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This study explored the potential application of polyphenols from Pinus halepensis bark in leather tanning. The primary objective was to characterize these polyphenols. The extraction and atomization processes proved efficient, reducing moisture content to 7.4%, increasing tannin content from 26.2% to 45.1%, and reducing insoluble substances by 77.5%. High-performance liquid chromatography (HPLC) coupled with mass spectrometry identified and quantified various polyphenolic compounds, including (+)-catechin, (+)-taxifolin, protocatechuic acid, and procyanidin B2. Notably, tannic catechin dimers were detected. Lignin was effectively removed through filtration. Concentrations of protocatechuic acid, (+)-catechin, (+)-taxifolin, (−)-epicatechin, and procyanidin B1 were significantly higher in the extract than in the powder, with the extract showing 1214.3 mg/kg of protocatechuic acid, 2098.0 mg/kg of (+)-catechin, 4017.0 mg/kg of (+)-taxifolin, 2163.0 mg/kg of (−)-epicatechin, and 917.0 mg/kg of procyanidin B1. [ABSTRACT FROM AUTHOR]

The present study aimed to determine the influence of organic amendments (OAs) on neutralizing the harmful effect of copper (Cu) on black mustard (Sinapis nigra L.). In a pot experiment, three levels of copper pollution were used: 200, 400, and 600 mg Cu kg−1, against a control without Cu. The soil was amended with three types of OAs: pine bark (PB), peat moss (PM), and cattle manure (CM). Our research showed that plant condition depends on the Cu content in the soil. Increasing soil contamination significantly affected the plant yield, leaf greenness index, and dry matter content. The type of OA had no significant effect on the condition of black mustard (BM); however, each had a different effect on neutralizing the harmful effects of Cu. CM reduced Cu accumulation, PM showed no effect, while PB contributed to a significant increase in Cu content in BM plants. The chemical composition of BM depended on the Cu content in the soil. With increased soil contamination with Cu, the contents of Ntot, K, Mg, Ca, and Na in BM increased, while the content of P decreased. In terms of mitigating the harmful effects, CM was more beneficial than PM and PB. Among the analyzed OAs, CM, and PM contributed to Cu immobilization, while PB promoted Cu mobilization in contaminated soils. [ABSTRACT FROM AUTHOR]

Phosphorus (P) fertilizers applied to container-grown nursery crops readily leach through pine bark-based substrates and can subsequently runoff and contribute to surface water contamination. The objectives of this research were to determine the effect of adding a layer of FeSO4·7H2O-amended pine bark (FSB) to the bottoms of nursery containers on P leaching characteristics. Phosphorus and iron (Fe) leaching in response to FSB layer height (4 or 7.5 cm), FeSO4·7H2O rate (0.3, 0.6, or 1.2 kg·m−3 Fe), and form (i.e., granular versus liquid) used to formulate the FSB layer, and the inclusion of dolomite in the FSB layer were also investigated. Greenhouse studies lasting 15 and 19 weeks were conducted, in which 2.5 L nursery containers containing the FSB layer treatments below non-amended pine bark substrate were fertilized with 199 or 117 mg P from a soluble or controlled-release fertilizer, respectively. Leachate resulting from daily irrigation was collected and analyzed for P and Fe weekly. All FSB treatments leached less P than the control (non-amended pine bark only), with P reductions ranging from 22% (4 cm FSB with 0.3 kg·m−3 Fe) to 73% (7.5 cm FSB with 1.2 kg·m−3 Fe). Phosphorus leaching decreased linearly with an increase in Fe rate or layer height. The amount of Fe that leached from containers with FSB was <5% of that applied, regardless of the Fe rate. Granular- and liquid-applied FeSO4·7H2O with or without dolomite were equally effective at reducing P leaching. Adding 0.6 kg·m−3 Fe to the bottom 500 cm3 of pine bark increased P adsorption by 0.053 mg·cm−3 P, which equates to 17.9 mg P adsorbed per gram of FeSO4·7H2O added. Results from this research suggest that including an FSB layer in the bottom of nursery containers is an effective strategy for reducing P runoff from container-based nursery production sites. [ABSTRACT FROM AUTHOR]

