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BACKGROUND: During the processing of spent coffee grounds (SCGs) several residues are obtained, which are mostly disposed of in landfills. There is an urgent need for a comprehensive waste management strategy for these residues. This study evaluates the potential of SCGs as a biofertilizer by assessing their effects on lettuce leaves and the release of antioxidants following in vitro digestion and fermentation. RESULTS: Lettuce plants were grown with different amounts of SCGs (0-150 g kg-1) in the substrate. High SCG concentrations in the soil generated lighter colored tissues, a decrease in the green color, less root development, and lower dry weight of leaves (P < 0.05). The SCG levels also affected the release of antioxidants by the final product. This effect was more pronounced in the digested fraction: applying the Ferric Reducing Antioxidant Power (FRAP) method, the addition of SCGs from 10 g kg-1 to 125 g kg-1 increased the amount of antioxidant from 43.88 ± 4.81 to 105.96 ± 29.09 µmol Trolox g-1 of dry weight (P < 0.05). The Indigo Carmine Reducing Capacity (ICRED) method also showed a similar trend, but in this case the highest value was obtained with 150 g kg-1 of SCGs (16.41 ± 3.93 mmol catechin g-1 of dry weight) (P < 0.05). Moreover, in the fermented fraction a significant increase in the antioxidant released was found with low levels of SCG(P<0.05), while lettuces fertilized with intermediate amounts of SCGs (25 and 50 g kg-1) presented the highest amount of insoluble antioxidant (P < 0.05). CONCLUSION: A compromise should be found in order to achieve a product with a high antioxidant capacity and an acceptable visual quality. © 2024 Society of Chemical Industry.

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In recent years, natural alternatives have been sought for the control of beekeeping pathologies; in the case of American Foulbrood (AFB) disease, the use of synthetic antibiotics was prohibited due to honey contamination and the generation of resistant bacteria. The significant increase in population growth worldwide has led to great concern about the production of large amounts of waste, including those from agribusiness. Among the most important beverages consumed is coffee, generating thousands of tons of waste called spent coffee grounds (SCG). The SCG is a source of many bioactive compounds with known antimicrobial activity. The aims of the present work were: (1) to obtain and chemically analyse by HPLC of SCG extracts (SCGE), (2) to analyse the antimicrobial activity of SCGE against vegetative form of Paenibacillus larvae (the causal agent of AFB), (3) to evaluate the toxicity in bees of SCGE and (4) to analyse the effect of the extracts on the expression of various genes of the immune system of bees. SCGs have a high content of phenolic compounds, and the caffeine concentration was of 0.3%. The MIC value obtained was 166.667 µg/mL; the extract was not toxic to bees, and interestingly, overexpression of abaecin and hymenoptaecin peptides was observed. Thus, SCGE represents a promising alternative for application in the control of American Foulbrood and as a possible dietary supplement to strengthen the immune system of honeybees. Therefore, the concept of circular bio-economy could be applied from the coffee industry to the beekeeping industry.

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Supplementary cementitious materials are considered a viable and affordable way to reduce CO2 emissions from the cement industry's perspective since they can partially or nearly entirely replace ordinary Portland cement (OPC). This study compared the impact of adding spent coffee grounds (SCGs), fly ash (FA), and volcanic ash (VA) to two types of cement: OPC and calcium sulfoaluminate cement (CSA). Cement samples were characterized using compressive strength measurements (up to 210 days of curing), scanning electron microscopy with energy dispersive X-ray spectroscopy (SEM-EDS), X-ray diffraction (XRD), attenuated total reflection infrared spectroscopy, and hydration temperature measurements. In all the studied systems, the presence of SCGs reduced compressive strength and delayed the hydration process. CSA composite cement containing 3.5% SCGs, 30% FA, and 30% VA showed compressive strength values of 20.4 MPa and 20.3 MPa, respectively, meeting the minimum requirement for non-structural applications. Additionally, the results indicate a formation of cementitious gel, calcium silicate hydrate (C-S-H) in the OPC-based composite cements, and calcium alumino-silicate hydrate (C-A-S-H) as well as ettringite in the CSA-based composite cements.

