MgO Nanoparticles Research Articles

Sol-gel synthesis of magnesium oxide nanoparticles and their evaluation as a therapeutic agent for the treatment of osteoarthritis.

Aim: We synthesized MgO NPs via sol-gel reaction and investigated them as carriers to deliver Mg2+ to the affected joint for osteoarthritis (OA).Materials & methods: The physicochemical properties of samples were characterized by transmission electron microscope (TEM), dynamic light scattering (DLS) and x-ray diffraction (XRD). The release of Mg2+ was monitored by ICP-MS. The potential cytotoxicity was evaluated using MTT assay. The efficacy and biosafety were evaluated in a rabbit OA model.Results: MgO NPs can prolong the Mg2+ release time from 0.5 h to 12 h. No significant cytotoxicity was observed when concentrations below 250μg/ml. Intra-articular samples could effectively alleviate the degeneration and destruction of the cartilage.Conclusion: this study demonstrates the potential of MgO NPs as a safe and effective treatment of OA. Simultaneously, the size of the particles may play a significant role in influencing the therapeutic outcome.

Bond strength and surface roughness assessment of novel antimicrobial polymeric coated dental cement

Resin cement integrated with zein-incorporated magnesium oxide nanoparticles has previously been found to inhibit oral microbes and decrease bacterial biofilm. However, the bond strength and surface features of this biomaterial have yet to be investigated. The objective of this study was to evaluate the shear bond strength, mode of fracture, and surface roughness of resin cement modified with zein-incorporated magnesium oxide nanoparticles. Characterization of the cement was performed by X-ray diffraction, field emission scanning electron microscopy, and Fourier transform infrared spectroscopy. 126 human teeth were divided into 3 groups and cemented to lithium disilicate ceramic using resin cement with zein-incorporated magnesium oxide nanoparticles at concentrations of 0%, 1%, and 2% (n = 42). 21 samples of each group were subjected to the shear bond strength test, while the other 21 underwent thermocycling for 10,000 cycles before the test, after which all samples were evaluated for the mode of fracture. To assess surface roughness, resin cement disks were analyzed by a profilometer before and after undergoing thermocycling for 10,000 cycles. The shear bond strength of the cement with 1% and 2% nanoparticles was significantly higher than the control before thermocycling. The mode of fracture was found to be mainly adhesive with all groups, with the unmodified cement presenting the highest cohesive failure. There was no significant difference in surface roughness between the groups before or after thermocycling. The addition of zein-incorporated magnesium oxide nanoparticles to resin cement improved or maintained the shear bond strength and surface roughness of the resin cement.Graphical abstract

Radiation and nanoparticle interaction for enhanced light absorption and heat conversion

The photo-thermal conversion performance (PTCP) of water-based nanofluids in a volumetrically heated solar collector (VHSC) is evaluated. The influences of nanoparticle volume concentration (NVC), particle size, Reynolds number, operating temperature, and collector geometry on the PTCP are numerically investigated. The utilization of nanoparticles and improving their concentration is found to enhance the photo-thermal conversion efficiency (PTCE) of the collector by augmenting the energy from the sun of the working fluid due to the radiation-nanoparticle interactions. The PTCE enhancement of Graphite, TiO2 and Ag nanoparticles dispersed in water is respectively 1.37, 1.33 and 1.29 times better, compared to that of water. This is further augmented by adding MgO nanoparticles to TiO2, Graphite and Ag particles, resulting in 1.69, 1.67 and 1.59x improved efficiency. Enhancing the flow rate of the fluids also contributes to the PTCE by reducing the losses of thermal from the collector to the ambient. Besides, the enhanced nanoparticle size increases the overall PTCP as it enables the nanofluid to absorb more solar energy. Enhancing the collector length accelerates the thermal loss to the ambient, as a result the system performance diminishes. It is also regarded that the heat absorption of the nanofluid declines with improving inlet temperature of the working fluid.

