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At least 145 records · Page 8
All sourcesNASA NTRSDOE OSTI
DOE OSTI2023

Adsorptive recovery of volatile fatty acids from wastewater fermentation broth

This work developed an adsorptive separation and recovery process for anaerobically generated volatile fatty acids (VFAs) from swine wastewater fermentation. Batch adsorption studies were conducted using several weak anion exchange resins and synthetic adsorbent resins to identify a suitable candidate. Relite RAM2 with a tertiary amine functional group showed the highest adsorption (over 98%) of hexanoic acid. Under the different pH values (1.8–7.5) and extraction temperatures (30–50 ↑C), the highest adsorption was observed at a pH of 3.2, which is below the pKa of hexanoic acid, at all the evaluated temperatures. A full factorial design was used to optimize the resin and VFA concentrations, wherein over 99% adsorption could be achieved when the ratio of VFA to resin was below 0.23 (g/g) for model solution (VFAs in water). The adsorption equilibrium could be achieved within 30 min of contact time and 0.5% w/v NaOH was identified as a suitable desorption agent. Under the optimal conditions, 65–72% of VFAs present in fermentation broth was adsorbed, which was increased to 72–76% by using fresh resins. The VFAs adsorption and recovery efficiency were maintained for 9 successive cycles without requiring extensive resin washing and regeneration for both model solution and fermentation broth. Altogether, the work presents a comprehensive study for resin adsorption to recover VFAs and provides a potential industrial-relevant process to recover VFAs from fermentation broth.

09 BIOMASS FUELS↗
DOE OSTI2024

Expanding the synthetic biology toolbox of Cupriavidus necator for establishing fatty acid production

Abstract The Gram-negative betaproteobacterium Cupriavidus necator is a chemolithotroph that can convert carbon dioxide into biomass. Cupriavidus necator has been engineered to produce a variety of high-value chemicals in the past. However, there is still a lack of a well-characterized toolbox for gene expression and genome engineering. Development and optimization of biosynthetic pathways in metabolically engineered microorganisms necessitates control of gene expression via functional genetic elements such as promoters, ribosome binding sites (RBSs), and codon optimization. In this work, a set of inducible and constitutive promoters were validated and characterized in C. necator, and a library of RBSs was designed and tested to show a 50-fold range of expression for green fluorescent protein (gfp). The effect of codon optimization on gene expression in C. necator was studied by expressing gfp and mCherry genes with varied codon-adaptation indices and was validated by expressing codon-optimized variants of a C12-specific fatty acid thioesterase to produce dodecanoic acid. We discuss further hurdles that will need to be overcome for C. necator to be widely used for biosynthetic processes.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Qualitative and quantitative profiles of jejunal oligosaccharides and cecal short–chain fatty acids in broiler chickens receiving different dietary levels of fiber, protein and exogenous enzymes

Dietary supplemental carbohydrases are able to degrade non-starch polysaccharides and generate oligosaccharides in the gastrointestinal tract. This study was conducted to investigate the influence of dietary fiber and protein levels on growth performance, nutrient utilization, digesta oligosaccharides profile and cecal short-chain fatty acid (SCFA) profile in broilers receiving diets supplemented with xylanase or protease individually or in combination. Enzyme supplementation had no effect on growth performance. There was significant (P < 0.05) fiber × protein × xylanase interaction for ileal nitrogen digestibility and significant (P < 0.01) protein × xylanase × protease interaction for nitrogen-corrected apparent metabolizable energy. Birds fed high-fiber diets had higher (P < 0.05) jejunal oligosaccharides and cecal SCFA concentrations. Xylanase and protease combination produced the greatest pentose (Pent) levels in low fiber–adequate protein diets but lowest levels in highfiber–low protein diets. There was significant (P < 0.05) fiber × xylanase × protease interaction explained by the digesta concentrations of (Pent) 3 , (Pent) 4 and (Pent) 5 being greatest (P < 0.5) in protease-only supplemented high-fiber diets but lowest in protease-only supplemented low-fiber diets. Furthermore, these results suggest that, of all the factors investigated, dietary fiber level had the greatest effect on modulating digesta concentration of oligosaccharides and cecal SCFA. Evidence points to the fact that there is considerable capacity for generating pentose oligosaccharides in the digestive tract of broilers receiving diets rich in fibrous feedstuffs, and that this may have a beneficial effect on microbial profile in the digestive tract.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Overcoming barriers to medium-chain fatty alcohol production

