Hollow-like three-dimensional structure of methyl orange-delaminated Ti3C2 MXene nanocomposite for high-performance electrochemical sensing of tryptophan.

Tryptophan(Trp) is being explored as a potential biomarker for various diseases associated with decreased tryptophan levels; however, metabolomic methods are expensive and time-consuming and require extensive sample analysis, making them urgently needed for trace detection. To exploit the properties of Ti3C2 MXenes a rational porous methyl orange (MO)-delaminated Ti3C2 MXene was prepared via a facile mixing process for the electrocatalytic oxidation of Trp. The hollow-like 3D structure with a more open structure and the synergistic effect of MO and conductive Ti3C2 MXene enhanced its electrochemical catalytic capability toward Trp biosensing. More importantly, MO can stabilize Ti3C2 MXene nanosheets through noncovalent π-π interactions and hydrogen bonding. Compared with covalent attachment, these non-covalent interactions preserve the electronic conductivity of the Ti3C2 MXene nanosheets. Finally, the addition of MO-derived nitrogen (N) and sulfur (S) atoms to Ti3C2 MXene enhanced the electronegativity and improved its affinity for specific molecules, resulting in high-performance electrocatalytic activity. The proposed biosensor exhibited a wide linear response in concentration ranges of 0.01-0.3 µM and 0.5-120 µM, with a low detection limit of 15 nM for tryptophan detection, and high anti-interference ability in complex media of human urine and egg white matrices. The exceptional abilities of the MO/Ti3C2 nanocatalyst make it a promising electrode material for the detection of important biomolecules.

An economical preparation strategy of magnetic biochar with high specific surface area for efficient removal of methyl orange.

Magnetic biochar (MBC) was obtained from pepper straw by impregnation-microwave pyrolysis method. The pyrolysis temperature and FeCl3 impregnation concentration were investigated on the structural properties of MBC and the adsorption of methyl orange (MO) in water. Characterization results showed that pyrolysis temperature and iron species significantly increased the specific surface area of MBC, which could reach the maximum of 2038.61 m2/g, and also provided more active adsorption sites by promoting the generation of graphitized structures and surface polar functional groups. MBC0.2-900 was selected as the adsorbent for MO with the maximum adsorption capacity reached 437.18 mg·g-1, 3.4 times higher than the virgin biochar. The adsorption process was dominated by chemisorption as well as spontaneous and exothermic. The adsorption mechanisms included pore-filling interaction, π-π EDA interaction, electrostatic interaction, hydrogen bonding, and Lewis acid-base electron interaction. In addition, MBC also exhibited excellent separability and reusability as a low-cost adsorbent. This study provided some theoretical foundation and technological support for producing high-performance biochar and developing pollutant removal technology in wastewater.

Deciphering the catalytic activity of nickel anchored on Fe3O4@SiO2@3-CPMS@L as a magnetically recoverable nanocatalyst for the efficacious reduction of 4-nitrophenol, nitrobenzene, and methyl orange.

In this paper, a versatile heterogeneous nanocatalyst was fabricated employing a self-assembly technique. To commence, Fe3O4 MNPs were coated with a thin layer of SiO2 using the stobbers method. Subsequently, the surface was further functionalized with 3-CPMS, followed by a reaction with a Schiff base. Finally, nickel NPs were deposited on the surface through in situ deposition, forming the Fe3O4@SiO2@3-CPMS@L-Ni magnetic nanocatalyst. The architecture of this magnetic nanocatalyst was meticulously characterized through an array of sophisticated techniques: XRD, FT-IR, SEM, TEM, BET and VSM. The XRD diffraction pattern confirmed the presence of Fe3O4 MNPs, SiO2, and Ni peaks, providing evidence for successful synthesis. Moreover, the successful functionalization with a Schiff base was demonstrated by the presence of an azomethane peak in the FTIR spectra of the synthesized nanocatalyst. The fabricated nanocatalyst was adeptly utilized for the reduction of 4-NP, NB, and MO demonstrating a remarkably elevated rate of catalytic efficacy. Moreover, this catalyst was effortlessly retrievable through the application of an external magnet, and it maintained its catalytic prowess across at least six consecutive cycles. The utilization of water as an environmentally friendly solvent, coupled with the utilization of abundant and cost-effective nickel catalyst instead of the costly Pd or Pt catalysts, along with the successful recovery and scalability of the catalyst, render this method highly advantageous from both environmental and economic perspectives for the reduction of 4-NP, NB, and MO.

