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    Individual variability in foraging success of a marine predator informs predator management

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    Albedo changes caused by future urbanization contribute to global warming

    Land coverUrban landscapes are characterized by small clusters of patches, forming land mosaics that are distinct from natural landscapes. An accurate estimation of climate forcing requires a land cover dataset at high resolutions that does not omit small urban patches. In this study, the RF estimates are based on 500-m and 1-km land cover datasets. This fine resolution is necessary to preserve spatial details of small urban patches while avoiding the large underestimation of urban land areas at coarse resolution (e.g., ~19% underestimation at 10 km compared to that at 1 km)3. We used 500-m resolution MODIS Land Cover product (MCD12Q1v006) for historical land cover changes. For future urban land cover distributions, we used the global urban land expansion products simulated under the SSPs for 2030–2100 (i.e., Chen-2020)4. The simulation performance was tested using historical urban expansion from 2000 to 2015 based on Global Human Settlement Layer51, where the agreement between simulated and observed urban land was evaluated using the Figure of Merit (FoM) indicator52 that has showed similar or better values than those reported in other existing land simulation applications4. The high-resolution Chen-2020 also shows very high spatial consistency with the prominent coarse resolution global urban land projection LUH2 that is recommended in CMIP64. Considering different scenarios is also necessary to account for the uncertainties of future socioeconomic and environmental conditions, so we included simulated urban lands under three scenarios (Supplementary Table 1): Sustainability -SSP1, Middle of the Road – SSP2, and Fossil-fueled Development – SSP553. Within each SSP scenario, the product provides a likelihood map of each grid becoming urban, based on 100 urbanization simulations. We used the likelihood map to account for spatial uncertainties of urban expansion by deriving 90% confidence intervals of projected urban land demand within a SSP scenario. We used the MODIS IGBP Land Cover classes (Supplementary Table 2) and resampled the original 500-m resolution MODIS products in 2018 to 1-km resolution to match the future simulations when it was used as a baseline year. To isolate the independent effect of urbanization (vs other types of land uses) in future estimates, land covers that are not converted to urban are assumed to have the same cover types as in 2018 (i.e., the baseline year). Though there are other global land cover products for current periods, we choose the MODIS IGBP land cover products because the albedo look-up maps (LUMs) were based on IGBP land cover types (see Albedo Look-Up Maps).To further evaluate the uncertainties caused by different projections of future urbanization, we also included the other two SSPs from Chen-2020, and another two 1-km resolution urban land cover products projected for the future for the purpose of comparison. The other two products include four projections of SRES scenarios (i.e., A1, B1, A1B, and B2) (i.e., Li-2017 mentioned above)3 and one without scenario description but assumed historical development would continue (i.e., Zhou-2019 mentioned above)2. These projections of future urban land expansion were calibrated with different historical urban land products and can be regarded as independent.Albedo look-up maps (LUMs)Albedo Look-Up Maps (LUMs)31 were derived from the intersection of MODIS land cover54 and surface albedo55 products, which are used to determine the albedo values for each IGBP land cover type by month and by location. Monthly means of white-sky (i.e., diffuse surface illumination condition) and black sky (i.e., direct surface illumination condition) during 2001–2011 were processed for snow-free and snow-covered periods for each of the 17 IGBP land cover classes at spatial resolutions of 0.05°−1°31. The LUMs have been verified by comparing the reconstructed albedo using the LUMs with the original MODIS albedo, which shows very similar values31. We used the LUMs at a resolution of 1° due to the significantly fewer missing values, to assure the spatial continuity of albedo changes at a global scale while keeping the