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This work investigated the addition of spent coffee grounds (SCG) as a valuable resource to produce biocomposites based on polylactic acid (PLA). PLA has a positive biodegradation effect but generates poor proprieties, depending on its molecular structure. The PLA and SCG (0, 10, 20 and 30 wt.%) were mixed via twin-screw extrusion and molded by compression to determine the effect of composition on several properties, including mechanical (impact strength), physical (density and porosity), thermal (crystallinity and transition temperature) and rheological (melt and solid state). The PLA crystallinity was found to increase after processing and filler addition (34-70% in the 1st heating) due to a heterogeneous nucleation effect, leading to composites with lower glass transition temperature (1-3 °C) and higher stiffness (~15%). Moreover, the composites had lower density (1.29, 1.24 and 1.16 g/cm3) and toughness (30.2, 26.8 and 19.2 J/m) as the filler content increased, which is associated with the presence of rigid particles and residual extractives from SCG. In the melt state, polymeric chain mobility was enhanced, and composites with a higher filler content became less viscous. Overall, the composite with 20 wt.% SCG provided the most balanced properties being similar to or better than neat PLA but at a lower cost. This composite could be applied not only to replace conventional PLA products, such as packaging and 3D printing, but also to other applications requiring lower density and higher stiffness.

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BACKGROUND: Spent coffee grounds (SCGs) are a good source of chlorogenic acid (CGA), which can be hydrolyzed to quinic acid (QA) and caffeic acid (CA). These molecules have antioxidant and neuroprotective capacities, benefiting human health. The hydrolysis of CGA can be done by biotechnological processes, such as solid-state fermentation (SSF). This work evaluated the use of SSF with Aspergillus sp. for the joint release of the three molecules from SCGs. RESULTS: Hydroalcoholic extraction of the total phenolic compounds (TPCs) from SCGs was optimized, obtaining 28.9 ± 1.97 g gallic acid equivalent (GAE) kg-1 SCGs using 0.67 L ethanol per 1 L, a 1:9 solid/liquid ratio, and a 63 min extraction time. Subsequently, SSF was performed for 30 days, achieving the maximum yields for CGA, QA, and TPCs on the 16th day: 7.12 ± 0.01 g kg-1 , 4.68 ± 0.11 g kg-1 , and 54.96 ± 0.49 g GAE kg-1 respectively. CA reached its maximum value on the 23rd day, at 4.94 ± 0.04 g kg-1 . The maximum antioxidant capacity was 635.7 mmol Trolox equivalents kg-1 on the 14th day. Compared with unfermented SCGs extracts, TPCs and CGA increase their maximum values 2.3-fold, 18.6-fold for CA, 14.2 for QA, and 6.4-fold for antioxidant capacity. Additionally, different extracts' profiles were obtained throughout the SSF process, allowing us to adjust the type of enriched extract to be produced based on the SSF time. CONCLUSION: SSF represents an alternative to produce extracts with different compositions and, consequently, different antioxidant capacities, which is a potentially attractive fermentation process for different applications. © 2022 Society of Chemical Industry.

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Spent coffee grounds (SCG) are a current subject in many works since coffee is the second most consumed beverage worldwide; however, coffee generates a high amount of waste (SCG) and can cause environmental problems if not discarded properly. Therefore, several studies on SCG valorization have been published, highlighting its waste as a valuable resource for different applications, such as biofuel, energy, biopolymer precursors, and composite production. This review provides an overview of the works using SCG as biopolymer precursors and for polymer composite production. SCG are rich in carbohydrates, lipids, proteins, and minerals. In particular, carbohydrates (polysaccharides) can be extracted and fermented to synthesize lactic acid, succinic acid, or polyhydroxyalkanoate (PHA). On the other hand, it is possible to extract the coffee oil and to synthesize PHA from lipids. Moreover, SCG have been successfully used as a filler for composite production using different polymer matrices. The results show the reasonable mechanical, thermal, and rheological properties of SCG to support their applications, from food packaging to the automotive industry.

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As coffee consumption is on the rise, and the global coffee production creates an excess of 23 million tons of waste per year, a revolutionary transition towards a circular economy via the transformation and valorization of the main by-products from its cultivation and preparation (Coffee Husk (CH), Coffee Pulp (CP), Coffee Silverskin (CS), and Spent Coffee Grounds (SCG)) is inspiring researchers around the world. The recent growth of scholarly publications in the field and the emerging applications of coffee by-products published in these scientific papers encourages a systematic review to identify the knowledge structure, research hotspots, and to discuss the challenges and future directions. This paper displays a comprehensive scientometric analysis based on 108 articles with a high level of influence in the field of coffee by-products and their applications. According to our analysis, the research in this field shows an explosive growth since 2017, clustered in five core applications: bioactive compounds, microbial transformation, environmental applications, biofuels from thermochemical processes, and construction materials.

