a higher linear slope of 0.270±0.030 mF cm−2, which means Ni0.75V0.25-LDH has a more electroactive surface than Ni0.75Fe0.25-LDH  of Ni0.25-LDH . The big0.25V0.75-, Ni0.5V0.5-, Ni0.75V0.25- and Ni0.83V0.17-LDH electrodes were 0.123±0.009, 0.166±0.012, 0.270±0.030 and 0.144±0.015 mF cm−2, respectively, in −2, while the corresponding current density increases from ∼4.0 to 27.0 mA cm−2, showing significantly almost seven times improvement. This is a strong clue that incorporating V into Ni hydroxide can not only change the active surface area, but also enhance the intrinsic catalytic property. As a comparison, we also investigated current density-dependent ECSA of different NiFe-LDHs in −2) with 1.3-fold increase of ECSA (from ∼0.157 to 0.199 mF cm−2). This linear correlation indeed indicates that the improvement of catalytic activity of NiFe-LDHs is mainly due to the increase of actively accessible surface area. We attempted to make a comparison of the catalytic activities between NiFe-LDH and NiV-LDH with the same ECSA value. The linear correlation in −2 of ECSA  is still lower than Ni0.75V0.25-LDH (∼27.0 mA cm−2), although they have been estimated to have the same ECSA value. These results suggest in addition to the increased accessible surface area, probably the V in the Ni-based LDHs can also lead to the increased intrinsic catalytic activity than NiFe-LDHs in our case.To verify this point, the ECSAs of NiV-LDH with different Ni content were further examined in details, as shown in 0.75Fe0.25-LDH and Ni0.75V0.25-LDH in 0.75V0.25-LDH (∼50 mV dec−1) than that of Ni0.75Fe0.25-LDH (∼64 mV dec−1) indicates the higher catalytic activity and superior type of the active sites in Ni0.75V0.25-LDH. In addition, Ni0.75V0.25-LDH also shows lower onset potential  than that of Ni0.75Fe0.25-LDH , implying the facile kinetics of Ni0.75V0.25-LDH for the water oxidation reaction, which is consistent with the Tafel slope investigation. These results indicate that Ni0.75V0.25-LDH possesses superior intrinsic catalytic activity in our case.Although it is difficult to correctly normalize the catalytic current to the number of active sites in the LDH electrocatalysts to compare the intrinsic catalytic performances accurately, the kinetic parameters of the Tafel slope and onset potential can be selected to reflect the intrinsic activity of catalysts300.75V0.25-LDH may be due to the higher conductivity than Ni0.75Fe0.25-LDH. To test the conductivity, electrochemical impedance spectroscopies (EIS) of Ni0.75Fe0.25-LDH and Ni0.75V0.25-LDH electrodes were carried out in the three-electrode configuration in 1 M KOH. The Nyquist diagrams of both electrodes show an apparent semicircle in the high frequency range (Rct) in the LDH catalysts320.75V0.25-LDH decreases comparing to that of Ni0.75Fe0.25-LDH, indicating lower charge transfer resistance, that is, improved conductivity in LDH catalysts. A reported electrical analogue was used to fit the EIS dataRct of Ni0.75V0.25-LDH is estimated to be ∼62 Ω cm2, lower than that of Ni0.75Fe0.25-LDH (∼94 Ω cm2). This is also reflected in the Bode plots in 0.75V0.25-LDH shows smaller resistance than Ni0.75Fe0.25-LDH. It is known that the LDHs suffer from their low conductivity, in the current case, the higher electron conductivity of Ni0.75V0.25-LDH benefits the charge transfer efficiently, which is also in a good agreement with the lower Tafel slope in The higher intrinsic catalytic activity of Nicy range , which s0.75Fe0.25-LDH, the slightly more three-dimensional structured Ni0.75V0.25-LDH can expose more edges of the nanosheets to the electrolyte , but after that the trend is much less pronounced. This change of trend of the activity dependent on ECSA for NiV-LDHs indicates that there should be other factors that can influence the activity of NiV-LDHs besides ECSA. It was reported that the catalytic performance of the layered catalyst for water splitting more strongly depends on the edge-state length than ECSA. The catalytic activity can be linear to the edge-state lengthOn the other hand, comparing to the relatively planar structure in Ni0.75V0.25-LDH, density functional theory (DFT) calculations were performed based on the following mechanism for water oxidation:To further understand the advantages of NiG1–ΔG5, ΔG5 is defined as 4.92 eV–ΔG1–ΔG2–ΔG3–ΔG4 to avoid the calculation of the energy of O2 molecule. The overpotential η is defined in Here ‘*' presents the adsorption site that is usually on the top of the doping element. The calculation of the reaction free energy with the zero-point energy and entropy corrections followed the same procedure in ref. G3+ΔG4)35G3) or formation of *OOH from *O (ΔG4). The calculated free-energy landscape shows that the rate-limiting step is the formation of *OOH when the sample is doped with V with η=0.62 V, whereas the rate-limiting step becomes the formation of *O when the sample is doped with Fe with η=1.28 V. Because ΔG3+ΔG4=3.22 eV, the lower limit of the η can be reached when ΔG3=ΔG4=1/2 × 3.22 eV=1.61 eV if a proper element was doped. Then η=1.61–1.23=0.38 V, which suggests it has a large space to further optimize the activity of Ni-based LDH by doping with other elements.The optimized structures of the intermediates in the free-energy landscape are shown in 0.75V0.25-LDH shows superior catalytic performance than other reported LDHs as well, to the best of our knowledge. 0.75V0.25-LDH with recently reported state-of-the-art LDH catalysts without exfoliation or coupling with other materials, which were loaded on GC electrodes for water splitting, including NiFe-, NiCo-, ZnCo-, CoCo- and MnCo-LDHs. Although the catalytic performances of LDH electrodes are strongly dependent on the preparation methods in literatures, to make a general overview, the current density at 350 mV overpotential and the corresponding mass activity (A g−1) are still used to compare the intrinsic activities between different LDH catalysts. To make the comparison more reasonable, ohmic-drop correction was also performed for Ni0.75V0.25-LDH, as most of the literature has done  before long-term anodic conditioning that was reported as one of the most active LDH catalyst for water oxidationBesides being better than the prepared NiFe-LDH in our case, Nihas done 89262938.0.75V0.25-LDH outperforms Ni0.75Fe0.25-LDH in our case, we cannot claim Ni0.75V0.25-LDH has higher catalytic activity than all the NiFe-LDHs in the literature. For instance, Louie and Bell reported NiFe-LDH that requires <300 mV overpotential to deliver current densities of 10 mA cm−2 with a Tafel slope of ∼40 mV dec−1 (with ohmic-drop correction), which outperforms our Ni0.75V0.25-LDH probably due to the different preparation method0.75V0.25-LDH is still highly efficient among the listed catalysts, and comparable to the best-performing NiFe-LDHs0.75V0.25-LDH is one of the best water oxidation catalysts.It is worth pointing out that although Ni2 evolution and Faradaic efficiency of Ni0.75V0.25-LDH electrode, the experimental