Evaluation of Anticancer and Antibacterial Activities for Pluronic F-127 Encapsulated Magnesium Oxide Nanoparticles Using Alstoniayunnanensis Leaf Extract

Primary brain tumors caused by gliomas are most frequently found in the neurons of the central nervous system. The present study explores the effects of Pluronic F-127 coated magnesium oxide nanoparticles (PF-127 MgO NPs) using Alstonia yunnanensis leaf extract (A. yunnanensis) on C6 glioma cells (C6 cells). The vital part of nanotechnology is the formulation of environmentally friendly methods for making NPs. Because of their small size, NPs can pass easily through the holes in intracellular cell membranes and into living tissue. Numerous integrated protein therapies have shown enhanced medicinal qualities and thermo stability owing to PF-127. MgO is employed in cell therapy, tissue repair and the creation of cancer medicines. In this research, we synthesized PF-127 MgO NPs using A. yunnanensis leaf extract to study the activity of their anti-bacterial and anti-cancer activities on C6 cells. We examined the synthesized NPs by the UV-Vis, FT-IR, PL, TEM, SAED, FE-SEM, EDAX, XRD, and DLS techniques. The anti-bacterial effectiveness and anti-cancer activity C6 cells on MTT assay, AO/EtBr, DAPI, DCFH-DA and Rh-123 staining methods of synthesized PF-127 MgO NPs. Moreover, the synthesized NPs characterization studies can be used high activity anti-bacterial and significant anti-cancer activity were noticed. The study finds that the NPs increased ROS accumulation and resulting in apoptotic cell death. Overall, our findings show that synthesized PF-127 coated MgO NPs using A. yunnanensis leaf extract possess anti-bacterial and anti-cancer properties in C6 cells.

Biomedical potential of green-engineered chitosan-magnesium oxide nanoparticles: An in vitro study on antibacterial and anticancer activities

The development of low-cost, highly effective, environmentally friendly, multi-functional bio-cidal substances has become a top priority in the current scenario. This study aimed to create low-cost materials, such as green-engineered magnesium oxide (GEMgO) and chitosan-coated magnesium oxide (GECsMgO) nanoparticles (NPs), using Chara zeylanica Klein ex Willd extract as a reducing agent. The X-ray diffraction results confirmed the cubic face-centered structure of the GEMgO and GECsMgO NPs. The field emission scanning electron microscopy and transmission electron microscopy results showed that GEMgO and GECsMgO NPs exhibit a spherical structure and nanoflakes with spherical structures, respectively. The Mg–O stretching frequencies were measured at 779 and 456 cm−1 for GEMgO and 543 cm−1 for GECsMgO NPs in the Fourier-transform infrared spectra. The oxygen vacancies responsible for the biocidal activities of GEMgO and GEC-MgO NPs were observed in their photoluminescence spectra at 501 and 520 nm and at 506 and 524 nm, respectively. The antioxidant activity of GEMgO and GECsMgO NPs was investigated using a DPPH assay. The GECsMgO NPs exhibited higher scavenging activity than the GEMgO NPs. Antibacterial studies on gram-negative bacteria treated with GEMgO and GECsMgO NPs showed that GECsMgO NPs exhibited the highest antibacterial activity. The ultrastructural morphological results of E. coli showed that GECsMgO NPs can damage it better than GEMgO NPs. An MTT assay determined that GEC-MgO NPs had better anticancer properties than GEMgO NPs when tested against a blood cancer cell line (MOLT-7). Fibroblast L929 cell toxicity tests showed that GEC-MgO NPs were less toxic than GEMgO NPs. In the future, GECsMgO NPs can be used as biocidal agents in clinical and industrial applications.

Phytoassisted synthesis, characterization, and evaluation of biological properties of magnesium oxide nanoparticles obtained using Cinnamomum tamala leaf extracts

Phytoassisted synthesis, characterization, and evaluation of biological properties of magnesium oxide nanoparticles obtained using Cinnamomum tamala leaf extracts

Adsorptive removal of phosphate from water with biochar from acacia tree modified with iron and magnesium oxides