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OBJETIVO: El objetivo de este trabajo fue establecer el efecto de la borra de café sobre la movilidad y los parámetros funcionales de los espermatozoides humanos in vitro. MATERIALES Y MÉTODOS: La borra de café, un subproducto obtenido en establecimientos especializados en la preparación de café soluble a base de grano, se diluyo en tampón fosfato salino y se mezcló en proporciones iguales con las muestras de semen de 16 voluntarios aparentemente sanos. A cada muestra se le determinó el efecto sobre la movilidad espermática en función del tiempo (30, 60, 90 y 120 minutos, n=16) y sobre los parámetros funcionales (n=6) por medio de citometría de flujo: potencial de membrana mitocondrial, producción de especies reactivas de oxígeno y lipoperoxidación de la membrana espermática. RESULTADOS: La incubación de los espermatozoides con la borra de café evidencio un cambio positivo en la movilidad espermática. Adicionalmente, la incubación con la borra de café incremento significativamente el potencial de membrana mitocondrial en los espermatozoides. CONCLUSIÓN: La borra de café, seguramente debido a los compuestos antioxidantes, afecta positivamente la movilidad espermática aumentando el potencial de membrana mitocondrial. Por lo tanto, esto es un paso inicial en la búsqueda de un suplemento de origen natural que aumente la calidad seminal.

OBJECTIVE: The objective of this work is to establish the effect of spent coffee grounds on the motility and functional parameters of human spermatozoa, in vitro. MATERIALS AND METHODS: Spent coffee grounds, a by-product obtained in specialized establishments in the preparation of soluble coffee based on grain, was diluted in saline phosphate buffer and mixed in equal proportions with semen samples from 16 apparently healthy volunteers. Each sample was determined the effect on sperm motility as a function of time (30, 60, 90 and 120 minutes, n=16) and on functional parameters (n=6) by means of flow cytometry: mitochondrial membrane potential, reactive oxygen species production and membrane lipoperoxidation. RESULTS: The incubation of the spermatozoa with the spent coffee grounds showed a positive change in sperm motility. Additionally, incubation with spent coffee grounds significantly increased the mitochondrial membrane potential in human sperm cells. CONCLUSION: Spent coffee grounds, probably due to antioxidant compounds, positively affects sperm motility by increasing mitochondrial membrane potential. Therefore, this is an initial step in the search for a supplement of natural origin that increases seminal quality.

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Due to the increasing coffee production, Spent Coffee Grounds' (SCGs) generation has grown dramatically, hence appropriate management of this solid biomass waste is imperative. SCGs can be used as feedstocks for renewable energy and fuel generation provided that a stable feeding of powders to reactors is maintained. Recently, a non-mechanical spouted bed feeder proved itself an excellent alternative in feeding SCGs to a pilot-scale circulating fluidized bed reactor. Nonetheless, further studies are necessary for the feeder's implementation in commercial applications. Here the feeding of SCGs with the spouted bed feeder is addressed by using Computational Fluid Dynamics. Firstly, a Two-Fluid Model (TFM) is validated against experimental data, and then the effects of five operating and design parameters were analyzed aiming at improving the handling of SCGs. The solids flowrate (WS) in the reactor could be stably controlled from 4 to 30 g/s depending on the settings. The feeder performance is enhanced by operating it under high gas flowrate (Q), high entrainment length (z), and high mass of solids in the feeder (HS). Using feeders with low cone angle (γ) or reactors with large diameter (DR) increases WS, which is appealing for the operation of medium-to large-scale units. The proposed TFM is a cost-effective tool for implementing spouted bed feeders in commercial applications. With the feeder coupled to the process, SCGs are treated continuously in the reactor for energy generation, thus reducing the disposal problems associated with this waste and improving the management of SCGs globally.