Medium-chain fatty alcohols ( mc FaOHs) are aliphatic primary alcohols containing six to twelve carbons that are widely used in materials, pharmaceuticals, and cosmetics. Microbial biosynthesis has been touted as a route to less-abundant chain-length molecules and as a sustainable alternative to current petrochemical processes. Several metabolic engineering strategies for producing mc FaOHs have been demonstrated in the literature, yet processes continue to suffer from poor selectivity and mc FaOH toxicity, leading to reduced titers, rates, and yields of the desired compounds. Here, this opinion examines the current state of microbial mc FaOH biosynthesis, summarizing engineering efforts to tailor selectivity and improve product tolerance by implementing engineering strategies that circumvent or overcome mc FaOH toxicity.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Superior performance biodiesel from biomass-derived fusel alcohols and low grade oils: Fatty acid fusel esters (FAFE)

We demonstrated production of a superior performance biodiesel referred to here as fatty acid fusel alcohol esters (FAFE) – by reacting fusel alcohols (isobutanol, 3-methyl-1-butanol, and (S)-(-)-2-methyl-1-butanol) with oil (glyceryl trioleate) using lipase from Aspergillus oryzae. Reaction conditions corresponding to a molar ratio of 5:1 (fusel alcohols to oil), enzyme loading of 2% w/w, reaction temperature of 35 °C, shaking speed of 250 rpm, and reaction time of 24 h achieved >97% conversion to FAFE. Further, FAFE obtained from reacting a fusel alcohol mixture with corn oil were evaluated for use as a fuel for diesel engines. FAFE mixtures showed superior combustion and cold-flow properties, with the derived cetane numbers up to 4.8 points higher, cloud points up to -6 °C lower, and the heat of combustion up to 2.1% higher than the corresponding FAME samples, depending on the fusel mixture used. This represents a significant improvement for all three metrics, which are typically anti-correlated. Finally, FAFE provides a new opportunity for expanded usage of biodiesel by addressing feedstock limitations, fuel performance, and low temperature tolerance.

09 BIOMASS FUELS↗
DOE OSTI2020

Life-cycle energy use and greenhouse gas emissions of palm fatty acid distillate derived renewable diesel

This study aims to quantify life-cycle fossil energy use and greenhouse gas (GHG) emissions for palm fatty acid distillate (PFAD) derived renewable diesel (RD) taking into consideration different feedstock classifications that are applicable to PFAD (residue, byproduct, or coproduct), and incorporating updated data for key processes. Under the three classifications, the PFAD to RD pathway was modeled using the Greenhouse gases, Regulated Emissions, and Energy Use in Technologies (GREET®) model. PFAD-derived RD could reduce fossil energy consumption by 77%–88%, relative to petroleum diesel. GHG emissions are very sensitive to PFAD classification and coproduct handling methods. Considering the production of palm oil and PFAD and economic value, we maintain that PFAD should be treated as a byproduct in palm oil refineries. With this treatment, PFAD-derived RD could achieve 84% GHG emissions reductions, compared to the emissions of petroleum diesel. We also employed a substitution method to address the substitution of PFAD by other materials in the marketplace. Compared to coproduct allocation results, we found substituting PFAD by tallow, soy oil, barley, and canola oil results in lower GHG emissions. Due to high induced land-use change emissions associated with palm farming, if PFAD is treated as a coproduct with refined palm oil, PFAD-derived RD may not deliver GHG reductions. A sensitivity analysis identified key parameters such as palm fruit yield, oil extraction efficiency in oil mills, and energy use intensity for RD production affects LCA results significantly; future efforts to improve these parameters could result in further GHG reductions.