matches with the 1° resolution of radiation data and RF kernels. The underlying assumption is that albedo of the same land cover type varies insignificantly within a 1° grid.Snow and radiation productSnow cover can significantly change the albedo of land regardless of cover types (Supplementary Fig. 4). In this study, we tally monthly albedo using snow-free and snow-covered categories in estimating RF. Past and present snow-free and snow-covered conditions were derived from level 3 MODIS/Terra Snow Cover (MOD10CM.006)56 at 0.05° spatial resolution and resampled to a 1° spatial resolution. Monthly means of 2001–2005 vs 2015–2019 were used for 2001 and 2018 respectively. For future periods, ensemble mean snow cover for each year and month, projected under the CMIP5 framework for three Representative Concentration Pathway (RCP) scenarios (i.e., RCP2.6, RCP4.5, and RCP8.5) were used (for more details see Supplementary Note 2B). By comparing the model outputs with MODIS observations for a recent decade (2006–2015), we found that the multi-model mean snow cover was systematically biased compared to MODIS observations. Consequently, we calibrated the ensemble mean projections by subtracting the biases for the grids. In each 10th year of the future (e.g., 2030, 2040, etc.), the decadal monthly mean snow cover (e.g., 2026–2035 for 2030, and 2036–2045 for 2040, etc.) was used for the year.We used the long-term monthly averages (1981–2010) of diffuse and direct incoming surface solar radiation reanalysis Gaussian grid product from National Centers for Environmental Prediction (NCEP)57. Visible and near infrared beam downward radiation and diffuse downward radiation from NCEP were used to compute the white-sky and black-sky fractions. As for snow cover, ensemble mean shortwave radiation at surface (SWSF) and at top-of-atmosphere (SWTOA) projected from CMIP5 models (Supplementary Note 3C) for RCP2.6, RCP4.5, and RCP8.5 were collected for empirically computing future albedo kernels (see section 3.4 below).Radiative kernelsRadiative kernels were used to compute top-of-atmosphere RF due to small perturbations of temperature, water vapor, and albedo. We used the latest state-of-the-art albedo kernels calculated with CESM v1.1.258 to compute RF in 2018 relative to 2001. In brief, the albedo kernel is the change in top-of-atmosphere radiative flux for a 0.01 change in surface albedo. The CESM1.1.2 kernels are separated into clear- and all-sky illumination conditions. We used the all-sky kernels because we include both black-sky and white-sky albedos. For future periods, because there are no available radiative kernels produced from general circulation models, we approximated the future kernels using an empirical parameterization following Bright et al.59:$${K}_{m}left(iright)={{SW}}^{{SF}}(i)times {sqrt}left(frac{{{SW}}^{{SF}}(i)}{{{SW}}^{{TOA}}(i)}right)/(-100)$$
    (1)
    where m is the month, i is the location, and SWSF and SWTOA are the surface and top-of-atmosphere shortwave radiation; dividing by −100 is for matching the CESM1.1.2 kernel definition of a 0.01 change in surface albedo.Estimation of albedo change and RFWe analyzed the RF in 2018 due to albedo changes caused by urbanization since 2001 (2018–2001), and in the future from 2030 to 2100 at decadal intervals (i.e., 2030, 2040, 2050, …, and 2100) since 2018 under three illustrative scenarios: SSP1-2.6, SSP2-4.5, and SSP5-8.5, which combine SSP-based urbanization projections and RCP-based climate projections. The three illustrative scenarios were selected following the scenario designation of the latest IPCC report50 and represent low greenhouse gas (GHG) emissions with CO2 emissions declining to net zero around or after 2050, intermediate GHG emissions with CO2 emissions remaining around current levels until the mid-century, and very high CO2 emissions that roughly double from current levels by 2050, respectively. We selected 2018 as the baseline year to divide the past from the future because 2018 was the latest year with available MODIS land cover products at the time of this study. We used ArcGIS 10.6 to produce spatial maps of all variables, including area of each land cover type within a 1° × 1°-grid, snow cover, albedo, radiation, and kernels, and R 3.6.1 to compute the RF.We focused only on albedo changes induced by urbanization, including the conversions from all other 16 IGBP land cover types to urban land. The changes of albedo for each grid (x, y) of a month (m) were obtained by computing the difference between albedo of that grid in the baseline year (t = t0) and in a later year (t = t1) with urban expansion:$${triangle alpha }_{m,t1-t0}(x,y)={alpha }_{m,t=t1}(x,y)-{alpha }_{m,t=t0}(x,y)$$