and theoretical O2 evolution amount by Ni0.75V0.25-LDH at a constant oxidative current of 1 mA in 1 M KOH were performed as shown in 2 evolution determined by gas chromatography exhibited 101±7% Faradaic efficiency, when the electrolysis time was 60 min. Moreover, to further explore the highly catalytic activity of Ni0.75V0.25-LDH, we drop-casted Ni0.75V0.25-LDH suspension onto Ni foam as well, which can supply excellent conductivity and very high surface area for the electrode, and tested the catalytic activity of resulted electrode in 1 M KOH for water oxidation. The Ni0.75V0.25-LDH on Ni foam exhibits the promising activity for practical applications, with ∼44 mA cm−2 at 350 mV overpotential, and just ∼300 mV overpotential required to reach 10 mA cm−2  was obtained by mixing NiCl2 and VCl3 in 80 ml H2O, while the total amount of metal ions (Ni2++V3+) was kept to 3.2 mmol. Afterwards, 0.3 g of urea was added, and the above mixture solution was transferred to a stainless-steel Teflon-lined autoclave, and heated in an oven at 120 °C for 12 h. After cooling the autoclave to room temperature, the resulting powder was washed by deionized water and ethanol three times, collected and then dried at 70 °C overnight. As a reference, for preparation of NiFe-LDH, the same processes were followed except FeCl3 was used as iron source instead of VCl3, which was analogous to the literatureA one-step hydrothermal method was employed to synthesize the bulk LDHs. In brief, for preparation of NiV-LDH, different mole ratios of Ni/V solution 2O, 0.25 ml 2-propanol and 10 μl 5% Nafion (ethanol solution) by sonication for >1 h. A measure of 2.5 μl of the above suspension were drop-casted to a pre-polished GC electrode (diameter: 3 mm), and dried at 50 °C for 15 min to evaporate the solvent.A measure of 5 mg of the obtained LDH powders were dispersed in the mixture solution of 1 ml Hα radiation. SEM images were taken on JEOL JSM-7000F instrument with energy-dispersive X-ray spectroscopy (EDS). TEM images were taken on JEOL JEM2100 TEM. Tapping mode AFM was performed to examine the surface morphology by a Dimension Icon AFM  on silicon wafer. Rectangular cantilevers with approximate dimensions of 125 μm in length and 40 μm in width (BudgetSensors Tap300Al-G) were used to perform the tapping mode experiments. The NanoScope Analysis software  was used to analyse the recorded AFM data. A first-order polynomial-flattening algorithm was employed to remove surface tilt from height images. XPS measurements were conducted with an in-house spectrometer (PHI 5500) using monochromatized Al Kα radiation . The pressure in the analysis chamber was ∼5 × 10−9 mbar during the measurement. Core peaks were analysed using a nonlinear Shirley-type background. The peak positions and areas were optimized by a weighted least-squares fitting method using 70% Gaussian/30% Lorentzian lineshapes. The powder was grinded and deposited on a conductive FTO glass. An electron flood gun was used to compensate the charging effects. The XPS spectra were energy calibrated by setting the adventitious carbon peak to 285 eV.X-ray diffraction measurements were carried out on Bruker X-ray diffraction diffractometer D5000 with Cu KERHE=EAg/AgCl+0.059 pH+0.197 V, and overpotential η=ERHE–1.23 V.Electrolysis experiments were carried out in a polytetrafluoroethylene cell with an Autolab potentiostat with GPES electrochemical interface (Eco Chemie) in a standard three-electrode configuration, which was composed of working electrode (LDHs deposited on GC electrodes), counter electrode (Pt net) and reference electrode (Ag/AgCl). The electrolyte was 1 M KOH, and the applied potentials were converted with respect to reversible hydrogen electrode (RHE), −2 was performed until a stable potential was obtained. Then LSV were measured from 0.2 to 0.7 V versus Ag/AgCl with a slow scan rate of 2 mV s−1. By plotting overpotential η against log (J) from LSV curves, Tafel slopes can be obtained. To test the stability of LDHs, a galvanostatic measurement at a fixed current density (J) of 10 mA cm−2 was performed. ECSA were measured by CV at the potential window 0.2–0.3 V versus Ag/AgCl, with different scan rates of 20, 40, 60, 80, 100 and 120 mV s−1. By plotting the ΔJ=(Ja−Jc) at 0.25 V versus Ag/AgCl against the scan rate, the linear slope that is twice of the double layer capacitance (Cdl) is used to represent ECSA. All the above measurements were carried out without ohmic-drop correction unless noted otherwise.First, before all the electrochemical measurements, a galvanostatic measurement at a fixed current density of 5 mA cmThe TOF of LDH catalysts were calculated according to the following equation (2)J is the current density at a given overpotential, for example, in our cases η=350 mV, A is the surface area of the electrode (0.07 cm2), F is Faraday constant  and m is the number of moles of the metal on the electrodes. In our cases, we assumed all the metal sites were actively involved in the electrochemical reaction.where 5 Hz, with an a.c. amplitude of 10 mV and overpotential bias of 350 mV.To compare the conductivities of LDHs, EISs were carried out in 1 M KOH with a three-electrode configuration, the frequency ranged from 0.1 to 102 were performed in an air-tight H shape cell, which was divided by a glass frit to two chambers. The working electrode, the Ag/AgCl reference electrode and a magnetic stirring bar were inserted in one chamber of the cell, the Pt counter electrode was inserted in the other chamber. The cell was filled with 1 M KOH and degassed with helium for >1 h. The headspace of the compartment containing the working electrode was 23.6 ml. The electrolysis was carried out with a constant oxidation current of 1 mA for 60 min. A measure of 500 μl of the gas sample in the compartment containing the working electrode was transferred by a specific syringe to the gas chromatography, (Shimadzu) where the amount of O2 evolution was determined. The Faradaic efficiency was determined from the total amount of charge Q (C) passed through the cell and the total amount of the produced O2nO2 (mol): Faradaic efficiency=4F × nO2/Q, where F is the Faraday constant, assuming the four electrons are needed to produce one oxygen molecule.The measurements of O0.75V0.25-LDH. Perdew-Burke-Ernzerhof (PBE)2+ and U−J=4.3 eV for Fe3+ and U−J=3.4 eV for V3+ species. One proton of the material is removed to create the metal ion at 3+ oxidation state. The structures were optimized until the maxima force on the atoms was smaller than 0.02 eV/Å.Calculations were carried out with DFT implanted in the Vienna Ab initio Simulation Package (VASP)394041The data that support the findings of this study are available from the corresponding author upon request.How to cite this article: Fan, K. et al. Nickel–vanadium monolayer double hydroxide for efficient electrochemical water oxidation. Nat. Commun. 7:11981 doi: 10.1038/ncomms11981 (2016).Supplementary Figures 1-9, Supplementary Table 1 and Supplementary References.