A novel urease-assisted ratiometric fluorescence sensing platform based on pH-modulated copper-quenched near-infrared carbon dots and methyl red-quenched red carbon dots for selective urea monitoring.

A novel and sensitive fluorescence ratiometric method is developed for urea detection based  on the pH-sensitive response of two fluorescent carbon dot (CD) systems: R-CDs/methyl red (MR) and NIR-CDs/Cu2+. The sensing mechanism involves breaking down urea using the enzyme urease, releasing ammonia and increasing pH. At higher pH, the fluorescence of NIR-CDs is quenched due to the enhanced interaction with Cu2+, while the fluorescence of R-CDs is restored as the acidic MR converts to its basic form, removing the inner filter effect. The ratiometric signal (F608/F750) of the R-CDs/MR and NIR-CDs/Cu2+ intensities changed in response to the pH induced by urea hydrolysis, enabling selective and sensitive urea detection. Detailed spectroscopic and morphological investigations confirmed the fluorescence probe design and elucidated the sensing mechanism. The method exhibited excellent sensitivity (0.00028 mM LOD) and linearity range (0.001 - 8.0 mM) for urea detection, with successful application in milk samples for monitoring adulteration, demonstrating negligible interference and high recovery levels (96.5% to 101.0%). This ratiometric fluorescence approach offers a robust strategy for selective urea sensing in complicated matrices.

Decolorization and detoxification of direct blue 5B by a Marinobacter-dominated halo-thermoalkalophilic consortium.

Azo dye-containing sewage is commonly detected at high salinity, temperature and pH. In this study, a halo-thermoalkalophilic azo dye decolorization consortium was enriched and named "consortium HL". Consortium HL which was dominated by Marinobacter (84.30%), Desulfocurvibacter (1.89%), and Pseudomonas (1.85%), was able to completely decolorize Direct Blue 5B (DB5) during incubation with the material at 5% salinity, 50 °C, and pH 9 for 30 h. The decolorization mechanism was proposed based on combined metagenomic analysis, GC‒MS, and enzymatic activity detection. The action of the consortium HL showed great tolerance to variations in salinity, temperature and pH. A phytotoxicity study indicated that the metabolic intermediates showed no significant toxicity to the generation of Cucumis sativus and Oryza sativa seeds. This study, in which azo dye decolorization and degradation under high-salt, high-temperature and high-alkalinity conditions were investigated and deeply analyzed by metagenomic information, is the first report regarding the ability of Marinobacter to decolorize azo dyes at high temperatures.

Discovery, isolation, and characterization of diazeniumdiolate siderophores.

The C-diazeniumdiolate (N-nitrosohydroxylamine) group in the amino acid graminine (Gra) is a newly discovered Fe(III) ligand in microbial siderophores. Graminine was first identified in the siderophore gramibactin, and since this discovery, other Gra-containing siderophores have been identified, including megapolibactins, plantaribactin, gladiobactin, trinickiabactin (gramibactin B), and tistrellabactins. The C-diazeniumdiolate is photoreactive in UV light which provides a convenient characterization tool for this type of siderophore. This report details the process of genomics-driven identification of bacteria producing Gra-containing siderophores based on selected biosynthetic enzymes, as well as bacterial culturing, isolation and characterization of the C-diazeniumdiolate siderophores containing Gra.

Photophysics in Biomembranes: Computational Insight into the Interaction between Lipid Bilayers and Chromophores.