    (2)
    where αm, t = t1 (x, y) and αm, t = t0) (x, y) is the albedo for each grid (x,y) of a month (m) at the base year and later year respectively; the grid-scale albedo is computed as the weighted sum of albedo by land cover types with the weighing factor corresponding to areal percentage of a land cover within the grid. The albedo for each land cover type of a grid was then obtained by applying the albedo LUMs that provide spatially continuous black-sky, white-sky, snow-covered, and snow-free albedo maps for a given month for each land cover. Firstly, monthly mean albedo is computed as:$${alpha }_{m,t}(x,y)=mathop{sum }limits_{l=1}^{17}mathop{sum }limits_{s=0}^{1}mathop{sum }limits_{r=0}^{1}{{f}_{l,t}(x,y){f}_{s,m,t}(x,y)f}_{r,m,t}(x,y)left({alpha }_{l,s,r,m}(x,y)right)$$
    (3)
    where m is the month, t is the year, l is the land cover type, fl is the proportion of a cover type within the grid, fs,m,t is the fraction for snow-covered (s = 0) and snow-free (s = 1) conditions of the time (m, t), fr,m,t (x, y) is the fraction for white-sky (r = 0) or black-sky (r = 1) conditions of the time, and αl,s,r,m (x, y) is the albedo for land cover type l in month m that is extracted from the albedo LUMs corresponding to snow condition (s) and radiation condition (r). The annual mean albedo change is reported as the mean of monthly albedo change:$${triangle alpha }_{t1-t0}(x,y)=frac{1}{12}mathop{sum }limits_{m=1}^{m=12}({alpha }_{m,t=t1}(x,y)-{alpha }_{m,t=t0}(x,y))$$
    (4)
    The conversion of other land covers to urban land can contribute differently to the global RF, as the total area that is converted into urban land is different among non-urban land covers and the albedo differences between urban land and non-urban land cover types vary. To estimate the proportional contributions of different land conversions, we first decomposed the total albedo of each grid into the proportion of each land cover type:$${alpha }_{l,m,t}(x,y)={f}_{l,m,t}(x,y)mathop{sum }limits_{s=0}^{1}mathop{sum }limits_{r=0}^{1}{f}_{s,m,t}(x,y){f}_{r,m,t}(x,y)left({alpha }_{l,s,r,m}(x,y)right)$$
    (5)
    The global RF due to albedo change caused by conversion from each non-urban land cover type (l ≠ 13) to urban land (l = 13) (see Supplementary Table 2 land cover labels) was calculated as:$${{RF}}_{triangle alpha ,l(lne 13),{global}}=frac{1}{{A}_{{Earth}}}mathop{sum }limits_{i=1}^{n}mathop{sum }limits_{m=1}^{12}{({alpha }_{13,m,t=t1}left(iright)-{alpha }_{l,m,t=t0}left(iright))Delta p}_{lto 13}left(iright){Area}left(iright){K}_{m}(i)$$
    (6)
    where i refers to a grid, n is the total number of pixels on global lands, AEarth is the global surface area (5.1  ×  108 km2), α13,m,t = t1) (i) is the albedo of urban land in month m in the later year with urban expansion, αl,m,t = t0 (i) is the albedo of a targeted non-urban land cover type in the base year t0, Δpl→13 is the percentage of the non-urban land cover type that is converted to urban land in the year t1 compared to year t0, Area(i) is the area of the pixel, and Km (i) is the radiative kernel at the grid.The global RF due to urbanization-induced albedo changes was then calculated as:$${{RF}}_{triangle alpha ,{global}}=mathop{sum }limits_{l=1}^{17}{{RF}}_{triangle alpha ,l,{global}}(l,ne, 13)$$
    (7)
    GWP: CO2-equivalentWe followed GWP calculations by explicitly accounting for the lifetime and dynamic behavior of CO2 to convert RF to CO2 equivalent60,61:$${GWP}[{kg},{of},{{CO}}_{2}-{eq}]=frac{{int }_{t=0}^{{TH}}{{RF}}_{triangle alpha ,{global}}(t)}{{k}_{{CO}_2}{int }_{t=0}^{{TH}}{y}_{{{CO}}_{2}}(t)}$$
    (8)