Puffinus puffinus, as a sentinel of environmental health, by determining contaminant levels  from carcass tissues and by isolating Vibrio spp. and Aeromonas spp. from live specimens. To this end, 35 Puffinus puffinus carcasses wrecked on the north-central coast of the state of Rio de Janeiro, Brazil, and two carcasses recovered in Aracruz, on the coast of the state of Espírito Santo, Brazil, were sampled, and fragments of muscle and hepatic tissues were collected for contaminant analyses. Swabs from eleven birds found alive at the north-central coast of Rio de Janeiro were collected for isolation of the aforementioned bacteria.Seabirds have been historically used to monitor environmental contamination. The aim of the present study was to test the suitability of a species belonging to the Procellariiformes group, the Manx shearwater, -1(liver) and 1.23 mg kg-1 (muscle); selenium 34.66 mg kg-1 (liver) and 7.98 mg kg-1 (muscle); cadmium 22.33 mg kg-1 (liver) and 1.11 mg kg-1 (muscle); and lead, 0.1 mg kg--1 (liver) and 0.16 mg kg-1 (muscle). Organochlorine compounds were detected in all specimens, and hexachlorbiphenyls, heptachlorbiphenyls and DDTs presented the highest levels. Regarding microbiological contamination, bacteria from the Vibrio genus were isolated from 91% of the analyzed specimens. Vibrio harveyi was the predominant species. Bacteria from the Aeromonas genus were isolated from 18% of the specimens. Aeromonas sobria was the only identified species.The average concentration in dry weight (dw) of the trace metals were: mercury 7.19 mg kgPuffinus puffinus seems to be a competent ocean health sentinel. Therefore, the monitoring of contaminant levels and the isolation of public health interest bacteria should proceed in order to consolidate this species importance as a sentinel.The results indicate that  Oceans cover approximately 70% of the earth surface and abouAs environmental degradation accelerates, science has increasingly focused on the influence of the environment on human health. Environmental degradation has direct impacts on life quality and health conditions of the human population. ConsequVibrio and Aeromonas genera/S, where SDbr is the standard deviation from 10 blank readings, and S is the inclination of the calibration curve. This formula provides the instrumental LQ. In order to calculate the final LQ, this formula was multiplied by the dilution factor.The organochlorine pesticides (OCPs) analyzed in this study were: hexachlorocyclohexanes (HCH) , hexachlorobenzene (HCB), heptachlor, heptachlor epoxide A and B, chlordanes , drins , dichlorodiphenyltrichloroethane  and its metabolites, dichlorodiphenyldichloroethylene  and dichlorodiphenyldichloroethane , endosulfan II, methoxychlor and mirex. The analyzed polychlorinated biphenyls (PCBs) congeners were the following IUPAC numbers: 8, 28, 31, 33, 44, 49, 52, 56/60, 66, 70, 74, 77, 81, 87, 95, 97, 99, 101, 105, 110, 114, 118, 123, 126, 128, 132, 138, 141, 149, 151, 153, 156, 157, 158, 167, 169, 170, 174, 177, 180, 183, 187, 189, 194, 195, 203, 206 and 209.et al. (1986)[et al. (2012)[The analytical procedure followed the protocol described by MacLeod . (1986), with mi. (2012).-1. The extract was concentrated to a volume of 0.9 mL in hexane. The internal standard 2,4,5,6-tetrachlorometaxylene (TCMX) was added before the gas chromatographic analysis was undertaken and a procedural blank was included in the set of samples.At first, 2,2’,4,5’,6-pentachlorobiphenyl (PCB 103) and 2,2’,3,3’,4,5,5’,6-octachlorobiphenyl (PCB 198) were added to all samples, blanks and reference material as surrogates for OCPs and PCBs. Then, approximately 2.5 g of each lyophilized muscle sample were extracted in a Soxhlet apparatus for 8 h using 80 mL of n-hexane and methylene chloride . The determination of extractable lipids was made by gravimetric analyses. The extracts were cleaned-up through the use of column chromatography with 8 g of silica and 16 g of alumina, both 5% water deactivated, eluted with 80 mL of n-hexane and methylene chloride . The fraction was purified once again, in order to remove lipid excess, now through high-performance liquid chromatography (HPLC), using methylene chloride as eluent with a flow of 5 mL min-1. The carrier gas used was hydrogen (constant pressure of 40 kPa at 100°C), and the makeup gas was nitrogen, at a rate of 60 mL min-1. Injector and detector temperatures were 280°C and 320°C, respectively. Oven temperature was programmed as follows: 70°C for 1 min, raised at 40°C.min-1 until 170°C, then raised at 1.5°C.min-1 until 230°C (held for 1 min), and at 20°C.min-1 until 300°C with a final hold of 5 min.Identification and quantification analyses of the organochlorine pesticides were performed with a 6890 N gas chromatograph with an electron capture detector (GC-ECD) (Agilent Technologies), using a 30 m × 0.25 mm i.d. capillary column coated with 5% phenyl-substituted dimethylpolysiloxane phase (0.5 μm film thickness). Automatic splitless injections of 2 μL were applied and the total purge rate was adjusted to 50 mL min-1). The interface, source and quadrupole temperatures were 280°C, 300°C and 200°C, respectively. Oven temperature was programmed as follows: 75°C for 3 min, raised at 15°C.min-1 until 150°C, then raised at 2.0°C.min-1, until 260°C and at 20°C.min-1, until 300°C with a final hold of 10 min.The quantitative PCBs analyses were performed by a 5973 N gas chromatograph coupled to a mass spectrometer (GC–MS) (Agilent Technologies), in a selected ion mode (SIM 70 eV), using a 30 m × 0.25 mm i.d. capillary column coated with 5% phenyl-substituted dimethylpolysiloxane phase (0.25 μm film thickness). Injections were made with 1 μL in automatic splitless mode. The carrier gas was helium (constant flow of 1.1 mL min-1 to 8.5 ng.g-1 dry weight (dw).The analytical methodology was validated through the use of a standard reference , from the National Institute of Standards and Technology, USA, for quality assurance and quality control. This material was analyzed in parallel, in duplicate, with an average analyte recovery inside the range accepted by the NS&T. The samCloacal, oral, ocular and tracheal swabs were carefully collected in order to avoid external contamination and accommodated in Cary-Blair media for transportation.Samples were enriched with Alkaline Peptone Water (APW) containing 1% sodium chloride (NaCl) (37°C/18-24 hours). Samples were then streaked onto Thiossulfate Citrate Bile Salts Sucrose Agar (TCBS) and onto Glutamate Starch Phenol Red Agar (GSP) and incubated to 37°C over night . Suspected colonies were transferred to Kligler Iron Agar, Lysine Iron Agar and Nutrient Agar with 1% NaCl. Posteriorly, biochemical tests were performed in order to identify species from the Vibrionaceae and Aeromonadaceae families, according to Noguerola & Blanch (2008) and JandVibrio spp. and Aeromonas spp. were calculated, as well as the frequency of isolation of each species.Statistical analyses were performed using the SPSS Statistics 17.0 software (IBM). Results from metal, organochlorine and bacteriological analysis were analyzed separately. Basic descriptive statistics was conducted. Data were tested for normal distribution using the Shapiro-Wilk’s test. Since most data were not normally distributed, non-parametric tests were used. Spearman’s rank correlation coefficient was used to verify the strength of the association between the concentration of the same metal in hepatic and muscular tissue and to verify the strength of the association in interelement relationship. A p-value of less than 0.05 was used to indicate statistical significance. For bacteriological analyses, the frequencies of isolation of The authors declare that they have no competing interests.MDC conceived of the study, carried out metal and statistical analyses and drafted the manuscript; JFM and DCT collected samples in the field, carried out necropsies and helped in several steps, including metal and statistical analyses and in the drafting of the manuscript; RAG helped in the metal analyses; FIC and RCM carried out the organochlorine compound analyses; EMR, RLS and DPR carried out the microbiological analyses; SS conceived the study, participated in its design and coordination and helped draft the manuscript. All authors read and approved the final manuscript.