ConspectusLight is ubiquitously available to probe the structure and dynamics of biomolecules and biological tissues. Generally, this cannot be done directly with visible light, because of the absence of absorption by those biomolecules. This problem can be overcome by incorporating organic molecules (chromophores) that show an optical response in the vicinity of those biomolecules. Since those optical properties are strongly dependent on the chromophore's environment, time-resolved spectroscopic studies can provide a wealth of information on biosystems at the molecular scale in a nondestructive way. In this work, we give an overview on the multiscale computational strategy developed by us in the last eight years and prove that theoretical studies and simulations are needed to explain, guide, and predict observations in fluorescence experiments. As we challenge the accepted views on existing probes, we discover unexplored abilities that can discriminate surrounding lipid bilayers and their temperature-dependent as well as solvent-dependent properties. We focus on three archetypal chromophores: diphenylhexatriene (DPH), Laurdan, and azobenzene. Our method shows that conformational changes should not be neglected for the prototype rod-shaped molecule DPH. They determine its position and orientation in a liquid-ordered (Lo) sphingomyelin/cholesterol (SM/Chol) bilayer and are responsible for a strong differentiation of its absorption spectra and fluorescence decay times in dioleoylphosphatidylcholine (DOPC) and dipalmitoylphosphatidylcholine (DPPC) membranes, which are at room temperature in liquid-disordered (Ld) and solid-gel (So) phases, respectively. Thanks to its pronounced first excited state dipole moment, Laurdan has long been known as a solvatochromic probe. Since this molecule has however two conformers, we prove that they exhibit different properties in different lipid membrane phases. We see that the two conformers are only blocked in one phase but not in another. Supported by fluorescence anisotropy decay simulations, Laurdan can therefore be regarded as a molecular rotor. Finally, the conformational versatility of azobenzene in saturated Ld lipid bilayers is simulated, along with its photoisomerization pathways. By means of nonadiabatic QM/MM surface hopping analyses (QM/MM-SH), a dual mechanism is found with a torsional mechanism and a slow conversion for trans-to-cis. For cis-to-trans, simulations show a much higher quantum yield and a so-called "pedal-like" mechanism. The differences are related to the different potential energy surfaces as well as the interactions with the surrounding alkyl chains. When tails of increased length are attached to this probe, cis is pushed toward the polar surface, while trans is pulled toward the center of the membrane.

Synthesis of Azo Analogs for Investigating Phosphatidic Acid-Mediated Signaling.

Phosphatidic acid (PA) is a key signaling lipid that plays a crucial role in regulating various cellular processes. Studies have shown that azobenzene-containing PA analogues can be used as an all-chemical strategy for light-mediated control of PA signaling. These photoswitchable lipids offer a solution to the limitations of traditional bulk dosing methods by allowing for light- and shape-dependent interactions with protein effectors and lipid-metabolizing enzymes. This chapter describes how to synthesize AzoPA and dAzoPA.

An optimised protocol for the detection of lipofuscin, a versatile and quantifiable marker of cellular senescence.

Lipofuscin is a yellow-brown pigment typically found in the lysosomes that contains a mixture of molecules including lipids, metals and misfolded proteins. The use of Sudan black B to detect lipofuscin accumulation, a well described marker of cellular senescence and ageing, was first described in 2013 by Georgakopoulou, et al. Here, we provide an optimisation of the original protocol. Firstly, we adjusted the staining methodology for increased ease of use on cultured cells. Secondly, we show that Sudan black B-stained lipofuscin emits strong fluorescence in the far-red channel making it suitable for fluorescence microscopy detection and quantification. Moreover, we also demonstrate that this optimised protocol can be utilised in conjunction with standard immunofluorescence staining techniques, making possible the simultaneous detection of lipofuscin and other cellular proteins of interest, like additional markers of senescence. This is a significant advantage over the most commonly used method for senescence detection, based on beta galactosidase enzymatic activity. We therefore believe that these findings and the provided optimised protocol will represent a useful tool for the scientific community in the field of cellular senescence.

Facile synthesis of silver doped manganese oxide nanocomposite with superior photocatalytic and antimicrobial activity under visible spectrum.

Water pollution and antimicrobial resistance (AMR) have become two global threats; 80% of diseases and 50% of child deaths are due to poor water quality. In this study, hydrothermal processing was employed to manufacture manganese oxide nanorods. Silver dopant was deposited on the surface of manganese oxide. XRD diffractogram confirmed the facile synthesis of Ag/Mn2O3 nanocomposite. XPS survey analysis demonstrated silver content of 9.43 atom %. Photocatalytic measurements demonstrated the outstanding efficiency of the Ag-Mn2O3 compared to virgin oxide particles under visible radiation. Degradation efficiencies Mn2O3 and Ag/Mn2O3 on methyl orange (MO) dye was found to be 53% and 85% under visible spectrum. Silver dopant was found to decrease the binding energy of valence electrons; this action could support electron-hole pair generation under visible spectrum and could promote catalytic performance. Ag/Mn2O3 NPs demonstrated most effective performance (95% removal efficiency) at pH 3; this could be ascribed to the electrostatic attraction between positively charged catalyst and the negatively charged MO. Ag/Mn2O3 demonstrated enhanced antibacterial activity against Gram-positive Staphylococcus aureus (S. aureus) (19 mm ZOI), and Gram-negative Escherichia coli (E. coli) (22 mm ZOI) respectively; the developed nanocomposite demonstrated advanced anti-film activity with inhibition percentage of 95.5% against E. coli followed by 89.5% against S. aureus.