Greenhouse tomato production faces multiple challenges, including the excessive use of nonrenewable substrates that are difficult to dispose of after use. Currently, most growers propagate tomatoes in rockwool, but there is an increasing demand for sustainable media. The objective of this research was to evaluate sustainable and organic alternatives for greenhouse propagation of tomato seedlings intended for high-wire production. Different organic and inorganic substrates were evaluated in three experiments, using a nutrient solution composed of a complete water-soluble fertilizer. Germination and growth parameters, including height, stem diameter, number of leaves, leaf area, foliar chlorophyll levels (SPAD), and shoot fresh and dry weight, were measured. In the first experiment, which employed overhead irrigation, rockwool, coir, wood fiber–coir mix, medium-grade pine bark, pine bark < 0.64 cm, and pine bark < 0.32 cm were evaluated. Tomato germination was faster and achieved higher percentages with pine bark < 0.64 cm compared to other substrates. However, growth performance was similar or better in coir than in rockwool four weeks after transplantation. For the second experiment with sub-irrigation only, rockwool, coir, wood fiber–coir mix, pine bark < 0.32 cm bark, and peat were evaluated at different container heights. Peat resulted in greater growth across all parameters, followed by wood fiber–coir mix in all container heights, while pine bark had the least growth across all measured parameters. In the third experiment with overhead irrigation, rockwool, wood fiber–coir mix, pine bark < 0.32 cm, and a commercial peat-based mixture were evaluated under different fertilizer rates (electrical conductivity of 1.1 and 2.2 mS·cm−1). Wood fiber–coir mix, peat-based mix, and rockwool were the substrates with the highest values for all evaluated parameters. While all the organic substrates showed potential for use in tomato propagation, pine bark < 0.32 cm bark and wood fiber–coir mix provided the best media for germination. Peat and wood fiber–coir mix showed the best media for subsequent seedling growth and demonstrated potential to be used as substitutes for rockwool. [ABSTRACT FROM AUTHOR]

Hexavalent chromium (Cr(VI)) is one of the most toxic heavy metals found in industrial wastewater, so many researchers are working to develop efficient and environmentally friendly removal methods. It has been reported that natural biomass and its derivatives can be used to treat wastewaters containing Cr(VI). However, biomass with sufficient Cr(VI) removal performance to replace the existing chemical method, which is cheap and simple, has not been reported yet. This study reports that inexpensive, abundant, and commercially available pine bark has the highest Cr(VI) removal capacity (i.e., 376.3 mg/g) compared to biomass reported elsewhere. This value is six times higher than the theoretical value of an inorganic reducing agent (iron(II) sulfate heptahydrate). The main mechanism of Cr(VI) removal by pine bark was clearly identified through kinetic experiments, Fourier-transform infrared spectrometer, and X-ray photoelectron spectroscopy analyses, which were used to study the compositions, functional groups, and bonding states of pine bark. It was found that pine bark consists of various acidic functional groups that can act as electron donors to promote the removal of Cr(VI) through redox reactions. In conclusion, pine bark may be a promising candidate for the removal of Cr(VI) from wastewater, owing to its excellent removal capacity. [ABSTRACT FROM AUTHOR]

Alzheimer's disease is the most common type of dementia with cognitive impairment. Various plant-derived phenolics are known to alleviate cognitive impairment in Alzheimer's disease by radical scavenging and strengthening synaptic plasticity activities. Here, we examined the cognition-improving effect of Pinus densiflora Sieb. et Zucc. bark extract (PBE). We identified and quantified phenolics in the PBE using a UHPLC-Orbitrap mass spectrometer. To evaluate the cognition-enhancing effects of PBE, scopolamine-induced amnesic Sprague-Dawley (SD) rats (5 weeks old) and ion channel antagonist-induced organotypic hippocampal slices of SD rats (7 days old) were used. Twenty-three phenolics were tentatively identified in PBE, 10 of which were quantified. Oral administration of PBE to the scopolamine-induced SD rats improved cognitive impairment in behavioral tests. PBE-fed SD rats showed significantly improved antioxidant indices (superoxide dismutase and catalase activities, and malondialdehyde content) and reduced acetylcholinesterase activity in hippocampal lysate compared with the scopolamine group. PBE increased the long-term potentiation (LTP) induction and rescued LTP from blockades by the muscarinic cholinergic receptor antagonist (scopolamine) and N-methyl-D-aspartate channel antagonist (2-amino-5-phosphonovaleric acid) in the organotypic hippocampal slices. These results suggest that polyphenol-rich PBE is applicable as a cognition-improving agent due to its antioxidant properties and enhancement of LTP induction. [ABSTRACT FROM AUTHOR]