    where kCO2 is radiative efficiency of CO2 in the atmosphere (W/m2/kg) at a constant background concentration of 389 ppmv, which is taken as 1.76  ×  1015 W/m2/kg62, and RF∆α,global is the global RF caused by albedo changes (W/m2). ({y}_{{{CO}}_{2}}) is the impulse-response function (IRF) for CO2 that ranges from 1 at the time of the emission pulse (t = 0) to 0.41 after 100 years, and here it is set to a mean value of 0.52 over 100 years60. The time horizon (TH) of our GWP calculations was fixed at 100 years following IPCC standards and previous studies60,63,64.Global mean surface air temperature changeWe estimated the 100-year global mean surface temperature change for the estimated RF by adopting an equilibrium climate sensitivity (ECS), defined as the global mean surface air temperature increase that follows a doubling of pre-industrial atmospheric carbon dioxide (RF = 3.7 W/m2). Given a value of RF induced by a forcing agent, the temperature change is estimated as RF/3.7 × ECS. To consider the uncertainties of ECS, we adopted a mean value of 3 °C and a very likely (90% confidence interval) range of 2–5 °C following IPCC AR650. Without knowing the exact distribution shape of ECS and future albedo-change-induced RF, we created a log-normal distribution (Supplementary Note 4) to approximate their asymmetric distribution through numerical simulation. We then conducted Monte Carlo simulations that draw 5000 random samples from each distribution to jointly estimate the uncertainties of global mean surface air temperature changes. We report the mean and 90% interval ranges of the change in temperature. More

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    Sex differences in the winter activity of desert hedgehogs (Paraechinus aethiopicus) in a resource-rich habitat in Qatar

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    A sandponics comparative study investigating different sand media based integrated aqua vegeculture systems using desalinated water

    Study siteThe study was conducted at the Center for Applied Research on the Environment and Sustainability (CARES) at The American University in Cairo, New Cairo, Egypt (30°01′11.7″N 31°29′59.8″E) from 12/Nov/2019 until 31st/March/2020. The experiment was carried out in a greenhouse-controlled environment with temperatures ranging from 18 to 23 °C and relative humidity between 60 and 70% during the growing period.Experimental designThe proposed design starts by treating brackish water using RO membrane separation technology, powered by an on-grid 10 kW photovoltaic solar panel as shown in Fig. 1. The permeate (freshwater) from the RO facility is directed to the aquaculture units of capacity of 1 m3, where the fish effluents are used as irrigation water and as the sole source of fertilizers for the crops.Figure 1Schematic Integrated model design. T1 Deep water culture system without sand, T2 Sandponics system with sand from October, T3 Sandponics system with sand from Beni suef, T4 Sandponics system with sand from Fayoum.Full size imageThe study followed a completely randomized design with four variants, i.e., an aquaponic deep-water culture system (T1) and three sandponics systems (T2–T4). The three sandponics systems were established with different sand collected from different sand locations in Egypt during the period between September and October 2019.Initially, an exploratory field trip was set to six different locations in Egypt to collect sand samples for lab analysis aimed at sourcing the most suitable sand for the system under study with regards to both the physical and chemical parameters. These areas include Ismailia Governorate; 30°34′55.2″N 31°50′08.1″E, 6th October governorate; 29°54′49.8″N 31°05′51.5″E, Benu Suef governorate; 28°53′18.4″N 30°45′12.9″E, Al-Minya governorate; 28.725799, 30.630305, and two sites from Fayoum governorate; 29°05′07.4″N 30°49′39.9″E.From the six locations in Egypt, preliminary sand analysis was carried out, and sand samples were also collected for both physical and chemical lab analysis at the Soil and Water Lab at the Agricultural Research Center in Dokki, Egypt. Following a thorough technical, field, mechanical, and lab chemical evaluation of the six sand samples from six locations, three sand locations/types were selected for experimentation that seemed fit and suitable for the current study. The criteria parameters for the shortlisting of sand included water retention potential of the sand by the percolation process, testing the carbonates level in the soil, the turbidity of the sand, porosity percentage and drainage potential of the sand. The three locations included 6th October (T2), Benu Suef (T3), and Fayoum site 2 (T4). In the second week of November 2019, ten cubic meter tracks of sand from the three above locations were set to collect sand from these areas to the research facility at CARES where the experiment was carried out.The study was carried out with two