A novel biochar (BC) from Acaciatortilis trees pruning waste was synthesized and tested for the removal of phosphate from aqueous solutions. The BC was prepared by calcination at 600 °C and doped with Fe3O4 and MgO by hydrothermal process. The presence of iron and magnesium ions in the modified BC was confirmed by EDS analysis and X-ray diffraction (XRD) methods. Both unmodified and doped BCs were tested for phosphate removal from synthetic 1–500 ppm aqueous solutions. While the unmodified BC did not show any significant removal of phosphate from aqueous solutions, the modified BC almost completely removed phosphate from water. The enhancement in removal efficiency is due to an increase in the overall surface charge and surface area of BC as a result of doping with Fe3O4 and MgO salts. The average porosity and BET surface area corresponding to the plain BC increased by more than 20% from 322 to 394 m2/g after modification by impregnation with iron oxide and magnesium oxide. The modificaiton of BC with Fe3O4 and MgO nanoparticles was observed to increase the point of zero electric charge (PZC) from pH 3.4 (corresponding to plain BC) to pH 5.3 (corresponding to modified BC). The adsorption process was very fast and a phosphate removal value of 82.5% was reached only after 30 min of adsorption, while the removal efficiency after 4 h of adsorption was 97.5%. The rapid removal efficiency in short contact time is attributed to the high surface area of BC and strong bonding between the modified BC surface and PO43− ions. The highest adsorption capacity was observed to correspond to 98.5 mg/g which was achieved at PO43− concentration of 500 ppm and pH 8.5. Moreover, after fitting the adsorption data onto four of the most widely used adsorption isotherm models, the adsorption of PO43− onto BC can be better described by the Langmuir isotherm model.

Nanoparticle agglomeration in molten salt nanofluids: Free energy landscape and its impacts on thermal transport property degradation

The nanofluids consisting of molten salt and nanoparticles can gain enhanced thermal performance for heat transfer and energy storage processes. Agglomeration of nanoparticles is ubiquitous in this colloidal dispersion, deteriorating the dispersed states of the nanocomposite interiors, and its microscopic mechanisms are unsatisfactorily explored. In this work, molecular dynamics simulations combined with umbrella sampling technique to capture the relative dispersion state of nanoparticles in molten salts effectively were performed to reveal the free energy landscape of agglomeration and its influence on heat transport. The dominant deep minima in the free energy surface demonstrates that the agglomeration of MgO nanoparticles in the molten NaCl salt is principally induced by van der Waals attractions between MgO nanoparticles and aggravated by large surface area. The unstable dispersion of nanoparticles is the most fatal problem for enhanced thermal conductivity during agglomeration, and the relative distance of MgO nanoparticles has less impacts on the thermal conductivity of the melt. The stronger adsorption of ions around nanoparticles can improve their relative dispersion, and utilize interfacial effects to enhance the thermal conductivity. Combined with the benefit of ordered ionic layer on the surface, the MgO nanoparticles of 10 Å diameter (specific surface area of ∼ 938.80 m2/g) become the optimal choice for molten NaCl. With the mechanisms of interfacial microstructures preventing agglomerated nanoparticles and improving heat transfer of molten salts revealed, this work can enlighten the selection or feasible modification for nanoparticles in various high-temperature molten salts, since the mechanisms are universal to other nanofluids, especially for ionic liquid based nanofluids.

Maximizing dental composite performance: Strength and hardness enhanced by innovative polymer-coated MgO nanoparticles

IntroductionZein-incorporated magnesium oxide nanoparticles (zMgO NPs) can influence the mechanical properties of dental materials. However, the effect of this addition on the mechanical properties of resin composite has yet to be investigated. The objective of this study was to add various concentrations of zMgO NPs to conventional, flowable, and bulk-fill composite and assess the effect on the compressive strength, flexural strength, and microhardness. Methodology150 samples each of conventional composite, flowable composite, and bulk-fill composite (n = 450) were enhanced with concentrations of zMgO NPs at 0 %, 0.3 %, 0.5 %, 1 %, and 2 % (n = 30). 10 samples of each group were randomly allotted to the compressive strength, flexural strength, or hardness test. Characterization of the specimens was performed by X-ray diffraction, Fourier transform infrared spectroscopy, scanning electron microscopy, energy-dispersive X-ray spectroscopy, and transmission electron microscopy. Two-way ANOVA test was used to compare between groups, and one-way ANOVA followed by Tukey's test was done at p = 0.05 to determine significance. ResultsCharacterization yielded a uniform distribution of nanoparticles in the matrix and the formation of a new hybrid composite that maintained its properties. Composite of all types enhanced with 0.3 % and 0.5 % zMgO NPs demonstrated a statistically significant increase in compressive strength, flexural strength, and hardness when compared to the control (p < 0.05). The bulk-fill composite with zMgO NPs concentrations of all groups demonstrated a statistically significant increase (p < 0.05) in hardness when compared to the control. ConclusionThe modified composites’ compressive strength, flexural strength, and hardness improved or remained consistent. Clinical significanceAn improved dental resin composite will enhance the quality of care and patient experience. The augmented strength and hardness of resin composite is desirable in prolonging the durability of the restoration.