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We evaluated the effects of acute intake of biscuits (B) containing either spent coffee grounds (SCG), (added with fructooligosaccharides; SC-FOS) or SCG antioxidant dietary fiber (SCF), on satiety, energy intake as well as gastrointestinal tolerance of healthy overweight volunteers. The addition of SCG and SCF to the biscuits (SC-FOS-B and SCF-B) increased their protein (11.4% and 12.2%), and total dietary fiber (8.4% and 11.8%) contents. The SCF-B significantly increased satiety perception, impacting ad libitum energy intake compared to the traditional biscuits recipe (TB; no added fiber). Moreover, SC-FOS-B and SCF-B decreased (no-observed-adverse effect) the most frequently reported symptoms by the participants, demonstrating that doses up to 5 g of SCF (per biscuit portion; 45 g) are well tolerated. The use of SCF and SCG as functional ingredients represents a sustainable strategy for the coffee industry and also potentiates the reduction of overweight, one of the leading health problems among the population. PRACTICAL APPLICATIONS: The addition of SCF or SCG as food ingredients increases protein and the dietary fiber content of traditional biscuits. SCF can slow gastric emptying, modulate appetite and thus body weight. SCF attenuates carbohydrate digestion blunting post-prandial blood glucose spikes reducing the risk of type 2 diabetes. SCF can be used as a functional ingredient to formulate foods with health benefits.

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Spent coffee grounds are wastes generated annually worldwide in significantly large amounts in the soluble coffee industry and in household and commercial beverage preparation. Although spent coffee grounds are rich in several classes of compounds, predominantly polysaccharides, profitable applications have not yet been effectively implemented for such wastes. Thus, it was the aim of this study to verify the feasibility of producing biopolymeric films from the polysaccharide-rich fraction of spent coffee grounds, obtained by alkaline hydrogen peroxide treatment of the coffee waste. Produced films were characterized for their physicochemical, barrier and mechanical properties and these properties were comparable to those of similar polysaccharides films from the literature.

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Spent coffee grounds (SCG) immobilized in agarose gel are proposed as a novel binding agent for application in the Diffusive Gradients in Thin films (DGT) technique for the determination of Cd, Cu, Ni, Pb and Zn in waters. The SCG-agarose gel was characterized by Scanning Electron Microscopy, Energy Dispersive X-ray Spectrometry and Porosimetry by nitrogen adsorption. Elution of analytes from the binding agent was effectively performed with 2 mol L-1 HCl. The effects of key DGT parameters (e.g. immersion time, ionic strength and pH) were evaluated with a deployment of DGT devices (DGT-SCG) in synthetic solutions with ionic strengths between 0.005 mol L-1 and 0.1 mol L-1 and within a pH range of 3.5-8.0. The results were in excellent agreement with the predicted theoretical curve for mass uptake. Consistent results were found for solutions with ionic strengths between 0.005 mol L-1 and 0.1 mol L-1 and within a pH range of 3.5-8.0. The DGT-SCG performance was also evaluated in two spiked river water samples (Corumbataí and Piracicaba river) with satisfactory uptake values (CDGT-SCG/Csol) between 0.74 and 1.53. The proposed DGT-SCG opens opportunities for using residual biomass as binding phase in the DGT technique, showing low costs in production and complying with "green" technology approaches.

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Brazilian coffee is well known worldwide due to its quality and richness in taste. The aim of the present study is to provide the elemental characterization of Brazilian coffee along different stages of the drip brewing process. To that end, samples from roasted ground coffee, spent coffee, paper filters and the final beverage were analyzed with one single ion beam technique, namely particle-induced X-ray emission (PIXE). In total, over 140 samples from 8 different Brazilian brands of ground coffee were analyzed. Large differences in some elemental concentrations were observed among different brands and among different batches of a single brand, which leads to high variances in the data. Concerning the beverage preparation, the analysis of the spent coffee shows that the transfer ratio from the ground coffee to the beverage differs for each element. Our results indicate that potassium and chlorine have the highest transfer ratio. Moreover, the concentration of rubidium is relatively high in drinking coffee. Finally, there is no influence of the elemental composition of paper filter in the preparation of drinking coffee.