Soilless substrate stratification (i.e., layering unique substrates within a single container) is an emerging substrate management strategy that may provide opportunities to augment nursery resource use. As such, this research aimed to analyze water movement through containers during hydration events under different initial moisture conditions. The results indicated substrate stratification had minimal influence on water movement compared to non-stratified systems (uniformly filled nursery containers). Cyclic irrigation significantly increased the stratified substrates' ability to retain water when irrigated at 20% volumetric water content (p < 0.0001) and significantly decreased the total volume leached (p < 0.0001). Moreover, irrigating the substrate profile with shallow and more frequent irrigations facilitated stratified substrates ty reach effective container capacity conditions (p < 0.0001n compared to non-stratified systems. The stratified systems took longer to leach all gravitational pores (p = 0.0266). In dry moisture conditions, non-stratified substrates were more hydrated when cyclic irrigation applications were applied compared to single applications (p = 0.0492). This study demonstrated that cyclic irrigation scheduling enhanced water retention in both non-stratified and stratified profiles under different initial moisture conditions and can be used as an irrigation strategy when dry substrate conditions prevail. [ABSTRACT FROM AUTHOR]

Hydrangea macrophylla exhibiting blue sepals (versus purple or pink) have improved marketability; however, little research has been conducted to evaluate aluminum (Al), the element responsible for bluing, on crop growth, effectiveness of bluing sepals, and characteristics of flower clusters in an outdoor nursery. This study compared substrate Al availability, crop growth, flower color, number, and size over a 56-week period in two locations. A polymer coated (90-day release) or ground aluminum sulfate [Al2(SO4)3; water soluble] was either incorporated into a non-limed pine bark substrate, applied to the surface of the substrate as a top dress, or as a routinely applied Al2(SO4)3 drench (low concn.) or applied once (high concn.). In general, application of Al increased plant foliar Al concentration, but also decreased substrate pore-water pH and increased electrical conductivity (EC) with varying effects based on the applied product's solubility and subsequent longevity. Aluminum sulfate increased the potential of Al phytotoxicity negatively affecting root morphology and creating an undesirable rhizosphere electrochemistry due to the pH being continually acidic, <4, and the EC being temporarily increased to >1.5 mS·cm−1. These suboptimal rhizosphere conditions resulted in a lower quality or smaller plant. No plants exhibited clear, deep blue flower cluster sought by consumers. Neither the effect of pore water pH or EC could, alone or in combination, account for the lack of plant vigor or blue flower clusters when substrate and foliar Al concentrations were adequate in flowering H. macrophylla. More research is needed to investigate the effect of pore-water electrochemical properties, possible mineral nutrient co-factors that provide Al synergisms or toxicity protections, and holistic plant health on ensuring blue coloration of a vigorous H. macrophylla. [ABSTRACT FROM AUTHOR]

Lignocellulosic biomass is an abundant resource that can be valorized for the production of bioenergy. However, studies aiming to quantify the amount of biogas production potential per km forest road are scarce in the literature. In this study, fresh pine needles, pine needle litter, pine branches, and pine bark were digested in batch reactors under mesophilic conditions after a grinding/milling pre-treatment. All samples were collected from a low-altitude Mediterranean Pinus forest (North Greece) adjacent to a category G forest road with a gentle slope. The methane yield of fresh pine needles was between 115 and 164 NmL g−1 volatile solids (VS), depending on the Pinus tree size. Pine needle litter produced a significantly lower methane yield (between 58 and 77 NmL g−1 VS), followed by pine bark (85 NmL g−1 VS) and pine branches (138 NmL g−1 VS). Considering the quantity of pine needle litter accumulated on adjacent forest roads (600 ± 200 g m−2), it was possible to calculate the biomethane production potential per km of forest road (up to 500 Nm3 km−1) if the biomass collected was disposed of at an anaerobic digestion facility. The results of the study demonstrate that residual forest biomass represents an additional resource for bioenergy production. Moreover, harvesting residual forest biomass can decrease the incidence of devastating summer forest fires and their disastrous consequences for the environment, the economy, and the local populations. [ABSTRACT FROM AUTHOR]