Synthesis and Characterization of Biogenic Magnesium Oxide Nanoparticles as Anticancer Agent

Synthesis and Characterization of Biogenic Magnesium Oxide Nanoparticles as Anticancer Agent

Exploring Listeria monocytogenes in Ewe Milk: ssrA Gene-based Real-time PCR Identification, Phylogenetic Analysis, and Antibacterial Assessment of Magnesium Oxide Nanoparticles Synthesized with Myrtus communis Leaf Extract

Background: Listeria monocytogenes, a zoonotic pathogen affecting humans and animals, exhibits a global distribution, including Iraq. This study focused on the rapid identification and phylogenetic analysis of L. monocytogenes in freshly collected ewe milk samples from various farms in Al-Qadisiyah province, Iraq.Methods: The study was conducted with care and precision, involving 150 milk samples. These samples were subjected to traditional bacterial isolation and identification using the enrichment culture method and biochemical tests, with the PCR technique confirming the results. The antibacterial activity of MgONPs was then assessed using the disc diffusion method, ensuring a comprehensive and reliable approach to the study.Result: The results show that 150 ewe milk samples underwent real-time PCR (RT-PCR) targeting the ssrA gene, followed by partial 16S rRNA gene sequencing (PSGS) of purified conventional PCR products. Furthermore, the study entails the biosynthesis of magnesium oxide nanoparticles using Myrtus communis leaf extract, followed by a comprehensive characterization utilizing UV-spectra, FTIR, SEM, and TEM techniques. The Agar well diffusion method assessed the antibacterial efficacy of these Biosynthesized nanoparticles against L. monocytogenes. The RT-PCR results revealed the presence of L. monocytogenes in 36 out of 150 samples (24%). Subsequent PCR analysis confirmed the presence of the pathogen in 30 out of these 36 positive samples (83.33%). Sequencing of two purified PCR products demonstrated 100% nucleotide identity with global isolates from Iraq and Turkey. Furthermore, the study demonstrated that L. monocytogenes exhibited substantial sensitivity (24.66 ± 0.3) to the biosynthesized magnesium oxide nanoparticles. These findings underscore the speed and precision of the RT-PCR method for detecting L. monocytogenes in fresh ewe milk samples.Conclusion: This comprehensive investigation enhances our understanding of L. monocytogenes prevalence in ewe milk and highlights the potential of Myrtus communis -derived nanoparticles for combating this pathogen.Keywords: Antibacterial nanoparticles; Listeria monocytogenes; Magnesium oxide; Myrtus communis sheep milk; ssrA gene

Microstructural and Optical Effects of Zn-Doped Magnesium Oxide Nanoparticles Obtained by the Precipitation Method

Microstructural and Optical Effects of Zn-Doped Magnesium Oxide Nanoparticles Obtained by the Precipitation Method

Green Synthesis of MgO Nano Particle Loaded Onto Carbon for Effective Rhodamine B Dye Removal

ABSTRACTIn this study, Rhodamine B dye (RhB) is effectively removed from aqueous solutions by using nano‐MgO and nano‐MgO activated carbon as an adsorbent. First, potassium hydroxide was used in a chemical activation process to create activated carbon from the Anacardium occidentale shell, often known as the cashew nutshell. Rosa cymose extract was used in a quick precipitation process to create nano‐magnesium oxide in a sustainable way. Activated carbon composite impregnated with nano‐magnesium oxide was made using a dropwise process. The study examined the nanocomposite that removed the dye Rhodamine B from the aqueous solution. Using SEM, XRD, FTIR, and EDX, the nano‐Mgo and nano‐MgO‐AC were analyzed. Using a scanning electron microscope, an analysis was conducted on the evenly distributed accumulation of MgO nanoparticles added to the activated carbon. The capability of nano‐MgO‐activated carbon to decolorize RhB was investigated. The effects of beginning pH ranges of 2.0–9.0, initial dye concentrations of 10–40 ppm, biosorbent dosages of 0.2–1.2 g, and contact times ranging from 10 to 60 min were investigated. At pH 5, most dye was eliminated. The work has shown that RhB may be effectively removed from aqueous medium using nano‐MgO‐AC, it could potentially be used as an affordable adsorbent material. Equilibrium estimations were acknowledged strongly through Langmuir approximations with a correlation determination of 0.985.

