Cyrene™ as a tyrosinase inhibitor and anti-browning agent.

The tyrosinase pathway takes part in the enzymatic process of food browning and is primarily responsible for food spoilage - manifesting itself from a decrease in its nutritional value to a deterioration of taste, which consequently leads to a gradual loss of shelf life. Finding safe and bio-based tyrosinase inhibitors and anti-browning agents may be of great importance in agriculture and food industries. Herein, we showed that Cyrene™ exhibits tyrosinase inhibitory activity (IC50: 268.2 µM), the 1.44 times higher than ascorbic acid (IC50: 386.5 µM). Binding mode studies demonstrated that the carbonyl oxygen of Cyrene™ coordinates with both copper ions. Surprisingly, both hydroxyl groups of Cyrene gem-diol perform a monodentate binding mode with both copper ions, at similar distances. This fact suggests that both compounds could have a similar binding mode and, as consequence, similar biological activities in tyrosinase inhibition assays and anti-browning activities.

Effect of N-phenyl substituent on thermal, optical, electrochemical and luminescence properties of 3-aminophthalimide derivatives.

The seven N-phthalimide derivatives substituted with the amine group at the 3-C position in the phenylene ring were synthesized. The effect of N-substituent chemical structure was investigated. The thermal, electrochemical and optical studies were performed and supported by the density functional theory calculations (DFT). The electrochemical investigations of the synthesized low-molecular phthalimides revealed the one oxidation and reduction process with the HOMO energy level under - 5.81 eV and energy-band gap below 3 eV. The N-phthalimide derivatives were emitted light in a blue spectral region in solutions (in polar and non-polar) with the quantum yield between 2 and 68%, dependent on the substituent at the nitrogen atom, solvent and concentration. The N-phthalimide derivatives were emissive also in a solid state as a thin film and powder. They were tested as a component of the active layer with PVK:PBD matrix and as an independent active layer in the organic light-emitting diodes. The registered electroluminescence spectra exhibited the maximum emission band in the 469-505 nm range, confirming the possibility of using N-phthalimides with PVK:PBD matrix as the blue emitters.

Thalidomide derivatives as nanomolar human neutrophil elastase inhibitors: Rational design, synthesis, antiproliferative activity and mechanism of action.

Here, we rationally designed a human neutrophil elastase (HNE) inhibitors 4a-4f derived from thalidomide. The HNE inhibition assay showed that synthesized compounds 4a, 4b, 4e and 4f demonstrated strong HNE inhibiton properties with IC50 values of 21.78-42.30 nM. Compounds 4a, 4c, 4d and 4f showed a competitive mode of action. The most potent compound 4f shows almost the same HNE inhibition as sivelestat. The molecular docking analysis revealed that the strongest interactions occur between the azetidine-2,4-dione group and the following three aminoacids: Ser195, Arg217 and His57. A high correlation between the binding energies and the experimentally determined IC50 values was also demonstrated. The study of antiproliferative activity against human T47D (breast carcinoma), RPMI 8226 (multiple myeloma), and A549 (non-small-cell lung carcinoma) revealed that designed compounds were more active compared to thalidomide, pomalidomide and lenalidomide used as the standard drugs. Additionally, the most active compound 4f derived from lenalidomide induces cell cycle arrest at the G2/M phase and apoptosis in T47D cells.

Coumarin-Based Compounds as Inhibitors of Tyrosinase/Tyrosine Hydroxylase: Synthesis, Kinetic Studies, and In Silico Approaches.

Cancer represents the main cause of morbidity and mortality worldwide, constituting a serious health problem. In this context, melanoma represents the most aggressive and fatal type of skin cancer, with death rates increasing every year. Scientific efforts have been addressed to the development of inhibitors targeting the tyrosinase enzyme as potential anti-melanoma agents due to the importance of this enzyme in melanogenesis biosynthesis. Coumarin-based compounds have shown potential activity as anti-melanoma agents and tyrosinase inhibitors. In this study, coumarin-based derivatives were designed, synthesized, and experimentally evaluated upon tyrosinase. Compound FN-19, a coumarin-thiosemicarbazone analog, exhibited potent anti-tyrosinase activity, with an IC50 value of 42.16 ± 5.16 µM, being more active than ascorbic acid and kojic acid, both reference inhibitors. The kinetic study showed that FN-19 acts as a mixed inhibitor. Still, for this compound, molecular dynamics (MD) simulations were performed to determine the stability of the complex with tyrosinase, generating RMSD, RMSF, and interaction plots. Additionally, docking studies were performed to elucidate the binding pose at the tyrosinase, suggesting that the hydroxyl group of coumarin derivative performs coordinate bonds (bidentate) with the copper(II) ions at distances ranging from 2.09 to 2.61 Å. Then, MM/PBSA calculations revealed that van der Waals interactions are the most relevant intermolecular forces for complex stabilization. Furthermore, it was observed that FN-19 has a binding energy (ΔEMM) value similar to tropolone, a tyrosinase inhibitor. Therefore, the data obtained in this study will be useful for designing and developing novel coumarin-based analogs targeting the tyrosinase enzyme.