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Calcium Alginate/Spent-Coffee-Grounds composite beads (CA-SCGs beads), which were made of two different proportions of alginate and spent-coffee-grounds (3:3 and 3:10), respectively, were used to adsorb Cu2+ in aqueous solution. These beads were compared with calcium alginate beads (CA beads) and spent-coffee-grounds (SCGs) in terms of adsorption capacity and rate of adsorption. The experiments were carried out at an initial pH of 4 at 30 °C with initial concentrations of Cu2+ from 10 ppm to 100 ppm. Equilibrium data was fitted with Langmuir, Freundlich and Sips models, and a pseudo-second-order kinetic equation. The Sips model showed the best correlation with the experimental values. CA-SCGs (3:3) beads showed a faster adsorption rate versus the CA beads. Also, CA-SCGs (3:3) beads showed a larger capacity of adsorption according to the Sips model, but not in the Langmuir model. FT-IR spectra and SEM images were taken for characterization. This study has shown that the CA-SCGs (3:3) beads have a synergistic effect, combining the capacity of adsorption of CA beads with the kinetics of the SCGs. The CA-SCGs beads have proven to be an effective adsorbent of Cu2+. Therefore, they can provide a use for the SCGs; which are considered pollutants in landfills.

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Antioxidant dietary fiber extracted from spent coffee grounds (FSCG) was evaluated as a potential functional food ingredient when incorporated in a food model (biscuits), and digested in vitro under simulated human gastrointestinal conditions. FSCG added to biscuits increased its total dietary fiber, antioxidant capacity after in vitro digestion, bioaccessibility of phenolic compounds (gallic acid and catechin) and amino acids. Furthermore, advanced glycation end products (AGEs), involved in chronic diseases, decreased up to 6-folds in the biscuits containing FSCG when compared with the traditional biscuit. The digestible fraction of biscuits containing the highest amount of FSCG (5 g) displayed the higher inhibiting α-glucosidase activity, correlating with the bioaccessibility of ascorbic acid and catechin. Our study seems to indicate that anti-diabetic compounds may be released in the small intestine during FSCG digestion, where biscuits containing FSCG may be able to beneficially regulate sugar metabolism thereby helping in producing foods friendly for diabetes.

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Coffee brew presents sensory, stimulatory and antioxidant properties highly appreciated by consumers, despite being associated with an increase in the level of blood cholesterol due to the effects of the diterpenes, especially cafestol, present in the lipid fraction. Although it is believed that the paper filter retains the brew diterpenes, new studies have shown that sometimes coffee filtered through paper can also increase the blood cholesterol level, putting in doubt the efficiency of the paper filter in retaining the diterpenes. Thus the objective of the present study was to verify the distribution of cafestol between the paper filter, the spent coffee and the coffee brew itself, from two coffee samples containing high and low cafestol contents selected from 13 samples of different cultivars and from different locations. In addition, the effect of the roasting degree on the cafestol contents of the roasted coffee was evaluated and the relationship between particle size of the roasted coffee and the extraction of solids. The highest cafestol content was found in the lightly roasted coffee, and the coffee brew presented higher solids contents when the particle size of the coffee powder was below 500µm. The results showed that of the initial cafestol concentration present in the roasted coffee, the paper filter retained 12.41%, the spent coffee 87.45% and the brew 0.15%. Thus, one can conclude that the greater part of the coffee cafestol is retained by the spent coffee, due to the low extraction of the lipid fraction by the hot water.

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Spent coffee grounds (SCG), rich in dietary fiber can be fermented by colon microbiota producing short-chain fatty acids (SCFAs) with the ability to prevent inflammation. We investigated SCG anti-inflammatory effects by evaluating its composition, phenolic compounds, and fermentability by the human gut flora, SCFAs production, nitric oxide and cytokine expression of the human gut fermented-unabsorbed-SCG (hgf-NDSCG) fraction in LPS-stimulated RAW 264.7 macrophages. SCG had higher total fiber content compared with coffee beans. Roasting level/intensity reduced total phenolic contents of SCG that influenced its colonic fermentation. Medium roasted hgf-NDSCG produced elevated SCFAs (61:22:17, acetate, propionate and butyrate) after prolonged (24h) fermentation, suppressed NO production (55%) in macrophages primarily by modulating IL-10, CCL-17, CXCL9, IL-1ß, and IL-5 cytokines. SCG exerts anti-inflammatory activity, mediated by SCFAs production from its dietary fiber, by reducing the release of inflammatory mediators, providing the basis for SCG use in the control/regulation of inflammatory disorders. The results support the use of SGC in the food industry as dietary fiber source with health benefits.