An in-depth investigation into the utilization of nettle fiber-reinforced epoxy composites with embedded MgO nanoparticles

An in-depth investigation into the utilization of nettle fiber-reinforced epoxy composites with embedded MgO nanoparticles

Temporal and Dosage Impact of Magnesium Oxide Nanoparticles on Grass Carp: Unveiling Oxidative Stress, DNA Damage and Antioxidant Suppression

Magnesium oxide nanoparticles (MgO NPs) have gained significant importance in biomedicine and variety of nanotechnology-based materials used in the agriculture and biomedical industries. However, the release of different nanowastes in the water ecosystem becomes a serious concern. Therefore, this study was executed to evaluate the toxic impacts of MgO NPs on grass carp. A total of 60 grass carp were randomly divided in three groups (G0, G1, and G2). Fish reared in group G0 were kept as control while fish of groups G1 and G2 were exposed to 0.5mg/L and 0.7mg/L MgO NPs respectively, mixed in water for 21 days. The 96h median lethal concentration (LC50) of MgO NPs was found to be 4.5mg/L. Evaluation of oxidative stress biomarkers, antioxidant enzymes, DNA damage in different visceral organs and the presence of micronuclei in erythrocytes were determined on days-7, 14, and 21 of the trial. Results revealed a dose and time-dependent significantly increased values of reactive oxygen species, lipid peroxidation product, DNA damage in multiple visceral organs and formation of micronuclei in the erythrocytes of treated fish (0.7mg/L). The results on antioxidant profile exhibited significantly lower amounts of total proteins, catalase, superoxide dismutase and peroxidase in visceral organs of the fish exposed to MgO NPs (0.5 and 0.7mg/L) at day 21 of trial compared to control group. In conclusion, it has been recorded that MgO NPs severely influence the normal physiological functions of the grass carp even at low doses.

Erratum for the Research Article: "Engineered MgO nanoparticles for cartilage-bone synergistic therapy" by Zheng et al.

Erratum for the Research Article: "Engineered MgO nanoparticles for cartilage-bone synergistic therapy" by Zheng et al.

Green synthesis of Vitis vinifera extract-appended magnesium oxide NPs for biomedical applications

Abstract Biologically active magnesium oxide (MgO) nanoparticles were synthesised using green reduction with an extract derived from the Vitis vinifera plant. The investigation focused on examining the structure and carbon abundance resulting from the thermal degradation of adsorbed biomolecules. It was accomplished using powder X-ray diffraction, Raman spectroscopy, and FT-IR analysis techniques. X-ray photoelectron spectroscopy studies conducted on MgO nanoparticles indicate the absence of any supplementary peaks, thereby indicating the purity of the material. The morphological characteristics, which have been examined using field emission scanning electron microscopy and TEM methodologies, demonstrate the presence of particles with a spherical shape, exhibiting minimal agglomeration and a uniform distribution across the surfaces of MgO. The porous structure, porosity, and pore volume of the MgO particles were evaluated using Brunauer-Emmett-Teller surface analysis. The experimental findings reveal that the surface area of the MgO nanoparticles is 23.8742 m2/g, while the total pore volume is 0.12528 cm3/g. Additionally, the average pore diameter is determined to be 1.7 nm. These observations collectively suggest the presence of microporous structures within the MgO nanoparticles. This article discusses the biological studies to assess the antibacterial, antifungal, anti-inflammatory, and anti-diabetic activities of the synthesised MgO nanoparticles.