The chemical composition of Pinus roxburghii bark essential oil (PRO) was qualitatively and quantitatively determined using GC/FID and GC/MS. The anti-inflammatory activity was assessed in vitro by evaluating the binding percentages on the cannabinoids and opioids receptors. Bleomycin (BLM)-induced pulmonary inflammation in albino mice was adopted to assess PRO anti-inflammatory efficacy in vivo. In silico molecular modelling of its major components was performed on human glucocorticoids receptor (GR). Seventy-five components were identified in which longifolene (33.13%) and palmitic acid (9.34%) constituted the predominant components. No binding was observed on cannabinoid receptor type 1 (CB1), whereas mild binding was observed on cannabinoid receptor type 2 (CB2), delta, kappa, and mu receptors accounting for 2.9%, 6.9%, 10.9% and 22% binding. A significant in vivo activity was evidenced by reduction of the elevated malondialdehyde (MDA), nitric oxide (NO), myeloperoxidase (MPO), interleukin-6 (IL-6), and tumor necrosis factor-a (TNF-a) levels by 55.56%, 55.66%, 64.64%, 58.85% and 77.78% with concomitant elevation of superoxide dismutase (SOD) and catalase (CAT) activities comparable to BLM-treated group at 100 mg/kg body weight. In silico studies showed that palmitic acid exerted the fittest binding. PRO could serve as a potent anti-inflammatory natural candidate that should be supported by further clinical trials. [ABSTRACT FROM AUTHOR]

The consumption of polyphenols has frequently been associated with low incidence of degenerative diseases. Most of these natural antioxidants come from fruits, vegetables, spices, grains and herbs. For this reason, there has been increasing interest in identifying plant extract compounds. Polymeric tannins and monomeric flavonoids, such as catechin and epicatechin, in pine bark and green tea extracts could be responsible for the higher antioxidant activities of these extracts. The aim of the present study was to characterize the phenolic compounds in pine bark and green tea concentrated extracts using high-performance liquid chromatography coupled to electrospray ionization mass spectrometry (HPLC-ESI-QTOF-MS). A total of 37 and 35 compounds from pine bark and green tea extracts, respectively, were identified as belonging to various structural classes, mainly flavan-3-ol and its derivatives (including procyanidins). The antioxidant capacity of both extracts was evaluated by three complementary antioxidant activity methods: Trolox equivalent antioxidant capacity (TEAC), ferric reducing antioxidant power (FRAP) and oxygen radical absorbance capacity (ORAC). Higher antioxidant activity values by each method were obtained. In addition, total polyphenol and flavan-3-ol contents, which were determined by Folin-Ciocalteu and vanillin assays, respectively, exhibited higher amounts of gallic acid and (+)-catechin equivalents. [ABSTRACT FROM AUTHOR]

Nurseries rely on soilless substrates to provide suitable growing media for container grown crops. These soilless substrates have been developed to readily drain water to prevent issues with waterlogging and associated soil-borne disease. A negative consequence of high porosity and subsequent drainage throughout the container profile is the required high or frequent irrigation rates with poor retention of applied nutrients. Substrates with relatively high levels of moisture and nutrient retention placed on top of a coarse and freely draining substrate could further optimize water and nutrient retention, while allowing for needed gas exchange for plant establishment and growth. Containerized Red Drift® rose (Rosa 'Meigalpio' PP17877) plants were grown under 16 mm or 12 mm daily irrigation, utilizing a traditional pine bark substrate or stratified substrates with either a conventional bark, bark fines, or a bark–peat mixture on top of a coarse bark within a container. The stratified substrates received 20% less controlled-release fertilizer; however, the fertilizer in the stratified treatments was concentrated in the upper strata only. During the first growing phase or season, plants grown in stratified substrates outperformed those grown in conventional, non-stratified bark substrates under normal irrigation. The stratified substrates did not reduce growth under reduced irrigation regimes. Overall, crop growth was equal or superior for stratified substrates when compared to the non-stratified controls, even with a 20% reduction of fertilizer. This research suggests that stratified substrate systems can be used to reduce fertilizer and irrigation rates while producing crops of similar or superior quality to conventionally grown containerized crops. [ABSTRACT FROM AUTHOR]