Eco-Friendly g-C3N4/Carboxymethyl Cellulose/Alginate Composite Hydrogels for Simultaneous Photocatalytic Degradation of Organic Dye Pollutants.

The presented study was focused on the simple, eco-friendly synthesis of composite hydrogels of crosslinked carboxymethyl cellulose (CMC)/alginate (SA) with encapsulated g-C3N4 nanoparticles. The structural, textural, morphological, optical, and mechanical properties were determined using different methods. The encapsulation of g-C3N4 into CMC/SA copolymer resulted in the formation of composite hydrogels with a coherent structure, enhanced porosity, excellent photostability, and good adhesion. The ability of composite hydrogels to eliminate structurally different dyes with the same or opposite charge properties (cationic Methylene Blue and anionic Orange G and Remazol Brilliant Blue R) in both single- and binary-dye systems was examined through adsorption and photocatalytic reactions. The interactions between the dyes and g-C3N4 and the negatively charged CMC/SA copolymers had a notable influence on both the adsorption capacity and photodegradation efficiency of the prepared composites. Scavenger studies and leaching tests were conducted to gain insights into the primary reactive species and to assess the stability and long-term performance of the g-C3N4/CMC/SA beads. The commendable photocatalytic activity and excellent recyclability, coupled with the elimination of costly catalyst separation requirements, render the g-C3N4/CMC/SA composite hydrogels cost-effective and environmentally friendly materials, and strongly support their selection for tackling environmental pollution issues.

Investigating the Influence of Photoswitchable Lipids on the Structure and Dynamics of Lipid Membranes: Fundamentals and Potential Applications.

In this work, we delve into the impact of photoisomerization of photoswitchable lipids (PSLs) on the membrane structure and dynamics at a molecular level. Through all-atom molecular dynamics simulations, we explore how UV irradiation-induced trans-to-cis isomerization of these lipids, particularly the azobenzene-derivatized phosphatidylcholine (AzoPC) lipid, influences the structure and dynamics of a simplified lipid membrane, mimicking those of E. coli bacteria across different temperatures. Our findings align with previous experimental observations regarding membrane properties and offer insights into localized effects and microscopic heterogeneity. Additionally, we estimate the relaxation time scale of the lipid membrane following AzoPC photoisomerization. Moreover, we demonstrate the feasibility of photoactivated drug release, exemplified by the controlled liberation of doxorubicin, an anticancer agent, through the membrane, suggesting the potential of PSLs in engineering photoactivated liposomes, coined as photoazosomes, for precise targeted drug delivery applications.

Eco-friendly synthesis of N- cholyl mercapto histidine capped silver nanoparticles and its sensing of mercury (II) ions and photo catalytic degradation of methyl orange.

In this report, we have developed highly water soluble and stable silver nanoparticles (Ag NPs) utilizing N-Cholyl Mercapto Histidine (NCMH) as a reducing and stabilizing agent with near the primary critical micellar concentration (CMC) under ambient sunlight irradiation. Moreover, The NCMH was firstly synthesized by demonstrating the reaction between cholic acid and 2- Mercapto Histidine through a simple acid amine coupling approach. The primary and secondary CMC of NCMH surfactant was measured by pyrene (1 × 10-6 M) as a fluorescent probe, and values were found to be 3.2 and 13.1 mM respectively. The synthesized Ag NPs showed at neutral pH and highly stable for more than one year without any noticeable aggregation. The TEM analysis displays the synthesized Ag NPs having a spherical shape and average size of 9.6 ± 0.5 nm. The synthesis of stabilized Ag NPs was used for ultra-sensitive and selective detection of Hg2+ ions in aqueous medium were monitored by Uv-visible spectrometer and naked eyes with a lowest limit of detection (LOD) 7 nM. The photo-catalytic degradation of methyl orange (MO) by utilizing Ag NPs as nano-catalyst exhibits a potential degradation within a study period of 180 min. Concluding that, facile and cost effective green synthesis of NCMH capped Ag NPs possess excellent reducing ability towards the selective detection of Hg2+ ions along with photo-catalytic degradation of MO dye. These true findings detached an innovative pathway of Ag NPs towards the reactivity against the catalytic activity of dye degradation and selective sensing of Hg2+ ions. Thus it paves the way for extensive range of novel potential applications of Ag NPs in various environment friendly approaches of sensitive and analytical protocol in the future.