Discovery of New 3,3-Diethylazetidine-2,4-dione Based Thiazoles as Nanomolar Human Neutrophil Elastase Inhibitors with Broad-Spectrum Antiproliferative Activity.

A series of 3,3-diethylazetidine-2,4-dione based thiazoles 3a-3j were designed and synthesized as new human neutrophil elastase (HNE) inhibitors in nanomolar range. The representative compounds 3c, 3e, and 3h exhibit high HNE inhibitory activity with IC50 values of 35.02-44.59 nM, with mixed mechanism of action. Additionally, the most active compounds 3c and 3e demonstrate high stability under physiological conditions. The molecular docking study showed good correlation of the binding energies with the IC50 values, suggesting that the inhibition properties are largely dependent on the stage of ligand alignment in the binding cavity. The inhibition properties are correlated with the energy level of substrates of the reaction of ligand with Ser195. Moreover, most compounds showed high and broad-spectrum antiproliferative activity against human leukemia (MV4-11), human lung carcinoma (A549), human breast adenocarcinoma (MDA-MB-231), and urinary bladder carcinoma (UMUC-3), with IC50 values of 4.59-9.86 µM. Additionally, compounds 3c and 3e can induce cell cycle arrest at the G2/M phase and apoptosis via caspase-3 activation, leading to inhibition of A549 cell proliferation. These findings suggest that these new types of drugs could be used to treat cancer and other diseases in which immunoreactive HNE is produced.

New Borane-Protected Derivatives of α-Aminophosphonous Acid as Anti-Osteosarcoma Agents: ADME Analysis and Molecular Modeling, In Vitro Studies on Anti-Cancer Activities, and NEP Inhibition as a Possible Mechanism of Anti-Proliferative Activity.

Many organophosphorus compounds (OPs), especially various α-aminophosphonates, exhibit anti-cancer activities. They act, among others, as inhibitors of the proteases implicated in cancerogenesis. Thesetypes of inhibitors weredescribed, e.g., for neutral endopeptidase (NEP) expressed in different cancer cells, including osteosarcoma (OS). The aim of the present study isto evaluate new borane-protected derivatives of phosphonous acid (compounds 1-7) in terms of their drug-likeness properties, anti-osteosarcoma activities in vitro (against HOS and Saos-2 cells), and use as potential NEP inhibitors. The results revealed that all tested compounds exhibited the physicochemical and ADME properties typical for small-molecule drugs. However, compound 4 did not show capability of blood-brain barrier penetration (Lipinski and Veber rules;SwissAdme tool). Moreover, the α-aminophosphonite-boranes (compounds 4-7) exhibited stronger anti-proliferative activity against OS cells than the other phosphonous acid-borane derivatives (compounds 1-3),especially regarding HOS cells (MTT assay). The most promising compounds 4 and 6 induced apoptosis through the activation of caspase 3 and/or cell cycle arrest at the G2 phase (flow cytometry). Compound 4 inhibited the migration and invasiveness of highly aggressive HOS cells (wound/transwell and BME-coated transwell assays, respectively). Additionally, compound 4 and, to a lesser extent, compound 6 inhibited NEP activity (fluorometric assay). This activity of compound 4 was involved in its anti-proliferative potential (BrdU assay). The present study shows that compound 4 can be considered a potential anti-osteosarcoma agent and a scaffold for the development of new NEP inhibitors.

Novel Azocoumarin Derivatives-Synthesis and Characterization.