Particles in a substrate create a network of pore pathways for water to move through, with size and shape determining the efficacy of these channels. Reduced particle size diversity can lead to increased leachate, poor substrate hydration, and an inefficient irrigation practice. This research examined the hydration characteristics of three greenhouse substrate components at three preconditioned initial moisture contents using subirrigation under five different irrigation durations and three water depths (2 mm, 20 mm, and 35 mm). Sphagnum peatmoss, coconut coir, and aged pine bark were tested at 67%, 50%, and 33% initial moisture (by weight). The objectives were to determine the impact of varying irrigation event durations (5, 10, 20, 30, 60 min) over a 60-min period, and the further influence of water depth and initial moisture, on the water capture abilities of peat, coir, and pine bark. The number of irrigation events depended on the irrigation event time of that experimental unit divided by the total time of 60 min, varying from 12, 6, 3, 2, and 1 event. Hydration efficiency was influenced by initial moisture content (IMC), water depth, pulsing duration, and inherent substrate characteristics (hydrophobicity/hydrophilicity). Initial MC had the largest impact on peat, regardless of water level or irrigation duration. Lower IMCs increased the hydrophobic response of peat, further reducing the amount of water the substrate was able to absorb. Pine bark had a 5–10% decrease in initial hydration between 67%, 50%, and 33% IMC, while coir's hydrophilic nature reduced any IMC affects. At 50% IMC or less, coir had the highest volumetric water content (VWC) across all substrates, pulsing durations, and water depths. Water depth was found to increase initial hydration and final hydration 6–8% across all substrates. These three materials had altered and varied water capture responses depending on the combination of treatments employed. This work demonstrated the effects of intensity and exposure on substrates and the need for more integrated research for improving water use efficiency on container crops. [ABSTRACT FROM AUTHOR]

Environmental pollution due to antibiotics is a serious problem. In this work, the adsorption and desorption of the antibiotic cefuroxime (CFX) were studied in four by-products/residues from the forestry and food industries. For this, batch-type experiments were carried out, adding increasing concentrations of CFX (from 0 to 50 µmol L−1) to 0.5 g of adsorbent. The materials with a pH higher than 9 (mussel shell and wood ash) were those that presented the highest adsorption percentages, from 71.2% (23.1 µmol kg−1) to 98.6% (928.0 µmol kg−1). For the rest of the adsorbents, the adsorption was also around 100% when the lowest concentrations of CFX were added, but the percentage dropped sharply when the highest dose of the antibiotic was incorporated. Adsorption data fitted well to the Langmuir and Freundlich models, with R2 greater than 0.9. Regarding desorption, the materials that presented the lowest values when the highest concentration of CFX was added were wood ash (0%) and mussel shell (2.1%), while pine bark and eucalyptus leaves presented the highest desorption (26.6% and 28.6%, respectively). Therefore, wood ash and mussel shell could be considered adsorbents with a high potential to be used in problems of environmental contamination by CFX. [ABSTRACT FROM AUTHOR]

The recovery of critical and precious metals from waste electrical and electronic equipment (WEEE) is an environmental and economic imperative. Biosorption has been considered a key technology for the selective extraction of gold from hydrometallurgical liquors obtained in the chemical leaching of e-waste. In this work, the potential of tannin resins prepared from Pinus pinaster bark to sequester and recover gold(III) from hydrochloric acid and aqua regia solutions was assessed. Equilibrium isotherms were experimentally determined and maximum adsorption capacities of 343 ± 38 and 270 ± 19 mg g−1 were found for Au uptake from HCl and HCl/HNO3 (3:1 v/v) solutions containing 1.0 mol L−1 H+. Higher levels of acidity (and chloride ligands) significantly impaired the adsorption of gold from both kinds of leaching solutions, especially in the aqua regia system, in which the adsorbent underperformed. Pseudo-first and pseudo-second order models successfully described the kinetic data. The adsorbent presented high selectivity towards gold. Actually, in simulated aqua regia WEEE liquors, Au(III) was extensively adsorbed, compared to Cu(II), Fe(III), Ni(II), Pd(II), and Zn(II). In three adsorption–desorption cycles, the adsorption capacity of the regenerated adsorbent moderately decreased (19%), although the gold elution in acidic thiourea solution had been quite limited. Future research is needed to examine more closely the elution of gold from the exhausted adsorbents. The results obtained in this work show good perspectives as regards the application of pine bark tannin resins for the selective extraction of Au from electronic waste leach liquors. [ABSTRACT FROM AUTHOR]