Rapid eco-friendly selective dye removal using modified chitosan-based sponges: Synthesis, characterization, and application.

The ongoing challenge of water scarcity persists alongside a concerning rise in water pollution driven by population expansion and industrial development. As a result, urgent measures are imperative to address the pressing need for a clean and sustainable water supply. In this study, a sustainable and green approach was utilized to prepare four chitosan-based sponges from a chemically modified chitosan with different alkyl chains in aqueous medium and at room temperature. The resulting sponges displayed excellent stability in water with outstanding dye removal efficiency. The adsorption capacity was associated with the alkyl chain length incorporated to the polymer backbone. All sponges displayed a high adsorption capacity of methyl orange (MO) ranges between 238 and 380 mg g-1, while a low capacity were obtained for methylene blue (MB) and Rhodamine B (RB). Competitive adsorption experiments were conducted on binary and ternary mixtures to assess the selective removal of MO from a mixture of dyes in which the separation factor was found to be ranging between 1.6 and 32. The adsorption kinetics isotherms of all sponges followed the pseudo-second-order, and the Langmuir model was found to be more suitable than the Freundlich for the adsorption of MO on the sponges. The chitosan-based sponges showed stable performance, robustness and reusability over 5 adsorption-desorption cycles, indicating their great potential for water treatment applications.

Photoresponsive heparin ionic complexes toward controllable therapeutic efficacy of anticoagulation.

Controllable heparin-release is of great importance and necessity for the precise anticoagulant regulation. Efforts have been made on designing heparin-releasing systems, while, it remains a great challenge for gaining the external-stimuli responsive heparin-release in either intravenous or catheter delivery. In this study, an azobenzene-containing ammonium surfactant is designed and synthesized for the fabrication of photoresponsive heparin ionic complexes through the electrostatic complexation with heparin. Under the assistance of photoinduced trans-cis isomerization of azobenzene, the obtained heparin materials perform reversible athermal phase transition between ordered crystalline and isotropic liquid state at room temperature. Compared to the ordered state, the formation of isotropic state can effectively improve the dissolving of heparin from ionic materials in aqueous condition, which realizes the photo-modulation on the concentration of free heparin molecules. With good biocompatibility, such a heparin-releasing system addresses photoresponsive anticoagulation in both in vitro and in vivo biological studies, confirming its great potential clinical values. This work provides a new designing strategy for gaining anticoagulant regulation by light, also opening new opportunities for the development of photoresponsive drugs and biomedical materials based on biomolecules.

Cellulose-citric acid-chitosan@metal sulfide nanocomposites: Methyl orange dye removal and antibacterial activity.

In this study, to develop efficient adsorbents in removing water pollution, new cellulose-citric acid-chitosan@metal sulfide nanocomposites (CL-CA-CS@NiS and CL-CA-CS@CuS) were synthesized by one-pot reaction at mild conditions and characterized using X-ray powder diffraction (XRD), thermogravimetric analysis (TGA), scanning electron microscope (SEM), Energy Dispersive X-ray (EDX) and Brunauer-Emmett-Teller (BET) isotherm. The results of characterization techniques confirm that the desired compounds have been successfully synthesized. The as-prepared composites were applied for the removal of methyl orange (MO) dye from aqueous solutions using a batch technique, and the effect of key factors such as initial pH, shaking time, MO concentration, temperature and adsorbent dose were investigated and discussed. Adsorption results exhibited positive impact of temperature, shaking time and adsorbent dose on the MO removal percent. The MO removal percent has been increased over a wide range of pH from 2 (27.6 %) to 6 (98.8 %). Also, almost being constant over a wide range of MO concentration (10-70 mg/L). The results demonstrated that the maximum removal percentage of MO dye (98.9 % and 93.4 % using CL-CA-CS@NiS and CL-CA-CS@CuS, respectively) was achieved under the conditions of pH 6, shaking time of 120 min, adsorbent dose of 0.02 g, MO concentration of 70 mg/L and temperature of 35 °C. The pseudo-second-order (PSO) and Langmuir models demonstrated the best fit to the kinetic and equilibrium data. Also, the thermodynamic results showed that the MO removal process is endothermic and spontaneous in nature. The MO adsorption can be happened by different electrostatic attraction, n-π and π-π stacking and also hydrogen bonding interaction. In addition, antibacterial activity of CL-CA-CS@NiS and CL-CA-CS@CuS nanocomposites exhibited a superior efficiency against S. aureus.