The paper presents synthesis and characterization of nine new thiazolyl-(phenyldiazenyl)-2H-chromen-2-one dyes. The impact of substituent structure in thiazole ring in the synthesized azocoumarin derivatives on electrochemical properties, photoisomerization process and photovoltaic response was examined. The dyes were electrochemically active and undergo reduction and oxidation processes. They showed low electrochemically estimated energy band gap in the range of 1.71-2.13 eV. Photoisomerization process of the synthesized molecules was studied in various solvents such as ethanol, chloroform and N,N-dimethylformamide (DMF) upon the UV illumination. It was found that novel azodyes showed reversible trans-cis-trans isomerization and exhibited long thermal back to the trans form, that was even 7 days in DMF. Selected azocoumarin were molecularly dispersed in polystyrene for preparation of guest-host azopolymer systems to study the cis-trans thermal isomerization of obtained dyes in solid state. The photovoltaic activity of the azochromophores was tested in bulk-heterojunction solar cells. They acting as weak donors in device with structure ITO/PEDOT:PSS/dye:PC70BM/Al. No photovoltaic response of cells with azocoumarin derivatives bearing 4-fluorobenzene, 3,4-dichlorobenzene, or 4-(1-adamantyl) unit was found. Additionally, dye which showed the best activity was examined in three-component solar cells ITO/PEDOT:PSS/PTB7:PC70BM:dye/PFN/Al.

Synthesis, electrochemical, optical and biological properties of new carbazole derivatives.

Carbazole skeleton is the key structural motif of many biologically active compounds including synthetic and natural products. Based on the (E)-2-(2-(4-9H-carbazol-9-yl)benzylidene) hydrazinyl)thiazole as skeleton, three novel carbazole dyes were synthesized. The scientific analysis includes the effect of changing the strength of the activating substituents and their exchange for the deactivating substituent on the chemical and biological properties. The presented research showed a significant influence of the CH3, OCH3 and CH2COOC2H5 groups on the spectral properties of the tested derivatives. Their significant influence is also visible in electrochemical, nonlinear-optic and biological properties. The study also included the analysis of the use of the presented derivatives as potential fluorescent probes for in vivo and in vitro tests. Quantum-chemical calculations complement the conducted experiments.

Spectrofluorimetric and Computational Investigation of New Phthalimide Derivatives towards Human Neutrophil Elastase Inhibition and Antiproliferative Activity.

Herein, nine phthalimide-based thiazoles (4a-4i) were synthesized and investigated as new human neutrophil elastase (HNE) inhibitors using spectrofluorimetric and computational methods. The most active compounds containing 4-trifluoromethyl (4c), 4-naphthyl (4e) and 2,4,6-trichloro (4h) substituents in the phenyl ring exhibited high HNE inhibitory activity with IC50 values of 12.98-16.62 µM. Additionally, compound 4c exhibited mixed mechanism of action. Computational investigation provided a consistent picture of the ligand-receptor pattern of inter-actions, common for the whole considered group of compounds. Moreover, compounds 4b, 4c, 4d and 4f showed high antiproliferative activity against human cancer cells lines MV4-11, and A549 with IC50 values of 8.21 to 25.57 µM. Additionally, compound 4g showed high activity against MDA-MB-231 and UMUC-3 with IC50 values of 9.66 and 19.81 µM, respectively. Spectrophotometric analysis showed that the most active compound 4c demonstrated high stability under physiological conditions.

The newly synthesized thiazole derivatives as potential antifungal compounds against Candida albicans.