09 BIOMASS FUELS↗
DOE OSTI2024

Adaptive laboratory evolution and metabolic engineering of Cupriavidus necator for improved catabolism of volatile fatty acids

Bioconversion of high-volume waste streams into value-added products will be an integral component of the growing bioeconomy. Volatile fatty acids (VFAs) (e.g., butyrate, valerate, and hexanoate) are an emerging and promising waste-derived feedstock for microbial carbon upcycling. Cupriavidus necator H16 is a favorable host for conversion of VFAs into various bioproducts due to its diverse carbon metabolism, ease of metabolic engineering, and use at industrial scales. Here, in this study, we report that a common strategy to improve product titers in C. necator, deletion of the polyhydroxybutyrate (PHB) biosynthetic operon, results in a significant growth defect on VFA substrates. Using adaptive laboratory evolution, we identify mutations to the regulator gene phaR, the two-component response regulator-histidine kinase pair encoded by H16_A1372/H16_A1373, and the tripartite transporter assembly encoded by H16_A2296-A2298 as causative for improved growth on VFA substrates. Deletion of phaR and H16_A1373 led to significantly reduced NADH abundance accompanied by large changes to expression of genes involved in carbon metabolism, balance of electron carriers, and oxidative stress tolerance that may be responsible for improved growth of these engineered strains. These results provide insight into the role of PHB biosynthesis in carbon and energy metabolism and highlight a key role for the regulator PhaR in global regulatory networks. By combining mutations, we generated platform strains with significant growth improvements on VFAs, which can enable improved conversion of waste-derived VFA substrates to target bioproducts.

09 BIOMASS FUELS↗
DOE OSTI2025

Co-Catalyst Enabled Biotransformation of Polyunsaturated Fatty Acids for Biobased Monomers

Enzymatic lipid epoxidation offers a promising approach to obtain renewable intermediates for biomaterials, but regiochemical control of these reactions has remained elusive. Here we report the discovery and application of artificial cocatalysts to direct the regioselectivity of catalytic epoxidation with the lipase CpLIP2. The methyl esters of alkyl dicarboxylic acids show the unique ability to direct regioselective epoxidation of polyunsaturated fatty acids, major components of plant and algae-based oils. We apply this transformation to the conversion of linoleic acid into sebacic acid, a dicarboxylic acid precursor valuable for biobased polyester polyurethanes, through a six-step pathway involving a regioselective Meinwald rearrangement. To highlight the route’s significance, sebacic acid was used to prepare a 100% biobased thermoplastic polyurethane, illustrating the relevance of this pathway to industrial applications. Furthermore, this regioselective chemoenzymatic oxidation and rearrangement process can be used to access multiple dicarboxylic acids that have remained previously unexplored as biobased monomers.

09 BIOMASS FUELS↗
DOE OSTI2026

Technoeconomic and Life Cycle Analysis of an Integrated Fermentation and Microbial Electrochemical Process for Volatile Fatty Acid Production from Food Waste

Techno-economic analysis (TEA) and life cycle assessment (LCA) were conducted for an integrated system designed for the production of volatile fatty acid (VFA) from food waste. The TEA estimated a production cost of $\$$3.12/kg VFA, and the LCA predicted negative greenhouse gas (GHG) emissions of -0.4 kg CO 2 e/kg VFA, driven primarily by diverting organic waste from landfills and avoiding methane emissions while producing valuable chemical products. Hotspot analysis showed arrested methanogenesis (AM) fermentation as the largest contributor to costs (37%) and environmental burden (47%), driven by high sodium hydroxide (NaOH) consumption. Distillation and microbial electrosynthesis (MES) units were the next-largest environmental contributors (28% and 18%). Major cost drivers also included residuals management (biosolids and wastewater) and the equipment and operating costs for AM, MES, and sonication pretreatment units. Although the new integrated system is environmentally benign, its costs and environmental impacts can be further reduced by integrating alternative energy sources, minimizing chemical and energy inputs through process optimization, and improving efficiency. In conclusion, this work highlighted the viability of waste-derived VFA production and provided a clear, data-driven strategy to accelerate the commercialization of waste valorization technology.