Embryonic stem cells (ESCs) can self-renew or differentiate into any cell type, a phenomenon known as pluripotency. Distinct pluripotent states, termed naive and primed pluripotency, have been described. However, the mechanisms that control naive-primed pluripotent transition are poorly understood. Here, we perform a targeted screen for kinase inhibitors, which modulate the naive-primed pluripotent transition. We find that XMD compounds, which selectively inhibit Erk5 kinase and BET bromodomain family proteins, drive ESCs toward primed pluripotency. Using compound selectivity engineering and CRISPR/Cas9 genome editing, we reveal distinct functions for Erk5 and Brd4 in pluripotency regulation. We show that Erk5 signaling maintains ESCs in the naive state and suppresses progression toward primed pluripotency and neuroectoderm differentiation. Additionally, we identify a specialized role for Erk5 in defining ESC lineage selection, whereby Erk5 inhibits a cardiomyocyte-specific differentiation program. Our data therefore reveal multiple critical functions for Erk5 in controlling ESC identity.  •A kinase inhibitor screen identifies Erk5 as a key regulatory of ESC pluripotency•Erk5 suppresses transition to primed pluripotency and neural differentiation•Erk5 controls ESC identity by suppressing cardiomyocyte differentiation Williams et al. combine chemical screening and genetic approaches to identify Erk5 kinase as a critical regulator of the naive-primed pluripotent transition and cardiomyocyte differentiation. Embryonic stem cells (ESCs) can self-renew or potentially differentiate into all cell types, a phenomenon known as pluripotency . DistincUnderstanding the mechanisms that control naive-primed pluripotent transitions is fundamental to ESC biology. In this regard, mouse ESCs (mESCs) provide a tractable model, as they undergo dynamic transition between naive and primed pluripotency when cultured in leukemia inhibitory factor (LIF) and fetal bovine serum (FBS) . LIF-JakSmall-molecule screening represents a simple approach to elucidate kinase regulators of pluripotency. In a screen for modifiers of the naive-primed transition, we uncover XMD series compounds, which selectively inhibit the Erk5 kinase and BET bromodomain family, as drivers of primed pluripotency. Using rational inhibitor engineering and genome editing, we deconvolve individual roles of Erk5 and Brd4 in pluripotency regulation. Erk5 promotes expression of a network of naive pluripotency factors, which requires kinase activity, upstream activation by Mek5, and a C-terminal transcriptional domain. Furthermore, Erk5 signaling potently suppresses the transition of naive cells toward primed pluripotency and neuroectoderm differentiation. Finally, we show that Erk5 has a distinct function in suppressing late-stage cardiac gene expression and cardiomyocyte development.http://lincs.hms.harvard.edu) and identified those that modulate the naive-primed transition  and Brd4 genes. Multiple Erk5−/− mESC clones display elevated Dnmt3b expression, while Nanog remains relatively unperturbed  and upstream phosphorylation by Mek5 . Mek5DD expression has relatively minor effects on Oct4 but robustly maintains key naive markers Nanog, Klf2, Esrrb, and Rex1 following LIF withdrawal E, indica+/+ or Erk5−/− mESCs  mimics differentiation of pluripotent cells during development. We employed a panel of mESC lines generated by CRISPR/Cas9 .Many reagents generated for this study are available by request at the MRC-PPU reagents website (http://lincs.hms.harvard.edu). JWG-071 and JWG-045 were synthesized by the Gray lab.Antibodies used were Nanog (ReproCell Inc.), Dnmt3b (Imgenex), Klf4 (R&D Systems), Erk1/2 and Oct4 (Santa Cruz Biotechnology), phospho-p44/42 MAPK (Erk1/2 Thr202/Tyr204), Stat3α, phospho-Stat3 (Tyr705) and c-Myc , Erk5 , Klf2 (Millipore), CD309 (Flk1) APC, Clone Avas 12a1, and CD140a (PDGFRα) PE, Clone APA5 (eBioscience). AZD4547, PD173074, PD0325901, PD184352, ruxolitinib, and tofacitinib were from the DSTT . The 228 kinase inhibitor library was curated by the Gray lab .mESCs were cultured on gelatin coated plates in media containing LIF, 10% fetal calf serum (Gibco), and 5% knockout serum replacement (Invitrogen) unless otherwise stated. mESCs cells were transfected using Lipofectamine LTX (Life Technologies) and selected with puromycin for 48 hr. For CRISPR/Cas9, mESCs were transfected with pX335 and pKN7 (Addgene) and selected, then either lysed or clones isolated. To generate stable lines, Erk5−/− mESCs were electroporated with 30 μg linearized pCAGGS vector, plated at clonal density, and clones were analyzed by immunoblotting. Cell extracts were made in lysis buffer  and anti-mouse-HRP (Dnmt3b), respectively.3 × 10Genomic DNA was extracted using the DNeasy Blood and Tissue Kit (QIAGEN), and the Erk5 gene was analyzed by PCR sequencing .mESCs were cultured at 1,000 cells per six wells for 6–7 days prior to AP staining solution. For colony phenotyping, mESCs were plated at 200 cells per 10-cm dish and analyzed after 7 days. For proliferation assay, mESCs were seeded at 10,000 cells per six wells and counted.EBs were formed by aggregating 60,000 mESCs/ml in the absence of LIF for 4 days before transfer to gelatin-coated plates for an additional 4 days.At day 4, EBs were treated with VEGF , Activin A , and BMP4  and dissociated at day 7 by incubation with TrypLE (Invitrogen) and stained.RNA was extracted using the OMEGA total RNA kit and reverse transcribed using iScript reverse transcriptase (Bio-Rad). qPCR was performed using SsoFast EvaGreen Supermix (Bio-Rad). The ΔCt method using GAPDH as a reference gene was used to analyze relative expression and the 2-ΔΔCt (Livak) method used to normalize to control. Primers used are listed in 2, 2 mM DTT, and 0.1 mM [γ- 32P]-ATP [500 cpm/pmol]) and incubated at 30°C for 30 min. The assay was terminated by SDS sample buffer and heating and analyzed by SDS-PAGE and autoradiography.Erk5 was immunoprecipitated from mESC lysate using 5 μg anti-Erk5 antibody. Beads were washed three times in lysis buffer containing 0.5M NaCl, then resuspended in a total volume of 25 μl kinase assay buffer , and 250 μM PIMtide (ARKKRRHPSGPPTA) and incubated at 30°C for 20 min. The assay was terminated by applying the reaction mixture onto p81 paper and incorporated radioactivity measured.200 ng pure active ERK5 was incubated with the indicated inhibitor in 50 mM Tris-HCl (pH 7.5), 0.1 mM EGTA, and 1 mM 2-mercaptoethanol. The reaction was initiated by adding 10 mM magnesium acetate, 50 μM [γ-Escherichia coli and biotinylated acetylated peptides. Brd4-1 protein and inhibitors were preincubated for 30 min in 50 mM HEPES (pH 7.5), 100 mM NaCl, 0.05% CHAPS, and 0.1% BSA, then incubated for a further 30 min after addition of tetra-acetylated histone H4 peptide (H4 (1-21) K5/8/12/16(Ac)4-Biotin) and the streptavidin-coated donor beads. Ni-chelate acceptor beads were added and incubated for 1 hr, and signals were measured by Envision (Ex/Em = 680/520–620 nm).Brd4-1 binding assay was performed by Reaction Biology Corp. using His-tag Brd4-1 proteins expressed in Data are presented as the average with error bars indicating SD. Statistical significance of differences between experimental groups was assessed using a Student’s t test. Differences in averages were considered significant if p < 0.05. Representative western blots are shown.C.A.C.W., R.F.-A., J.W., and G.M.F. designed, performed, and analyzed experiments. R.T. and N.S.G. provided reagents and expertise. G.M.F. wrote the paper.


Lampaya medicinalis Phil. (Verbenaceae) is a plant used by Aymara and Quechua ethnic groups from Northern Chile as folk medicine in the treatment and cure of various diseases. The aim of this study was to investigate the in vitro antioxidant activity, total phenols content, total flavonoids content, total antioxidant activity, reducing power, brine shrimp cytotoxicity and identify the principal chemical constituents.3 colorimetric method, respectively. Reducing power was determined by phosphomolybdate and hexacyanoferrate (III) methods. Biotoxicity assays were performed on shrimps of Artemia salina. The EAF was fractionated using chromatographic methods.The crude hydroethanolic extract (HEE) and its partitioned fraction: hexane (HF), dichloromethane (DF), ethyl acetate (EAF), n-butanol (BF) and soluble residual aqueous fraction (RWF) were evaluated for their antioxidant activity using different assays namely, DPPH, ABTS, FRAP, β-carotene bleaching assay. The content of total phenolics and total flavonoids were measured by Folin-Ciocalteau and by the AlClConsiderable amount of phenolic and flavonoid contents were recorded in the hydroethanolic extract (HEE) and its derived fractions. Although HEE and all its derived fractions exhibited good antioxidant activities, the most distinguished radical scavenging potential was observed for ethyl acetate fraction (EAF). EAF showed the higher radical scavenging activity by DPPH (95%) and by ABTS (98%), antioxidant activity by FRAP , β-carotene bleaching assay (86.8%), the highest total phenols content (101.26 ± 1.07 mg GAE/g fraction), the highest total flavonoids content (66.26 ± 3.31 μg quercetin/g fraction). The EAF extract showed an reducing power of 78% and 65% using the phosphomolybdate and hexacyanoferrate (III) assays, respectively.Lampaya medicinalis for the first time.Four flavonoids, two p-hydroxyacetophenone derivatives and one iridoid were isolated from L.medicinalis can be used as an effective natural source of antioxidant, as ethnomedicine and as a commercial basis for the development of nutraceuticals.EtOAc soluble fraction (EAF) shows the strongest antioxidant activity, and it can be attributed to its high content in phenolic and flavonoid compounds. It can be concluded that  Free radicals are chemical species which contain one or more unpaired electrons in their outer shell. They are highly unstable and cause damage to other molecules by extracting electrons from them in order to attain stability. Free radicals result from an imbalance between the generation of Reactive Oxygen Species (ROS) and the antioxidant protection conferred by enzymatic systems. Since 1950s, free radical reactions are directly involved in aging and progression of several diseases . In the When there is a lack of materials to quench the excess of reactive free radicals, oxidative stress occurs which can lead to serious diseases. Free radicals then react with various cellular components causing serious damage. So there is a great need for antioxidant supplements to avoid conflict oxidative damage. In the food industry, an antioxidant is defined as a substance that, in low concentrations, is capable of preventing or delaying oxidation of easily oxidizable biomolecules. In mechanistic terms, an antioxidant can be defined as a hydrogen donor or an electron donor. The use of plants as food and medicinal remedies from ancient times is partially attributed to biological efficacy of secondary metabolites that possess antioxidant activities such as phenolic compounds . MedicinMedicinal plants comprise a source of biologically active compounds, which possess different chemical structures and diverse biological activities, and they may, due to the synergistic actions act as powerful antioxidants. Among these compounds, flavonoids and phenolic compounds, phytochemicals with a wide distribution in nature, have been the subject of several studies on their antioxidant activity, which is mainly due to their ability to act as free radical scavengers and/or a metal chelator. There is an increasing interest in the natural antioxidant contained in medicinal plants, which are candidates for the prevention of oxidative damage .Andean High Plateau in Northern Chile, known as “Puna atacameña”, is a particular biotope characterized by varying altitudes of between 3000–4200 meters above sea level, a very low relative humidity, no rains, and cloudless skies during most of the year. Around 5000 people live in this peculiar ecological system and they use medicinal plants as curatives or palliatives of health problems because the plants are recognized as a traditional way to treat ailments and diseases .Lampaya medicinalis Phil (Verbenaceae) commonly known, as “lampaya” is a small bush, with a height of 80–100 cm that grows in the “Puna atacameña” in