Recently, the occurrence of candidiasis has increased dramatically, especially in immunocompromised patients. Additionally, their treatment is often ineffective due to the resistance of yeasts to antimycotics. Therefore, there is a need to search for new antifungals. A series of nine newly synthesized thiazole derivatives containing the cyclopropane system, showing promising activity against Candida spp., has been further investigated. We decided to verify their antifungal activity towards clinical Candida albicans isolated from the oral cavity of patients with hematological malignancies and investigate the mode of action on fungal cell, the effect of combination with the selected antimycotics, toxicity to erythrocytes, and lipophilicity. These studies were performed by the broth microdilution method, test with sorbitol and ergosterol, checkerboard technique, erythrocyte lysis assay, and reversed phase thin-layer chromatography, respectively. All derivatives showed very strong activity (similar and even higher than nystatin) against all C. albicans isolates with minimal inhibitory concentration (MIC) = 0.008-7.81 µg/mL Their mechanism of action may be related to action within the fungal cell wall structure and/or within the cell membrane. The interactions between the derivatives and the selected antimycotics (nystatin, chlorhexidine, and thymol) showed additive effect only in the case of combination some of them and thymol. The erythrocyte lysis assay confirmed the low cytotoxicity of these compounds as compared to nystatin. The high lipophilicity of the derivatives was related with their high antifungal activity. The present studies confirm that the studied thiazole derivatives containing the cyclopropane system appear to be a very promising group of compounds in treatment of infections caused by C. albicans. However, this requires further studies in vivo. KEY POINTS: • The newly thiazoles showed high antifungal activity and some of them - additive effect in combination with thymol. • Their mode of action may be related with the influence on the structure of the fungal cell wall and/or the cell membrane. • The low cytotoxicity against erythrocytes and high lipophilicity of these derivatives are their additional good properties.

Synthesis and Physicochemical Characterization of Novel Dicyclopropyl-Thiazole Compounds as Nontoxic and Promising Antifungals.

There is a need to search for new antifungals, especially for the treatment of the invasive Candida infections, caused mainly by C. albicans. These infections are steadily increasing at an alarming rate, mostly among immunocompromised patients. The newly synthesized compounds (3a-3k) were characterized by physicochemical parameters and investigated for antimicrobial activity using the microdilution broth method to estimate minimal inhibitory concentration (MIC). Additionally, their antibiofilm activity and mode of action together with the effect on the membrane permeability in C. albicans were investigated. Biofilm biomass and its metabolic activity were quantitatively measured using crystal violet (CV) staining and tetrazolium salt (XTT) reduction assay. The cytotoxic effect on normal human lung fibroblasts and haemolytic effect were also evaluated. The results showed differential activity of the compounds against yeasts (MIC = 0.24-500 µg/mL) and bacteria (MIC = 125-1000 µg/mL). Most compounds possessed strong antifungal activity (MIC = 0.24-7.81 µg/mL). The compounds 3b, 3c and 3e, showed no inhibitory (at 1/2 × MIC) and eradication (at 8 × MIC) effect on C. albicans biofilm. Only slight decrease in the biofilm metabolic activity was observed for compound 3b. Moreover, the studied compounds increased the permeability of the membrane/cell wall of C. albicans and their mode of action may be related to action within the fungal cell wall structure and/or within the cell membrane. It is worth noting that the compounds had no cytotoxicity effect on pulmonary fibroblasts and erythrocytes at concentrations showing anticandidal activity. The present studies in vitro confirm that these derivatives appear to be a very promising group of antifungals for further preclinical studies.

Effect of the Chloro-Substitution on Electrochemical and Optical Properties of New Carbazole Dyes.

Carbazole derivatives are the structural key of many biologically active substances, including naturally occurring and synthetic ones. Three novel (E)-2-(2-(4-9H-carbazol-9-yl)benzylidene)hydrazinyl)triazole dyes were synthesized with different numbers of chlorine substituents attached at different locations. The presented research has shown the influence of the number and position of attachment of chlorine substituents on electrochemical, optical, nonlinear, and biological properties. The study also included the analysis of the use of the presented derivatives as potential fluorescent probes for in vivo and in vitro tests. Quantum-chemical calculations complement the conducted experiments.

Effect of the dichloro-substitution on antiproliferative activity of phthalimide-thiazole derivatives. Rational design, synthesis, elastase, caspase 3/7, and EGFR tyrosine kinase activity and molecular modeling study.

Phthalimide derivatives are a promising group of anticancer drugs, while aminothiazoles have great potential as elastase inhibitors. In these context fourteen phthalimido-thiazoles containing a dichloro-substituted phenyl ring with high antiproliferative activity against various cancer cell lines were designed and synthesized. Among the screened derivatives, compounds 5a-5e and 6a-6f showed high activity against human leukemia (MV4-11) cells with IC50 values in the range of 5.56-16.10 µM. The phthalimide-thiazoles 5a, 5b and 5d showed the highest selectivity index (SI) relative to MV4-11 with 11.92, 10.80 and 8.21 values, respectively. The antiproliferative activity of compounds 5e, 5f and 6e, 6f against human lung carcinoma (A549) cells is also very high, with IC50 values in the range of 6.69-10.41 µM. Lead compounds 6e and 6f showed elastase inhibition effect, with IC50 values about 32 µM with mixed mechanism of action. The molecular modeling studies showed that the binding energies calculated for all set of compounds are strongly correlated with the experimentally determined values of IC50. The lead compound 6e also increases almost 16 times caspase 3/7 activity in A549 cells compared to control. We have also demonstrated that compound 6f reduced EGFR tyrosine kinase levels in A549 cells by approximately 31%. These results clearly suggest that 3,4-dichloro-derivative 6e and 3,5-dichloro-derivative 6f could constitute lead dual-targeted anticancer drug candidates.