Preparation and potential of chitosan-based/Al2O3 green hydrogel composites for the removal of methyl red dye from simulated solution.

Different dyes are discharged into water streams, causing significant pollution to the entire ecosystem. The present work deals with the removal of acid red 2 dye (methyl red-as an anionic dye) by green sorbents based on chitosan derivatization. In this regard, two classes of chitosan derivatives-a total of six-were prepared by gamma irradiation at 30 kGy. The first group (group A) constitutes a crosslinked chitosan/polyacrylamide/aluminum oxide with different feed ratios, while the second group, identified as group B, is composed of crosslinked carboxymethyl chitosan/polyacrylamide/aluminum oxide with different ratios. Glycerol was added to soften the resultant hydrogels. The products were characterized by different tools, including FTIR for confirming the chemical modification, TGA for investigating their thermal properties, and XRD for verifying their crystalline structure. The morphology of the prepared derivatives was studied through SEM, while their topography before and after dye adsorption was monitored via the AFM. The removal efficiencies of the prepared sorbents were verified at different operation conditions, such as pH, temperature, adsorbent dose, initial concentration of dye solutions, and contact time. The data revealed that the optimum conditions for maximum dye uptake were as follows: pH 4, contact time 120 min, 0.1-g sorbent dose, and 50-ppm dye concentration. Additionally, the prepared sorbents demonstrated potent adsorption capacity and removal efficiency. It was found that the elements of the second group displayed higher performance than their counterparts. The data showed also that the adsorption process best fits with the Freundlich model and obeyed pseudo-first-order kinetic isotherm. In addition, the synthesized composites showed observable antibacterial potency toward E. coli as a Gram-negative bacterium and S. aureus as a Gram-positive bacterium.

A chemical platform for the efficient screening of arylazopyrazole-based photoswitchable CENP-E inhibitors using mild cyclization reactions.

A set of arylazopyrazole-based inhibitors targeting the mitotic motor protein CENP-E was discovered through the chemical platform using the quantitative cyclization of 1,3-diketone intermediate with various hydrazines under mild conditions. Through this efficient platform, the structure-activity relationship pertaining to the pyrazole photoswitch in photoswitchable CENP-E inhibitors not only in vitro but also in cells was successfully clarified.

Critical review on electrooxidation and chemical reduction of azo dyes: Economic approach.

Effective degradation technologies have been extensively investigated and used to remove azo dyes from wastewater for decades. However, no review dealing with both electrooxidation and chemical reduction of azo dyes from an economic and, therefore, application-relevant perspective has been found in the current literature. A novelty of this review article consists not only in the brief summarization and comparison of both methods but mainly in the evaluation of their economic side. Based on the literature survey of the last 15 years, the costs of treatment approaches published in individual research articles have been summarized, and the missing data have been calculated. A broad spectrum of advanced electrode materials and catalysts have been developed and tested for the treatment, specifically aiming to enhance the degradation performance. An outline of the global prices of electrode materials, reducing agents, and basic chemicals is involved. All additional costs are described in depth in this review. The advantages and disadvantages of respective methods are discussed. It was revealed that effective and cheap treatment approaches can be found even in advanced degradation methods. Based on the collected data, electrooxidation methods offer, on average, 30 times cheaper treatment of aqueous solutions. Concerning chemical reduction, only ZVI provided high removal of azo dyes at prices <100 $ per kg of azo dye. The factors affecting total prices should also be considered. Therefore, the basic diagram of the decision-making process is proposed. In the conclusion, challenges, future perspectives, and critical findings are described.