systems/setups, i.e., an aquaponic Deep Water Culture (DWC) and SP systems. The DWC model comprises a 1 m3 fish tank, a settlement tank, a mechanical filter, a biological filter, three grow beds, and a drainage tank. This system being the most practiced aquaponics technique was considered as the control. Fish effluent water flowed from the fish tank to the settlement tank to filter big solid wastes through the mechanical filter to remove the smaller solid wastes and the biological filter for the nitrification process. Then filtered water continues to the grow beds, where overflow drains into the drainage tank and back to the fish tank in a closed system.On the other hand, the variable in the three IAVS systems is the sand source. This system comprises three independent set-ups: a 1 m3 fish tank, three grow beds, and a drainage tank. Fish effluents flowed from the fish tank directly to the sand grow beds where water was supplied through irrigation drip lines using diaghram emitters connected with valves to ensure uniformity of water application to each grow bed.All the fish tanks were installed with the same fish stock size of 30 Nile tilapia (Oreochromis niloticus) from an existing fish stock at the research center with an average initial weight of 244 g and the same amount of water, initially 850L per tank. The fish was sourced from an already existing aquaponics system at the research center to avoid any transportation stress effects and related shocks on the small fish, leading to a lot of mortality cases. The fish were fed 3–4 times daily with commercial pellets containing 30% proteins, 5% crude lipid, 6% crude fiber, 13% Ash, and 9% moisture content supplied by Skretting Egypt. The feeding pattern and frequency were according to the fish body biomass percentage of 2–3% depending on the growth stage and upon reaching satiation.DesalinationThe experiment was entirely run with desalinated water produced from a desalination facility at the center. The desalination technology used was Reverse Osmosis (RO); in batch mode; using a Sea Water Pump with Energy Recovery Unit (model Danfoss-APP1.0/APM1.2). The RO membrane used is Hydraunatic SWC5-4040, from Lenntech company with an average salt rejection of 99.7%. Three modules were connected in a series arrangement (3 Pressure Vessels each equipped with a single module). Synthesized brackish water was prepared by dissolving industrial grade sodium chloride (sea salt) from El-Arish Governorate, Egypt. The salt chemical properties are presented in Table 1. Feedwater salinity was 10 mg/L, with an equivalent osmotic pressure equal to 8.61 bars. The osmotic pressure was calculated using Van’t Hoff relation. Permeate Total Dissolved Solids (TDS) was 192 mg/L, and brine TDS was 13.1 g/L as shown in Table 2.Table 1 Chemical properties of the used salt.Full size tableTable 2 Chemical properties of water samples used.Full size tableThe average pure water flux is 9.5 LMH and was calculated by dividing the permeate volume by the product of membrane surface area and time. Each batch run produced around 4 m3 of permeate, which was enough to irrigate the designated plant beds. The estimated average permeate recovery for the RO process is 22% and salt rejection exceeded 98.7%. The differential pressure between membrane inlet and outlet was equal to 1 bar, where membrane inlet pressure was 16 bars, and the outlet was 15 bars. The RO process operated at an average transmembrane pressure equal to 16 bars and an average permeate and brine flow rates equivalent to 3.49 and 12.41 Lpm, respectively. All experiment runs were performed at 25 °C.Plant materials and cultivation practiceSwiss chard bright lights (Beta vulgaris subsp. cicia) seeds were imported from Seed kingdom seed company in the USA. Seeds were sown in ¼ inch holes in a seed starting mix containing perlite and vermiculite and irrigated with a hand mist sprayer daily to keep the growing media always moist. Sowing was done on the 12th of November 2019, and seedlings were transplanted when they were 40 days old. Seedlings were transplanted into raised grow beds made of fiberglass material measuring 1.8 × 1.2 × 0.6 m for each of the four systems. The beds were raised off the ground by 0.5 m to allow drainage water from the bed to be collected and circulated back to the fish tank. Each bed was constructed with a drainage pipe at the bottom covered with a mesh net to prevent water blockage