Carboxylic Acid Production↗
DOE OSTI2025

Insights into genetic determinants of volatile fatty acid catabolism in Cupriavidus necator H16

The soil bacterium Cupriavidus necator H16 is a promising host for upgrading waste-derived volatile fatty acids (VFAs) into renewable biochemicals. While bacterial VFA metabolic pathways are well understood, the C. necator genome encodes multiple enzymes for each catabolic step, and the degree of substrate specificity among these homologs is currently unknown. To gain insight into the catabolism of VFA substrates in C. necator, we performed transcriptomics on cells grown with acetate, propionate, butyrate, valerate, or hexanoate as the sole source of carbon and energy. These data revealed that C. necator upregulates multiple sets of genes putatively involved in substrate activation and β-oxidation in response to VFAs. To better understand this redundancy, we performed biochemical and genetic deletion studies of acyl-CoA synthetase enzymes upregulated during growth on VFA substrates. These results demonstrated the functional redundancy of the C. necator VFA catabolism and led to the identification of a gene cluster, H16_B1332-H16_B1337, that contains several genes that are important for the efficient catabolism of hexanoate. Constitutive expression of a second copy of these hexanoate catabolism genes did not improve growth of C. necator on hexanoate, suggesting that other factors (e.g., redox, transport, or toxicity) may be limiting for growth. Collectively, this work provides new insight into how C. necator uses metabolic regulation to effectively utilize VFA substrates and uncovers the important role of the gene cluster H16_B1332-H16_B1337 in the catabolism of hexanoate.

09 BIOMASS FUELS↗
DOE OSTI2020

Mechanism and dynamics of fatty acid photodecarboxylase

The deposition contains diffraction patterns after hit-finding of time-resolved serial femtosecond crystallography measurements on fatty acid photodecarboxylase performed at LCLS XFEL (proposal ID: LT59). Five datasets are available. One dataset without pump laser excitation, labeled as "dark" and four time-resolved datasets with pump laser excitation at four different time-points labeled as "20 ps", "900 ps", "300 ns" and "2 μs". Please check the publication for more information about the datasets.

CXI↗
DOE OSTI2021

The influence of dietary intake of omega-3 polyunsaturated fatty acids on the association between short-term exposure to ambient nitrogen dioxide and respiratory and cardiovascular outcomes among healthy adults

Background: Short-term exposure to ambient nitrogen dioxide (NO 2 ) is associated with adverse respiratory and cardiovascular outcomes. Supplementation of omega-3 polyunsaturated fatty acids (PUFA) has shown protection against exposure to fine particulate matter. This study aims to investigate whether habitual omega-3 PUFA intake differentially modify the associations between respiratory and cardiovascular responses and short-term exposure to ambient NO 2 . Methods: Sixty-two healthy participants were enrolled into low or high omega-3 groups based on their habitual omega-3 PUFA intake. Each participant was repeatedly assessed for lung function, blood lipids, markers of coagulation and fibrinolysis, vascular function, and heart rate variability (HRV) in up to five sessions, each separated by at least 7 days. This study was carried out in the Research Triangle area of North Carolina, USA between October 2016 and September 2019. Daily ambient NO 2 concentrations were obtained from an area air quality monitoring station on the day of outcome assessment (Lag0), 4 days prior (Lag1-4), as well as 5-day moving average (5dMA). The associations between short-term exposure to NO 2 and the measured indices were evaluated using linear mixed-effects models stratified by omega-3 levels and adjusted by covariates including relative humidity and temperature. Results: The average concentration of ambient NO 2 during the study periods was 5.3±3.8 ppb which was below the National Ambient Air Quality Standards (NAAQS). In the high omega-3 group, an interquartile range (IQR) increase in short-term NO 2 concentrations was significantly associated with increased lung function [e.g. 1.2% (95%CI: 0.2%, 2.2%) in FVC at lag1, 2.6% (95%CI: 0.4%, 4.8%) in FEV1 at 5dMA], decreased blood lipids [e.g. -2.6% (95%CI: -4.4%, -0.9%) in total cholesterol at lag2, -3.1% (95%CI: -6.1%, 0.0%) in HDL at 5dMA, and -3.1% (95%CI: -5.5%, -0.7%) in LDL at lag2], improved vascular function [e.g. 8.9% (95%CI: 0.6%, 17.2%) increase in FMD and 43.1% (95%CI: -79.8%, -6.3%) decrease in endothelin-1 at 5dMA], and changed HRV parameters [e.g. -7.2% (95%CI: -13.6%, -0.8%) in HFn and 13.4% (95%CI: 0.2%, 28.3%) in LF/HF ratio at lag3]. In the low omega-3 group, an IQR increase in ambient NO 2 was associated with elevations in coagulation markers (von Willebrand Factor, D-dimer) and a decrease in HRV (very-low frequency); however, null associations were observed between short-term NO 2 exposure and changes in lung function, blood lipids, and vascular function. Conclusions: The results in this study imply that dietary omega-3 PUFA consumption may offer respiratory and vascular benefits in response to short-term exposure of healthy adults to NO 2 levels below the NAAQS.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2022