Northern Chile. There is little or no information about the chemical composition or medicinal properties of this plant in the scientific literature  × 100, where Asample is the absorbance of sample or standard, and Ablank is the absorbance of the ABTS solution only. All tests were run in triplicate and averaged. The results reported are expressed as their IC50 through Graph prism pad software.HEE, HF, DF, EF, BF and RWF were tested by using the ABTS assay. The assay involve the use of a pre-formed ABTS radical cation /Ablank] × 100, where Asample is the absorbance of sample or standard, and Ablank is the absorbance of the DPPH solution only and IC50 value was calculated by graph pad prism software. All test were run in triplicate and averaged.The free-radical scavenging activity of various fractions, gallic acid and ascorbic acid was measured with the stable radical diphenylpicryhydrazyl (DPPH) in terms of hydrogen donating or radical scavenging activity. DPPH  solution was added to 100 μL of different fractions (dissolved in respective solvent), at different concentration (1–250 μg/mL). After storage at room temperature for 30 min, the absorbance was measured at 517 nm against a blank  were then taken at 539 nm. The results were expressed as mg of Trolox equivalent/g extract [Ferric reducing/antioxidant power was measured according to the method described by Suarez et al. with some modification . The sto extract .OR). The rate of β-carotene bleaching (R) was calculated according to equation R = [ln(Ao/At)]/t, where ln is natural logarithm, Ao is Absorbance of the emulsion at time 0, At is Absorbance at time t, and t is 20, 40, 60, 80, 100 or 120 min. The AA% was calculated according to the following equation: AA % = [Rblank - Rsample/Rblank] × 100, where Rsample and Rblank represent the bleaching rates of β-carotene with or without the addition of sample, respectively. The ROR was calculated according to equation ROR = (Rsample/Rblank).The inhibition activity of β-carotene oxidation by peroxide radicals of samples was determined as described by Wu et al. . 1 mL ofblank - Asample)/Ablank] × 100 where Asample is the absorbance of sample, and Ablank is the absorbance of blank solution, containing all reagents except the test sample.Reducing power was determined by phosphomolybdate method, using ascorbic acid as standard . Reagent3 solution. The absorbance was measured at 700 nm against a blank. The control contained all reagents except the extract fraction while water was used as blank. Ascorbic acid was used as a standard. Higher absorbance indicates higher reducing power. The increase of reducing power by the extract and standard was calculated as follows: Percentage of increase reducing power = [(Atest - Acontrol)/Atest] × 100, where Atest is absorbance of test solution and Acontrol is absorbance of control [The reducing power of all fractions and ascorbic acid were determined based on the method described by Rahman et al. . Differe control .Artemia salina Leach naupliis according to previously described methods, in three independent 36-h exposure experiments at 22–25°C. Artemia salina eggs were incubated at room temperature for 48 h with the help of a light source, in seawater boiled and filtered [50 at 95% confidence limit by the Finney Probit analysis program. HgCl2 was used as a positive control, methanol was used as solvent, brine shrimp larvae in only sea water were taken as a negative control.The bioactivity of each extract was monitored by the brine shrimp lethality test against filtered ,25. Ten p < 0.05 was considered to be statistically significant.All data were expressed as means ± SD. Analysis of variance was performed by ANOVA procedure. Duncan’s new multiple-range test was used to determine the differences of means, and A sample of EAF was separated by successive column chromatography over silica gel using n-hexane and EtOAc gradient with increasing amounts of EtOAc to obtain three crystalline colourless and four crystalline yellow products.Structure assignments of natural products were done based on their physical and spectroscopic properties: UV, IR, NMR and MS analysis and comparison with data reported in the literature.1H NMR  δ 3.88 , 3.90 , 6.58 , 6.37 , 6.48 , 7.84 , 7.02 , 7.98 , 12.10 . 13C NMR  δ 163.35 (C-2), 104.27 (C-3), 182.30 (C-4), 157.60 (C-5), 98.20 (C-6), 165.60 (C-7), 92.90 (C-8) 152.60 (C-9), 105.00 (C-10), 122.59 (C-1′), 126.36 (C-2′), 114.48 (C-3′), 161.63 (C-4′), 114.48 (C-5′), 126.36 (C-6′), 56.10 , 55.00 .1. Yellow crystal, mp 173–174°C (EtOAc - n-hexane). 1H NMR  δ 3.82 , 3.85 , 3.89 , 6.50 , 6.29 , 6.41 , 7.25 , 6.91 , 7.45 , 12.10 . 13C NMR  δ 163.80 (C-2), 103.07 (C-3), 179.14 (C-4), 163.30 (C-5), 98.20 (C-6), 167.60 (C-7), 92.90 (C-8), 156.60 (C-9), 106.00 (C-10), 122.99 (C-1′), 114.90 (C-2′), 147.48 (C-3′), 149.00 (C-4′), 114.90 (C-5′), 121.00 (C-6′), 56.50 , 56.20 , 55.90 .2. Yellow needles, mp 163–165°C (EtOAc–n-hexane). 1H NMR  δ 3.81 , 3.93 , 6.42 , 6.30 , 6.51 , 7.25 , 6.97 , 7.43 , 12.72 . 13C NMR  δ 163.92 (C-2), 103.23 (C-3), 180.62 (C-4), 161.05 (C-5), 96.30(C-6), 165.15 (C-7), 92.70 (C-8) 155.90 (C-9), 106.50 (C-10), 123.59 (C-1′), 114.90 (C-2′), 147.48 (C-3′), 145.10 (C-4′), 119.00 (C-5′), 121.60 (C-6′), 55.75 , 55.90 ,3. Yellow needles, velutin, mp 216–218°C (EtOAc - n-hexane). 1H NMR  δ 3.75 , 3.91 , 3.93 , 6.86 , 6.94 , 7.59 , 6.95 , 7.58 , 12.95 . 13C NMR  δ 163.98 (C-2), 103.23 (C-3), 182.62 (C-4), 162.65 (C-5), 131.90 (C-6), 158.65 (C-7), 91.68 (C-8), 152.50 (C-9), 106.50 (C-10), 121.39 (C-1′), 103.10 (C-2′), 148.10 (C-3′), 150.78 (C-4′), 120.50 (C-5′), 118.70 (C-6′), 56.40 , 55.99 , 60.00 .4. Yellow needles, cirsilineol, mp 204–206°C (EtOAc–n-hexane). 1H NMR  δ 2.57 , 6.54  , 6.92 , 7.91 . 13C NMR  δ 26.25 (Me), 115.53 (C-3 and C-5), 129.66 (C-1), 131.19 (C-2 and C-6), 161.27 (C-4), 198.50 (CO).5. Colorless crystal, mp 109–111°C. 1H NMR  δ 2.38 , 3.20 , 3.30 , 3.32 , 3.70 , 4.41 , 4.17 , 5.07 . 13C NMR  δ 26.25 (Me), 116.07 (C-3 and C-5), 130.98 (C-1), 130.23 (C-2 and C-6), 160.92 C-4), 196.45 (CO), 63.30 (C-6′), 70.15 (C-4′), 73.23 (C-2′), 74.02, (C-5′), 76.43, (C-3′), 99.62 (C-1′).6. Colorless crystal, picein, mp 195°C. 2O). 