by the sand. Also, a 5 cm layer of small gravel was uniformly laid at the bottom of the beds to facilitate drainage, followed by sand with a height of 50 cm.In the IAVS systems, plants were irrigated using manually punched diaphragm emitters, and the irrigation flow rate was controlled using small plastic valves at the start of every irrigation tube. Emitters were installed in drip tubing at a 30 cm distance as well the tubing lines were also placed 30 cm between each other. Seedlings were transplanted 5 cm away from the emitters at 30 cm between rows and 30 cm within the row. Since the water was pumped with submersible pumps to the grow beds, regulatory pressure valves were installed in between the pump and the main irrigation line, and then water flows through the emitters into the row furrows. Water would then saturate in the sand and eventually drain at the bottom into drainage tanks and pumped back to the fish tanks.To maintain the water quality, two full cycles of water recirculation were run every day. Every irrigation cycle recirculated 25% of the fish tank, and complete drainage was allowed for a maximum of two hours. Plants were harvested upon reaching maturity for three cuts, except with the T1, which could not grow back after the second cut. Plants took 52 days from transplanting to reach the first cut, 20 days from cut 1 to cut 2, and as well 23 days from cut 2 to reach cut 3. Measurable crop parameters included plant height at harvesting/cutting, leaf area, number of leaves per plant, chlorophyll content, fresh weight per plant, and nutrient composition. Since the focus of SP is on the crops, fish were only measured to monitor their relative growth in terms of weight gained at harvesting/cutting time.Measurement of crop parametersPlants were cut 5 cm above the soil surface, and agronomical trait measurements from a representative sample of 12 plants per replicate were taken as follows.Plant heights were taken using a foot ruler and averages determined. Leaf number was obtained as the number of leaves counted per plant and averages determined. Leaf area was calculated according to the equation reported by Yeshitila and Taye16.$${text{Leaf}} , {text{ Area }}left( {{text{cm}}^{{2}} } right) = , – {422}.{973} + { 22}.{752}0{text{L }}left( {{text{cm}}} right) , + { 8}.{text{31W }}left( {{text{cm}}} right)$$where L and W represent the leaf length and Leaf width respectively, − 422.973 is a constant relating to the shape of the leaf of Swiss chard developed by the author under citation.Chlorophyll content was measured using MC-100 chlorophyll meter from Apogee Instruments, Inc, and data was expressed as SPAD averages. Fresh weight was measured using a digital weighing balance and data expressed as g/plant.Sand testSand samples were obtained and sent for analysis at the Soils, Water and Environment Research Institute, Agricultural Research Center, Giza, Egypt. The Electrical conductivity (EC) values were measured from the sand paste extract; pH values were taken from sand suspensions at ratio of 1:2.5 as described by Estefan17. The available nitrogen in the sand sample was extracted using potassium chloride (KCl) as an extractable solution with the ratio of (5gm sand to 50 ml KCl) and determined using the micro- kjeldahl method. Available potassium was determined using a flame photometer, and the other elements in the sand sample were determined by using inductively coupled plasma (ICP) Spectrometry (model Ultima 2 JY Plasma)18,19. The physical and chemical characteristics of the used sand are presented in Table 3.Table 3 (a): Chemical analysis of field sand samples, (b): Available macro, micronutrients, and heavy metals content of the sand samples.Full size tableWater analysisEvery 15 days, a measured amount of desalinated water was added to a standard mark of 850L in the fish tanks to compensate for the consumed amount of water in the system. Fish water quality parameters such as water temperature, pH, and dissolved oxygen (DO) was closely monitored using automated digital Nilebot technologies by Conative labs to fit the ideal required levels as reported by Somerville et al.20. In contrast, ammonia, nitrite, and nitrate were adjusted using an API test kit every week. These parameters’ recorded values were as follows: water temperature ranged between 25 and 28 °C, DO range between 6–7 mg/L, and pH between 6.5 and 7.0. Ammonia levels were kept below 1 