Systematic Analysis of the Role of Substituents in Oxiranes, Oxetanes, and Oxathietanes Chemical Shifts.

The important role substituents play on proton chemical shifts in heterocyclic compounds was investigated in detail. For this purpose, a considerable number of model oxiranes, oxetanes, and oxathietanes with different substituents were studied in a systematic way. In addition, the oxygen and sulfur heteroatom influence on the chemical shift values was analyzed. The density functional theory (DFT) approximation was employed together with the M06 and the B3LYP functionals and the aug-pcS-1 and the 6-311++G** basis sets. We carried out a careful analysis of the shift values and the changes in the corresponding molecular electrostatic potential surfaces due to substitution. We observed that chemical shift values for the protons closest to the substituents are larger for the chloro and fluoro derivatives than those for the cyano and ethynyl ones. The presence of oxygen as well as sulfur in the ring causes an increase of the chemical shift values, most pronounced for the atom closest to the substituent. A large decrease of the proton shifts was observed when going from methylenecyclopropane to methyleneoxirane that can be attributed to π-electron resonance. Protons diagonal to the substituents behaved in a different way depending on their cis or trans disposition with respect to them. The conclusions of the present study will be useful in theoretical and experimental work on NMR spectra of heterocyclic compounds.

Tropinone-Derived Alkaloids as Potent Anticancer Agents: Synthesis, Tyrosinase Inhibition, Mechanism of Action, DFT Calculation, and Molecular Docking Studies.

A new series of hybrid compounds with tropinone and thiazole rings in the structure was designed and synthesized as potential anticancer agents. They were tested against human multiple myeloma (RPMI 8226), lung carcinoma (A549), breast adenocarcinoma (MDA-MB-231), and mouse skin melanoma (B16-F10) cell lines. Toxicity was tested on human normal skin fibroblasts (HSF) and normal colon fibroblasts (CCD-18Co). The growth inhibition mechanism of the most active derivative was analyzed through investigation of its effect on the distribution of cell cycle phases and ability to induce apoptosis and necrosis in RPMI 8226 and A549 cancer cells. The tyrosinase inhibitory potential was assessed, followed by molecular docking studies. Compounds 3a-3h show high anticancer activity against MDA-MB-231 and B16-F10 cell lines with IC50 values of 1.51-3.03 µM. Moreover, the cytotoxic activity of the investigated compounds against HSF and CCD-18Co cells was 8-70 times lower than against the cancer cells or no toxicity was shown in our tests, with derivative 3a being particularly successful. The mechanism of action of compound 3a in RPMI 8226 cell was shown to be through induction of cell death through apoptosis. The derivatives show ability to inhibit the tyrosinase activity with a mixed mechanism of inhibition. The final molecular docking results showed for IC50 distinct correlation with experiment.

Towards understanding the interaction of (S)-thalidomide with nucleobases.

Interaction of (S)-thalidomide molecule with four nucleobases: adenine, guanine, cytosine and thymine, is investigated in details employing density functional theory methods. Different mutual positions of the molecules are considered, with the starting geometries enabling hydrogen bond interactions between the monomers. Optimization of geometrical parameters is carried out within the B3LYP/6-311G** approximation and followed by evaluation of vibrational frequencies. Binding and interaction energies are calculated employing exchange-correlation functionals including long-range corrections and properly diffuse basis sets. The strongest interaction exists within the (S)-thalidomide-guanine complex. Interestingly, in one of the investigated (S)-thalidomide-guanine complexes two bifurcated hydrogen bonds are observed. The two hydrogens involved in one of them are bonded to a carbon atom in the α position relative to carbonyl group. The present study can be useful in the design of new anticancer and antiviral drugs interacting selectively with DNA or RNA.