1H NMR  δ 2.08 , 2.55 , 2.90 , 3.22 , 3.74 , 4.33 , 4.83  5.88 , 7.53 . 13C NMR  168.17 (COOMe), 152.74 (C-3), 142.26 (C-8), 130.13 (C-7), 110.96 (C-4), 95.33 (C-1), 61.53 (C-10), 51.50 (Me), 48.38 (C-9), 39.24 (C-6), 36.22 (C-5).7. Colorless crystal, mp 120 – 122°C  and its partitioned fraction of Lampaya medicinalis, are summarized in Table 50 values). Overall, the IC50 of ABTS free radical scavenging capacity of six extracts was found between 5.16 ± 1.09 to 56.32 ± 1.70 μg/mL. EAF, HEE, and BF showed comparatively strong ABTS free radical scavenging capacity. Similarly, DF and RWF showed moderate activity. Figure Lampaya medicinalis. It was found that the radical-scavenging activities of all extracts increased with the concentration. All test samples exhibited potent scavenging activities in a concentration dependent manner. At a concentration of 50 μg/mL, scavenging activities of EAF reached a plateau of 98% while, at same concentration, scavenging effects of HEE, DF, BF, RWF and HF were 78%, 71%, 70%, 67%, and 39%, respectively. While at the same concentration the activity of Trolox was 99%. The antioxidant activity of HEE and its fractions and standard Trolox increased in the following order HF < RWF < BF < DF < HEE < EAF ≈ Trolox.ABTS radical scavenging activity involves a more drastic radical, chemically produced and is often used for screening complex antioxidant mixtures such as plant extracts, beverages and biological fluids. The actual version of this assay, a stable ABTS radical cation which has a blue-green chromophore absorption, was produced by oxidation of ABTS with potassium persulfate. Antioxidant activity of the natural products is determined by the discoloration of the ABTS, by measuring the reduction of the absorbance at 734 nm. The extension of discoloration, expressed as a percentage inhibition of ABTS50, which is the amount of antioxidant necessary to decrease the initial DPPH˙ by 50% expressed in μg/mL. The results of DPPH scavenging activity of hydroethanolic extract (HEE) and its partitioned fraction of Lampaya medicinalis, are summarized in Table 50 values). Overall, the IC50 of DPPH free radical scavenging capacity of six samples were found between 9.94 ± 0.18 to 27.17 ± 0.64 μg/mL. EAF, RWF, HEE and BF, showed comparatively strong DPPH free radical scavenging capacity. Similarly, DF and HF showed moderate activity. Figure Lampaya medicinalis. It was found that all test samples exhibited potent scavenging activities in a concentration dependent manner. At a concentration of 50 μg/mL, the scavenging activities of EAF reached a plateau of 95% while, at same concentration, the scavenging effects of BF, RWF, HEE, DF and HF were 81% , 79% , 74%, 71% and 60%, respectively.2,2-diphenyl-1-picrylhydrazyl (DPPH), in solution, is a stable free radical, which has been widely accepted as a tool for estimating free radical scavenging activities of antioxidants. One of the reasons is that this method is simple and highly sensitive. This assay is based on the theory that a hydrogen donor is an antioxidant. DPPH is one of the few stable and commercially available organic nitrogen radicals. The antioxidant effect is proportional to the disappearance of DPPH˙ in test sample. Various methods of monitoring the amount of DPPH˙ in the antioxidant test system have been reported, however monitoring with UV spectrometer has become the most widely used. DPPH˙ accepts an electron or hydrogen radical to become a stable diamagnetic molecule. This reaction is stoichiometric with respect to the number of hydrogen atoms absorbed, or to the number of electrons captured. The reduction capability of DPPH radical is determined by the decrease in absorbance at 517 nm induced by antioxidants. Then, color changing from purple to yellow is the consequence of the reducing ability of antioxidant toward DPPH stable free radical. The higher the discoloration of the DPPH methanol solution, the lower the absorbance of the reaction mixture, indicating thereby significant free radical scavenging capacity. The results have been reported as IC+3 to Fe+2 ions in the presence of TPTZ -s-triazine), forming an intense blue Fe-TPTZ complex with an absorbance maximum at 593 nm. Decreased absorbance is proportional to the antioxidant content. The FRAP assay results of the HEE and its fractions expressed as mg equivalent of Trolox/g fraction are shown in Table Ferric reducing antioxidant power assay is based on the ability of an antioxidant to reduce Fe2. In this antioxidant assay a mixture of β-carotene and linoleic acid is used. The peroxyl radical reacts with β-carotene, subsequently, the amount of β-carotene reduces in a testing solution, resulting in the rapid disappearance of color. If an antioxidant is present in a testing solution, it reacts competitively with peroxyl radical and color is maintained for a longer period. Therefore, antioxidant activities are monitored by bleaching the color of a test solution at 470 nm, which is the typical absorbance by β-carotene. The IC50 values of β-carotene bleaching assay of Lampaya medicinalis extract and various fractions is summarized in Table 50 values of 65.41 ± 3.56 μg/mL and 370.96 ± 3.67 μg/mL respectively. The antioxidant activity (%) of HEE and its fractions and ascorbic acid increased with an increasing concentration of the sample. The antioxidant activity of HEE and its fractions and ascorbic acid increased in the following order HF < RWF < DF < HEE < BF < EAF ≈ ascorbic acid. At a concentration of 250 μg/mL, EAF and ascorbic acid exhibited 86.78 ± 3.65% and 86.56 ± 2.98% as antioxidant activity, respectively. Figure Lampaya medicinalis extract and various fractions.It is widely know that β-carotene reacts with the peroxyl radical to produce β-carotene epoxides acting as a radical scavenger or antioxidant. The linoleic acid forms a peroxyl radical in the presence of ROS and OLampaya medicinalis extracts and various fractions was estimated from their ability to reduce de Mo (VI) to Mo (V) and resulting in formation of green phosphate/Mo (V) complex with maximum absorption at 695 nm. Increased Absorbance of the reaction mixture indicated increased total antioxidant capacity. In the