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The current study presents an application of Diffuse Reflectance Infrared Fourier Transform Spectroscopy for detection and quantification of fraudulent addition of commonly employed adulterants (spent coffee grounds, coffee husks, roasted corn and roasted barley) to roasted and ground coffee. Roasted coffee samples were intentionally blended with the adulterants (pure and mixed), with total adulteration levels ranging from 1% to 66% w/w. Partial Least Squares Regression (PLS) was used to relate the processed spectra to the mass fraction of adulterants and the model obtained provided reliable predictions of adulterations at levels as low as 1% w/w. A robust methodology was implemented that included the detection of outliers. High correlation coefficients (0.99 for calibration; 0.98 for validation) coupled with low degrees of error (1.23% for calibration; 2.67% for validation) confirmed that DRIFTS can be a valuable analytical tool for detection and quantification of adulteration in ground, roasted coffee.

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Introducción: Coffea arabica L., además de su importancia comercial, también se considera una planta medicinal, porque presenta propiedades biológicas diversas, pero por su comercialización como bebida genera muchos subproductos. La borra es uno de estos, que se obtiene por la preparación de la bebida, contiene una concentración significativa de compuestos polifenólicos y, por tanto, la recuperación de estas sustancias a partir de un residuo sin valor, sería potencialmente útil para la industria farmacéutica y alimentaria. Objetivos: recuperar compuestos fenólicos a partir de la borra de café y darle un valor agregado a un residuo de origen vegetal, como fuente de componentes con capacidad antirradicales libres in vitro. Métodos: la borra de café previamente secada se sometió a extracciones sólido-líquido usando diferentes sistemas de solventes. Se evaluaron las propiedades antioxidantes in vitro usando los métodos del catión radical del ácido 2,2´-azino-bis (3-etilbenzotiazoline-6-sulfónico) y del radical 1,1-difenil-2-picrilhidracilo. Se usó cromatografía líquida de alta resolución acoplada a espectrometría de masas para la caracterizaron de los principales componentes. Resultados: todos los extractos obtenidos mostraron buena capacidad antioxidante, con el extracto de etanol:agua como el mejor, seguido del extracto de metanol acidulado. Sin embargo, la capacidad antioxidante de la fracción en diclorometano del extracto etanol:agua resultó menor que la presentada por la taza de café. Se identificaron los ácidos clorogénico, isoclorogénico y feruloilquínico como los principales componentes de la borra de café. Conclusiones: todos los extractos presentaron buena capacidad protectora contra radicales libres. La borra de café, considerada un desecho obtenido del procesamiento industrial, se puede convertir en materia prima para la recuperación de sustancias antioxidantes; lo cual genera grandes expectativas sobre su posible uso en la industria farmacéutica y alimentaria, y le da al café un valor agregado importante.

Introduction: besides its commercial importance, Coffee arabica L. is also considered a medicinal plant due to its various biological properties, but its marketing produces a large amount of residues. Spent coffee grounds are one of these residues, which are obtained after the preparation of the drink and contain a significant concentration of polyphenolic compounds. Therefore, the recovery of these substances costless residual will be potentially useful for food and pharmaceutical industry. Objectives: to recover polyphenolic compounds from spent coffee grounds and to give an added value to a vegetable waste as a source of substances with free radicals capacity in vitro. Methods: dehydrated spent coffee grounds were subjected to solid-liquid extraction using different solvent systems. Antioxidant properties were evaluated in vitro using the radical monocation 2,2´-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS) and the stable free radical 2-2-diphenyl-1-picrylhydrazyl (DPPH). Liquid chromatography coupled to mass spectrometry was applied to characterize the main compounds. Results: all the extracts obtained showed a good antioxidant capacity with ethanol-water extract, followed by acidulated methanol extract. However, the antioxidant capacity of the ethanol-water extract was lower than coffee beverage. Chlorogenic, isochlorogenic and feruloylquinic acids were identified as the main compounds present in spent coffee grounds. Conclusions: all extracts showed a significant protection effect against free radicals; spent coffee ground, which is considered an undesirable solid waste from industrial processing, could be an add-value raw material in the recovery of antioxidant substances which generates great expectations about its possible use in the pharmaceutical and food industry and gives coffee an important added value.