Omega-3 fatty acids attenuate cardiovascular effects of short-term exposure to ambient air pollution

Background: Exposure to air pollution is associated with elevated cardiovascular risk. Evidence shows that omega-3 polyunsaturated fatty acids (omega-3 PUFA) may attenuate the adverse cardiovascular effects of exposure to fine particulate matter (PM 2.5 ). However, it is unclear whether habitual dietary intake of omega-3 PUFA protects against the cardiovascular effects of short-term exposure to low-level ambient air pollution in healthy participants. In the present study, sixty-two adults with low or high dietary omega-3 PUFA intake were enrolled. Blood lipids, markers of vascular inflammation, coagulation and fibrinolysis, and heart rate variability (HRV) and repolarization were repeatedly assessed in 5 sessions separated by at least 7 days. This study was carried out in the Research Triangle area of North Carolina, USA between October 2016 and September 2019. Daily PM 2.5 and maximum 8-h ozone (O 3 ) concentrations were obtained from nearby air quality monitoring stations. Linear mixed-effects models were used to assess the associations between air pollutant concentrations and cardiovascular responses stratified by the omega-3 intake levels. Results: The average concentrations of ambient PM 2.5 and O 3 were well below the U.S. National Ambient Air Quality Standards during the study period. Significant associations between exposure to PM 2.5 and changes in total cholesterol, von Willebrand factor (vWF), tissue plasminogen activator, D-dimer, and very-low frequency HRV were observed in the low omega-3 group, but not in the high group. Similarly, O 3 -associated adverse changes in cardiovascular biomarkers (total cholesterol, high-density lipoprotein, serum amyloid A, soluable intracellular adhesion molecule 1, and vWF) were mainly observed in the low omega-3 group. Lag-time-dependent biphasic changes were observed for some biomarkers. Conclusions: This study demonstrates associations between short-term exposure to PM 2.5 and O 3 , at concentrations below regulatory standard, and subclinical cardiovascular responses, and that dietary omega-3 PUFA consumption may provide protection against such cardiovascular effects in healthy adults.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Biophysical characterization and a roadmap towards the NMR solution structure of G0S2, a key enzyme in non-alcoholic fatty liver disease

In the United States non-alcoholic fatty liver disease (NAFLD) is the most common form of chronic liver disease, affecting an estimated 80 to 100 million people. It occurs in every age group, but predominantly in people with risk factors such as obesity and type 2 diabetes. NAFLD is marked by fat accumulation in the liver leading to liver inflammation, which may lead to scarring and irreversible damage progressing to cirrhosis and liver failure. In animal models, genetic ablation of the protein G0S2 leads to alleviation of liver damage and insulin resistance in high fat diets. The research presented in this paper aims to aid in rational based drug design for the treatment of NAFLD by providing a pathway for a solution state NMR structure of G0S2. Here we describe the expression of G0S2 in an E. coli system from two different constructs, both of which are confirmed to be functionally active based on the ability to inhibit the activity of Adipose Triglyceride Lipase. In one of the constructs, preliminary NMR spectroscopy measurements show dominant alpha-helical characteristics as well as resonance assignments on the N-terminus of G0S2, allowing for further NMR work with this protein. Additionally, the characterization of G0S2 oligomers are outlined for both constructs, suggesting that G0S2 may defensively exist in a multimeric state to protect and potentially stabilize the small 104 amino acid protein within the cell. This information presented on the structure of G0S2 will further guide future development in the therapy for NAFLD.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Association of Serum Bile Acid and Unsaturated Fatty Acid Profiles with the Risk of Diabetic Retinopathy in Type 2 Diabetic Patients