Plant water availability in soilless substrates is an important management consideration to maximize water efficiency for containerized crops. Changes in the characteristics (i.e., shrink) of these substrates at low water potential (<−1.0 MPa) when using a conventional pressure plate-base can reduce hydraulic connectivity between the plate and the substrate sample resulting in inaccurate measures of water retention. Soilless substrate components Sphagnum peatmoss, coconut coir, aged pine bark, shredded pine wood, pine wood chips, and two substrate composites were tested to determine the range of volumetric water content (VWC) of surface-bound water at water potentials between −1.0 to −2.0 MPa. Substrate water potentials were measured utilizing dewpoint hygrometry. The VWC for all components or composites was between 5% and 14%. These results were considerably lower compared to previous research (25% to 35% VWC) utilizing conventional pressure plate extraction techniques. This suggests that pressure plate measurements may overestimate this surface-bound water which is generally considered unavailable for plant uptake. This would result in underestimating available water by as much as 50%. [ABSTRACT FROM AUTHOR]

Substrate wettability is an important factor in determining effective and efficient irrigation techniques for container-grown crops. Reduced substrate wettability can lead to lower substrate water capture, excessive leaching and poor plant growth. This research examined substrate water capture using surface and subirrigation under three initial moisture contents (IMC). Sphagnum peat moss, coconut coir, and pine bark were tested at IMCs of 67% 50%, and 33%. Substrate water capture was influenced by both IMC and irrigation technique. Surface irrigation increased the water capture of coir and peat, regardless of IMC, whereas IMC influenced pine bark water capture more than irrigation method. Surface-irrigated coir at or above 50% IMC provided the greatest water capture across all treatments. The first irrigation had the highest capture rate compared to all other events combined. Container capacities of pine bark and coir were unaffected by IMC and irrigation type, but the CC of peat was less by ~ 40% volumetrically under low IMC conditions. Coir, had the greatest ability to capture water, followed by pine bark and peat, respectively. Moisture content, irrigation type and component selection all influence the water capture efficiency of a container substrate. [ABSTRACT FROM AUTHOR]

Many achievements have been made on the research of composite polyurethane foams to improve their structure and mechanical properties, and the composite foams have been widely utilized in building insulation and furniture. In this work, rigid polyurethane foams (RPUFs) with the addition of different fillers (nano-SiO2, peanut shell, pine bark) were prepared through the one-step method. The effects of inorganic nano-SiO2 and organic biomass on foam properties were evaluated by means of physical and chemical characterization. The characterization results indicate that the compressive strength values of prepared foams could fully meet the specification requirement for the building insulation materials. The inorganic and organic fillers have no effect on the hydrogen bonding states in composite RPUFs. Furthermore, compared to the biomass fillers, the addition of nano-SiO2 greatly influenced the final residual content of the fabricated foam. All composite foams exhibit closed-cell structure with smaller cell size in comparison with the parent foam. The prepared composite foams have the potential for utilization in building insulation. [ABSTRACT FROM AUTHOR]

Aiming to investigate the efficacy of different materials as bio-sorbents for the purification of As-polluted waters, batch-type experiments were employed to study As(V) sorption and desorption on oak ash, pine bark, hemp waste, mussel shell, pyritic material, and soil samples, as a function of the As(V) concentration added. Pyritic material and oak ash showed high sorption (90% and >87%) and low desorption (<2% and <7%). Alternatively, hemp waste showed low retention (16% sorption and 100% desorption of the amount previously sorbed), fine shell and pine bark sorbed <3% and desorbed 100%, the vineyard soil sample sorbed 8% and released 85%, and the forest soil sample sorbed 32% and desorbed 38%. Sorption data fitted well to the Langmuir and Freundlich models in the case of both soil samples and the pyritic material, but only to the Freundlich equation in the case of the various by-products. These results indicate that the pyritic material and oak ash can be considered efficient As(V) sorbents (thus, useful in remediation of contaminated sites and removal of that pollutant), even when As(V) concentrations up to 6 mmol L -1 are added, while the other materials that were tested cannot retain or remove As(V) from polluted media., Competing Interests: The authors declare no conflict of interest. The founding sponsors had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, and in the decision to publish the results.