XPS studies and ferromagnetism evaluations of pure MgO and Fe- and Co-doped MgO nanoparticles

Magnesium oxide nanoparticles (MgO-NPs), as well as Fe- and Co-doped MgO-NPs, were synthesized using a gelatin-based sol–gel method. The structure and morphology of the prepared samples were examined using X-ray diffraction (XRD) and transmission electron microscopy (TEM), respectively. The results indicated that pure and doped MgO-NPs, with particle sizes below 100 nm, crystallized in cubic structures. XRD data were analyzed using the size-strain plot (SSP) method, revealing that doping increased the lattice strain to 0.11 for pure MgO, 0.18 for Co-doped, and 0.13 for Fe-doped MgO nanoparticles. The crystallite sizes were found to be 60 nm for pure MgO, 48 nm for Co-doped, and 60 nm for Fe-doped MgO nanoparticles. X-ray photoelectron spectroscopy (XPS) was used to investigate the oxidation states of the prepared samples. The Fe-doped MgO-NPs exhibited the highest value of oxygen vacancy states, indicating more defects compared to the other samples. The saturation magnetization (Ms) was measured to be 0.026 emu/g for pure MgO, 0.45 emu/g for Co-doped, and 0.62 emu/g for Fe-doped MgO nanoparticles.

Aloe vera-assisted biogenic synthesis of tunable magnesium oxide nanoparticles for enhanced photocatalytic and antibacterial applications

In this study, magnesium oxide (MgO) nanoparticles (NPs) tuned via doping with transition metal ions, Co2+/Cu2+/Zn2+ have been synthesized biologically using Aloe vera leaf extract as an encapsulating agent. These pure and doped MgO NPs have been characterized by using various physico-chemical characterization techniques. X-ray diffraction (XRD) analysis confirms the formation of face-centered cubic structures of pure and doped MgO NPs with their average crystallite sizes ranging from 8.11-17.83 nm. X-ray photoelectron spectroscopic (XPS) analysis confirmed the presence of doped element in their +2 oxidation state. UV-Visible spectral analysis revealed a significant reduction in band gap energies in nano-dimensions which further gets modified upon doping (1.76 − 4.9 eV). Our findings also demonstrated the engagement of phytochemicals as capping agents in Fourier transform infrared (FTIR) examination. Scanning electron microscopic (SEM) and Transmission electron microscopic (TEM) analysis provided insights into the morphologies and particle sizes. Elemental analysis by Energy dispersive X-ray (EDX) confirms the presence of Co, Cu, and Zn elements in doped MgO NPs. The photocatalytic degradation of dyes has confirmed that Co and Cu doping substantially enhance the photocatalytic activity compared to both undoped and Zn-doped MgO NPs. Furthermore, these nanoparticles demonstrate superior efficiency in degrading the cationic dye Malachite green (MG) compared to the anionic dye Methyl orange (MO). Regarding their antibacterial properties, only Cu doping induces a notable antibacterial effect in MgO NPs, particularly against gram-positive bacteria. Additionally, these nanoparticles exhibit substantial antioxidant activities. These tailored MgO nanoparticles exhibit potential for applications in wastewater treatment.

Simultaneous removal of diclofenac sodium, cadmium and bacterial inactivation from aqueous solutions by activated MgO nanoparticles

This study investigated whether MgO nanoparticles (NPs) produced via the sol–gel process can simultaneously remove the pharmaceutical diclofenac sodium (DCF) and cadmium ions (Cd2+) from water. The UV spectroscopy, Fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), scanning electron microscopy (SEM), transmission electron microscopy (TEM), and energy-dispersive X-ray spectroscopy (EDX) were used to confirm the composition, morphology and particle size of the MgO NPs. The adsorption performance of MgO NPs was compared to that of bentonite-nano zero valent iron (B-nZVI) and activated carbon (AC). The impact of contact time, adsorbate concentration, pH, and temperature on the removal efficacy of DCF and Cd2+ via MgO NPs in aqueous solutions were investigated, and the removal data was best described by the kinetic model of pseudo-second-order and the Langmuir isotherm model (R 2 > 0.996). The maximum removal capacity of MgO for DCF and Cd2+ at pH 7 was 66.2 mg.g−1 (%R = 90) and 50.8 mg.g−1 (%R = 80), respectively. The average adsorption enthalpy for DCF and Cd2+ (ΔH° 50 kJ. mol−1) demonstrates the occurrence of physical adsorption process, while the negative value of ΔG° reveals the spontaneous nature of DCF and Cd2+ adsorption on the surface of MgO NPs. The DCF and Cd2+ adsorbed MgO surfaces were reused against various gram-positive bacteria. Finally, it was shown that MgO and sand column were more effective for removal of persistent substances and heavy metal residues than a column packed by a mixture of B-nZVI and sand.