mg/L. Elements in water samples were determined according to EPA methods18 using inductively coupled plasma (ICP) Spectrometry (model Ultima 2 JY Plasma) as presented in Table 4.Table 4 Water sample analysis for the different systems’ fish tanks and sump tanks.Full size tableNutritive composition analysisAccording to Official methods of analysis from the association of official analytical chemists (A.O.A.C) (1990), moisture content and Vitamin C were determined. Vitamin A was determined according to the procedures described by Aremu and Nweze21. Briefly, 100 g of the sample were homogenized, from which 1 g was obtained and soaked in 5 mL methanol for two hours at room temperature in the dark for complete extraction of a pro-vitamin A carotenoid, β-carotene. Separation of the β-carotene layer was achieved through the addition of hexane to the sample, and moisture was removed using sodium sulphonate. The absorbance of the layer was measured at 436 nm using hexane as a blank. β-carotene was calculated using the formula:$$beta {text{-carotene }}left( {{mu g}/{1}00{text{ g}}} right) , = {text{ Absorbance }}left( {text{436 nm}} right) , times {text{ V }} times {text{ D }} times { 1}00 , times { 1}00/{text{W }} times {text{ Y}}$$where: V = total volume of the extract; D = Dilution factor; W = Sample weight; Y = Percentage dry matter content of the sample.Vitamin A was then determined according to the concept of Retinol Equivalent (RE) of the β-carotene content of the vegetables using the standard conversion formula. Total hydrolyzable carbohydrates were determined as glucose using phenol–sulfuric acid reagent as described by Michel22.Vitamin C content was determined using dichlorophenol indophenol reagent. As such, 10 g of fresh leaf tissues, were crushed using a motor and pestle in the presence of 10 ml metaphosphoric acid 6% (Merck). This was followed by centrifugation at 4000×g for 5 min at 4 °C. Five mL of the supernatant were transferred into an Erlenmeyer flask, and 20 mL of 3% metaphosphoric acid were added. The extract was titrated by dichlorophenol indophenol (Sigma-Aldrich) until a rose color was observed. Vitamin C (mg/100 g FW) was then calculated and based on the standard curve of l-Ascorbic acid (Merck) concentrations.For the determination of protein and mineral content, 0.5 g of dried samples were digested using sulfuric acid (H2SO4) and hydrogen peroxide (H2O2) as described by Cottenie23. From the extracted sample, the following minerals were determined:Nitrogen was determined according to the procedures described by Plummer24. Briefly, 5 mL of the digestive solution was distilled with 10 mL of sodium hydroxide (NaOH) for 10 min to obtain ammonia. Back titration was then used to determine the amount of nitrogen present in ammonia. Protein content was calculated by multiplying total nitrogen by 6.25 according to methods of AOAC25.Phosphorus content was determined calorimetrically (660 nm) according to the procedures described by Jackson26. Potassium, Calcium, and Sodium were determined against a standard using a flame-photometer (JEN way flame photometer) as described by Piper27. Magnesium (Mg), Copper (Cu), Manganese (Mn), Zinc (Zn), and Iron (Fe) content were determined using Atomic Absorption Spectrophotometer, Pyeunican SP1900, according to methods described by Liu28.The moisture percentage of leaf samples was determined by weighing the fresh weight for each sample (Fw), then dried for 72 h at 80 °C. The dry matter weight was record as Dw. The leaf water content was then calculated as the following:$${text{Moisture}};{text{ content }}left( % right) , = , left( {{text{Fw}} – {text{Dw}}} right) , /{text{ Fw}} * {1}00$$Statistical analysisStatistical comparisons among means of more than two groups were performed with analysis of variance (ANOVA) using SPSS V22, and the difference in means was analyzed by Tukey’s test at α = 0.05. Statistical differences were considered significant at P ≤ 0.05 in triplicates and data expressed as mean ± S.D.Plant materialAll plant materials and related procedures in this study were done in accordance with the guidelines of the Institutional Review Board of the American University in Cairo and the Ministry of Agriculture and Land Reclamation in Egypt.Ethics approvalThis study followed the guidelines and approval of Committee of Animal Welfare and Research Ethics, Faculty of Agriculture, Kafrelsheikh University, Egypt. More