present assay all the different fractions showed good total antioxidant index, which was concentration-dependent. The results of reducing power of hydroethanolic extract (HEE) and its partitioned fraction are summarized in Table 50 values). Overall, the IC50 of five samples were found between 26.93 ± 2.09 to 48.31 ± 3.13 μg/mL. Figure Lampaya medicinalis. It was found that all test samples exhibited potent activities in a concentration dependent manner. At a concentration of 100 μg/mL, the antioxidant activity of HEE, its fractions and ascorbic acid increased in the following order HF < RWF < DF < HEE < EAF < BF ≈ Ascorbic acid.The total antioxidant potential of +3/Hexacyanoferrate (III) complex to Fe+2/Hexacyanoferrate (III) complex, so the reducing power of sample can be monitored by measuring the formation Perl’s Prussian Blue complex at 700 nm. Increased absorbance indicated increase antioxidant capacity.Reduction power, widely used in evaluating antioxidant activity of plants, was determined by the potassium hexacyanoferrate (III) reduction method. In this assay, the presence of reductans in the antioxidant sample causes the reduction of the Fe50 values of reducing power assay of Lampaya medicinalis extract and various fractions are summarized in Table 50 values of 40.23 ± 3.89 μg/mL and 162.45 ± 5.11 μg/mL respectively. The reducing activity (%) of HEE and its fractions and ascorbic acid increased with an increasing concentration of the sample in the following order HF < RWF < DF < HEE < BF < EAF ≈ ascorbic acid, as shown in Figure The IC50 is between 500–1000 μg/mL, moderate the LC50 is between 100–500 μg/mL, and strong when the LC50 is between 0–100 μg/mL [Artemia were alive after 36 h of experiment in the negative control. All larvae in HgCl2 control died at dose 3 μg/mL. Test samples showed LC50 in the range of 33.36 ± 1.20 - 7.29 ± 0.20 μg/mL, indicating a strong cytotoxicity activity. Among all tested fractions, EAF (7.29 ± 0.20 μg/mL), BF (7.44 ± 0.31 μg/mL) and HEE (8.99 ± 0.30 μg/mL) showed comparatively higher cytotoxic activity. While less bioactive fractions were HF and DF, with LC50 33.36 ± 1.20 μg/mL. This result indicates that all fractions of Lampaya medicinalis may contain chemical compounds with pharmacological properties.A general bioassay that seems capable to detecting a wide range of biological properties of crude extracts is the brine shrimp lethality test. The technique is simple, inexpensive and use a small amount of test material, and it has been used to evaluate plants for potential pharmacological activity. In bioactivity evaluation by brine shrimp bioassay, cytotoxic activity is considered weak when the LC00 μg/mL . The degLampaya medicinalis. The structures of the compounds were based on detailed analysis of UV, IR, 1DNMR, 2DNMR, EM spectral evidence and by comparing data with the literature values. Chromatographic fractionation of EAF led to identification of four compounds belonging to the flavonoid class (1–4), two p-hydroxyacetophenone derivatives (5–6) and one iridoid (7) . Compound 1 have oxygenation patterns as apigenin 5,7,4′, Compounds 2 and 3 have oxygenation patterns as luteolin 5,7,3′,4′ and compound 4 is a 6-luteolin substituted and by comparing the data with the literature values the structures were identified to be 5-hydroxy-7,4′-dimethoxyflavone 1 [This is the first report of the presence of seven compounds in  iridoid   shows the strongest antioxidants activity and it can be attributed to its high content in phenolic and flavonoid compounds. It can be concluded that L.medicinalis can be used as an effective natural source of antioxidant, as ethnomedicine and as a commercial basis for the development of nutraceuticals.The results obtained in this study demonstrate that the hydroethanolic extract of The authors declare that they have no competing interests.GM and AP performed all experiments and read and approved the final manuscript.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1472-6882/14/259/prepub


We report here the draft genomes of two pneumococcal isolates in Hong Kong, CU_SPNE1_05 and CU_SPNE32_06. Strain CU_SPNE1_05 had a cefotaxime MIC of 1 µg/ml, and CU_SPNE32_06 had an MIC of 32 µg/ml. Both strains belong to the multidrug-resistant serogroup 19, sequence type 271 . Streptococcus pneumoniae serogroup 19 multilocus sequence type 271 (ST271) is a multidrug-resistant (MDR) clone and belongs to clonal complex CC 320/271  PCV7, PCV10, and PCV13. Among the serotypes included in PCV7, serotype 19F was the least likely to evoke a protective immune response, according to both vaccine efficacy trials and  studies , 7. Ther studies . During S. pneumoniae strain CU_SPNE1_05 was isolated from a sputum sample from a 90-year-old male in 2005, and strain CU_SPNE32_06 was isolated from a sputum sample from a 52-year-old male in 2006 in a university teaching hospital in Hong Kong. Both strains belonged to serotype 19F and ST271 (http://spneumoniae.mlst.net). Both CU_SPNE1_05 and CU_SPNE32_06 were multidrug resistant, with resistance to lincomycin, erythromycin, and tetracycline and to chloramphenicol and linezolid (The genomes were sequenced using Illumina HiSeq platform (90-bp paired-end reads). Genome assembly was performed with metAMOS pipeline . Prokka (version 1.11) was used for genome annotation, including open reading frame (ORF) finding, RNA gene prediction, and gene function annotation. The genome of CU_SPNE1_05 was assembled into 68 contigs (≥500 bp), with a total length of 2,038,146 bp (sequenced to ~11 million reads with ~400× coverage). The predicted genes from this genome include 1,998 coding sequences (CDSs), 3 rRNAs, and 24 tRNAs. For CU_SPNE32_06, 88 contigs (≥500 bp) were assembled, with a total length of 2,040,265 bp (sequenced to ~15 million reads with ~600× coverage). The predicted genes from the genome include 1,977 CDSs, 3 rRNAs, and 40 tRNAs.S. pneumoniae.Further studies are under way with the two genomes to understand the evolution of CTX resistance in LCSI00000000 and LCSJ00000000, respectively.The draft genome sequences of CU_SPNE1_05 and CU_SPNE32_06 have been deposited in the NCBI database under the GenBank accession numbers