Aim: We aimed to identify the ability of serum bile acids (BAs) and unsaturated fatty acids (UFAs) profiles to predict the development of diabetic retinopathy (DR) in type 2 diabetes mellitus (T2DM) patients. Methods: We first used univariate and multivariate analysis to compare 15 serum BA and 11 UFA levels in healthy control (HC) group (n = 82), T2DM patients with DR (n = 58) and T2DM patients without DR (n = 60). Forty T2DM patients were considered for validation. Then, the receiver operating characteristic curve (ROC) and decision curve analysis were used to assess the diagnostic value and clinical benefit of serum biomarkers alone, clinical variables alone or in combination, and the area under the curve (AUC), integrated discrimination improvement (IDI), and net reclassification improvement (NRI) were used to further assess whether the addition of biomarkers significantly improved the predictive ability of the model. Results: Orthogonal partial least squares-discriminant analysis (OPLS-DA) of serum BAs and UFAs separated the three cohorts including HC, T2DM patients with or without DR. The difference in serum BA and UFA profiles of T2DM patients with or without DR was mainly manifested in the three metabolites of taurolithocholic acid (TLCA), tauroursodeoxycholic acid (TUDCA) and arachidonic acid (AA). Together, they had an AUC of 0.785 (0.918 for validation cohort) for predicting DR in T2DM patients. After adjusting for numerous confounding factors, TLCA, TUDCA, and AA were independent predictors that differentiated T2DM with or without DR. The results of AUC, IDI, and NRI demonstrated that adding these three biomarkers to a model with clinical variables statistically increased their predictive value and were replicated in our independent validation cohort. Conclusion: These findings highlight the association of three metabolites, TLCA, TUDCA and AA, with DR and may indicate their potential value in the pathogenesis of DR.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2026

Carbon Capture and Utilization for Protein and Fatty Acids

The unlimited release version of the final report for the "Carbon Capture and Utilization for Protein and Fatty Acids" project. This project advanced an integrated open raceway algae cultivation and processing system to engineering scale for carbon capture and utilization (CCU) from the flue gas of a naphtha-fired power plant.

02 PETROLEUM↗
DOE OSTI2022

Polyunsaturated Fatty Acids Mend Macrophage Transcriptome, Glycome, and Phenotype in the Patients with Neurodegenerative Diseases, Including Alzheimer’s Disease

Macrophages of healthy subjects have a pro-resolution phenotype, upload amyloid-β (Aβ) into endosomes, and degrade Aβ, whereas macrophages of patients with Alzheimer’s disease (AD) generally have a pro-inflammatory phenotype and lack energy for brain clearance of Aβ. To clarify the pathogenesis of sporadic AD and therapeutic effects of polyunsaturated fatty acids (PUFA) with vitamins B and D and antioxidants on monocyte/macrophage (MM) migration in the AD brain, MM transcripts in energy and Aβ degradation, MM glycome, and macrophage clearance of Aβ. We followed for 31.3 months (mean) ten PUFA-supplemented neurodegenerative patients: 3 with subjective cognitive impairment (SCI), 2 with mild cognitive impairment (MCI), 3 MCI/vascular cognitive impairment, 2 with dementia with Lewy bodies, and 7 non-supplemented caregivers. We examined: monocyte migration in the brain and a blood-brain barrier model by immunochemistry and electron microscopy; macrophage transcriptome by RNAseq; macrophage glycome by N-glycan profiling and LTQ-Orbitrap mass spectrometry; and macrophage phenotype and phagocytosis by immunofluorescence. MM invade Aβ plaques, upload but do not degrade Aβ, and release Aβ into vessels, which develop cerebrovascular amyloid angiopathy (CAA); PUFA upregulate energy and Aβ degradation enzyme transcripts in macrophages; PUFA enhance sialylated N-glycans in macrophages; PUFA reduce oxidative stress and increase pro-resolution MM phenotype, mitochondrial membrane potential, and Aβ phagocytosis (p < 0.001). Macrophages of SCI, MCI, and AD patients have interrelated defects in the transcriptome, glycome, Aβ phagocytosis, and Aβ degradation. PUFA mend macrophage transcriptome, enrich glycome, enhance Aβ clearance, and benefit the cognition of early-stage AD patients.

60 APPLIED LIFE SCIENCES↗