Benzo[b]thiophene-thiazoles as potent anti-Toxoplasma gondii agents: Design, synthesis, tyrosinase/tyrosine hydroxylase inhibitors, molecular docking study, and antioxidant activity.

Synthesis and investigation of anti-Toxoplasma gondii activity of novel thiazoles containing benzo [b]thiophene moiety are presented. Among the derivatives, compound 3k with adamantyl group shows exceptionally high potency against Me49 strain with IC50 (8.74 µM) value which is significantly lower than the activity of trimethoprim (IC50 39.23 µM). In addition, compounds 3a, 3b and 3k showed significant activity against RH strain (IC50 51.88-83.49 µM). The results of the cytotoxicity evaluation showed that Toxoplasma gondii growth was inhibited at non-cytotoxic concentrations for the mammalian L929 fibroblast (CC30 ∼ 880 µM). The most active compound 3k showed tyrosinase inhibition effect, with IC50 value of 328.5 µM. The binding energies calculated for compounds 3a-3e, 3k are strongly correlated with the experimentally determined values of tyrosinase inhibition activity. Moreover, the binding energies corresponding to the same ligands and calculated for both tyrosinase and tyrosine hydroxylase are also correlated with each other, suggesting that tyrosinase inhibitors may also have an inhibitory effect on tyrosine hydroxylase. Compounds 3j and 3k have also very strong antioxidant activity (IC50 15.9 and 15.5 µM), respectively, which is ten times higher than well-known antioxidant BHT.

Discovery of tropinone-thiazole derivatives as potent caspase 3/7 activators, and noncompetitive tyrosinase inhibitors with high antiproliferative activity: Rational design, one-pot tricomponent synthesis, and lipophilicity determination.

We have designed novel tropinone-thiazole derivatives that showed high antiproliferative activity against a variety of cancer cell lines via caspase 3/7 activation mechanism. Among the derivatives, compounds 3b-3h were found to exhibit high activity against human leukemia (MV4-11), human lung carcinoma (A549), human breast carcinoma (MCF-7), and skin melanoma (B16-F10) cancer cell lines, with IC50 values of 5.43-11.06 µM. The lead compound 3g increases caspase 3/7 activity in A549 cells 25 times more than the control, and 2 times more than reference drug camptothecin. We have also found that tropinone-thiazole derivatives exhibit high tyrosinase inhibitory activity. The lead compounds 3g and 3h showed tyrosinase inhibition effect, with IC50 values 3.22 and 3.51 µM, respectively. These inhibitory activities are 22 times higher than the activity of kojic acid (IC50 72.27 µM) and 120 times higher than activity of ascorbic acid (IC50 386.5 µM). For compounds 3g and 3h, the experimentally determined lipophilicity correlates very well with their enzymatic activities. These data suggest that presented compounds could constitute lead anticancer drug candidates.

Triazene salts: Design, synthesis, ctDNA interaction, lipophilicity determination, DFT calculation, and antiproliferative activity against human cancer cell lines.

Synthesis, characterization and investigation of antiproliferative activity of nine triazene salts against human cancer cells lines (MV-4-11, MCF-7, JURKAT, HT-29, Hep-G2, HeLa, Du-145 and DAUDI), and normal human mammary epithelial cell line (MCF7-10A) is presented. The structures of novel compounds were determined using 1H and 13C NMR, and GC-APCI-MS analyses. Among the derivatives, compound 2c, 2d, 2e and 2f has very strong activity against biphenotypic B myelomonocytic leukemia MV4-11, with IC50 values from 5.42 to 7.69 µg/ml. The cytotoxic activity of compounds 2c-2f against normal human mammary gland epithelial cells MCF-10A is 6-11 times lower than against cancer cell lines. Our results also show that compounds 2c and 2f have very strong activity against DAUDI and HT-29 with IC50 4.91 µg/ml and 5.59 µg/ml, respectively. Their lipophilicity was determined using reversed-phase ultra-performance liquid chromatography and correlated with antiproliferative activity. Our UV-Vis spectroscopic results indicate also that triazene salts tends to interact with negatively charged DNA phosphate chain. To support the experiment, theoretical calculations of the 1H NMR shifts were carried out within the Density Functional Theory.