Antiviral effects of umbelliferone against viral hemorrhagic septicemia virus in olive flounder (Paralichthys olivaceus).

Viral hemorrhagic septicemia virus (VHSV) poses a significant threat to global aquaculture, prompting ongoing efforts to identify potential drug candidates for disease prevention. Coumarin derivatives have recently emerged as a promising class of compounds effective against rhabdoviruses, which severely impact the aquaculture industry. In this study, we assessed the anti-VHSV activity of umbelliferone (7-hydroxycoumarin) in fathead minnow (FHM) cells and olive flounder Paralichthys olivaceus. Umbelliferone exhibited an EC50 of 100 µg/mL by reducing cytopathic effect, with a maximum cytotoxicity of 30.9 % at 750 µg/mL. Mechanistic analyses via a time-course plaque reduction assay revealed that direct incubation with the virus for 1 h resulted in 97.0 ± 1.8 % plaque reduction, showing excellent direct virucidal activity. Pretreatment for 4 h resulted in a 33.5 ± 7.8 % plaque reduction, which increased with longer incubation times. Cotreatment led to a 33.5 ± 2.9 % plaque reduction, suggesting interference with viral binding, whereas postinfection treatment proved less effective. Umbelliferone was prophylactically administered to the olive flounder through short-term (3 days) and long-term (14 days) medicated feeding, followed by a 4-day postinfection period. Short-term administration at 100 mg/kg body weight (bw)/day resulted in the highest relative percent survival (RPS) of 56 %, whereas long-term administration achieved a maximum RPS of 44 % at 30 mg/kg bw/day. Umbelliferone administration delayed mortality at these doses. Additionally, umbelliferone significantly inhibited the expression of the VHSV N gene during viral challenge, with no observed toxic effects in fish up to an administration dose of 30 mg/kg bw/day for 28 days. Our findings suggest that the protective mechanism of short-term administration of 100 mg umbelliferone against VHSV infection may involve the overexpression of TLR2, MDA5, STAT1, and NF-κB at 24 h postinfection (hpi). IL-8 and IFN II expression was upregulated, whereas TNF-α, IL-1ß, and IFN I expression was suppressed at 24 hpi. The upregulation of ISG15 at 48 hpi may contribute to the inhibition of VHSV replication, whereas the downregulation of Caspase 3 expression at 96 hpi suggests a possible inhibition of virus-induced apoptosis at later infection stages. Overall, umbelliferone exhibited anti-VHSV activity through multiple mechanisms, with the added advantage of convenient administration via medicated feed.

Mining for Metal-Organic Systems: Chemistry Frontiers of Th-, U-, and Zr-Materials.

The conceptual framework presented in this Perspective overviews the design principles of innovative thorium-based materials that could address urgent needs of the medicinal, nuclear energy, and waste remediation sectors from the lens of zirconium and uranium analogs. We survey the intersections of Zr, Th, and U chemistry with a focus on how the intrinsic behavior of each metal translates to broader material properties, including, but not limited to, structural and topological diversity, preferential metal-ligand binding, and reactivity. On the example of several classes of materials, including organometallic complexes, polyoxometalates, and the primary focus of this Perspective, metal-organic frameworks (MOFs), the design principles that govern the preparation of Zr-, Th-, and U-compounds, including oxophilicity, variation in oxidation states, and stable coordination environments have been considered. Further, we highlight how the impact of the mentioned variables may shift throughout the progression from discrete molecular systems to extended structures. We discuss the common assumption that zirconium-organic materials are typically considered a close analog of thorium-based congeners in areas such as material design and preparation. Through consideration of fundamental chemistry principles, we shed light on the relationships between Zr-, Th-, and U-based materials and highlight how a critical analysis of their distinct properties can be used to target a desired material performance. As a result, we provide a detailed understanding of Th-based materials chemistry by anchoring their fundamental properties between two well-studied reference points, zirconium- and uranium-containing analogs.

Clinical outcomes of colorectal neoplasm with positive resection margin after endoscopic submucosal dissection.

A positive resection margin after colorectal endoscopic submucosal dissection (ESD) is associated with an increased risk of recurrence. We aimed to identify the clinical significance of positive resection margins in colorectal neoplasms after ESD. We reviewed 632 patients who had en bloc colorectal ESD at two hospitals between 2015 and 2020. The recurrence rates and presence of residual tumor after surgery were evaluated. The rate of additional surgery after ESD and recurrence rate were significantly higher in patients with incomplete resection (n = 75) compared to patients with complete resection (n = 557). When focusing solely on non-invasive lesions, no significant differences in recurrence rates were observed between the groups with complete and incomplete resection (0.2% vs. 1.9%, p = 0.057). Among 84 patients with submucosal invasive carcinoma, 39 patients underwent additional surgery due to non-curative resection. Positive vertical margin and lymphovascular invasion were associated with residual tumor. Lymphovascular invasion was associated with lymph node metastasis. However, no residual tumor nor lymph node metastases were found in patients with only one unfavorable histological factor. In conclusion, a positive resection margin in non-invasive colorectal lesions, did not significantly impact the recurrence rate. Also, in T1 colorectal cancer with a positive vertical resection margin, salvage surgery can be considered in selected patients with additional risk factors.

Friends or Foes: Fundamental Principles of Th-Organic Scaffold Chemistry Using Zr-Analogs as a Guide.

The fundamental interest in actinide chemistry, particularly for the development of thorium-based materials, is experiencing a renaissance owing to the recent and rapidly growing attention to fuel cycle reactors, radiological daughters for nuclear medicine, and efficient nuclear stockpile development. Herein, we uncover fundamental principles of thorium chemistry on the example of Th-based extended structures such as metal-organic frameworks in comparison with the discrete systems and zirconium extended analogs, demonstrating remarkable over two-and-half-year chemical stability of Th-based frameworks as a function of metal node connectivity, amount of defects, and conformational linker rigidity through comprehensive spectroscopic and crystallographic analysis as well as theoretical modeling. Despite exceptional chemical stability, we report the first example of studies focusing on the reactivity of the most chemically stable Th-based frameworks in comparison with the discrete Th-based systems such as metal-organic complexes and a cage, contrasting multicycle recyclability and selectivity (>97%) of the extended structures in comparison with the molecular compounds. Overall, the presented work not only establishes the conceptual foundation for evaluating the capabilities of Th-based materials but also represents a milestone for their multifaceted future and foreshadows their potential to shape the next era of actinide chemistry.

Breaking the photoswitch speed limit.

The forthcoming generation of materials, including artificial muscles, recyclable and healable systems, photochromic heterogeneous catalysts, or tailorable supercapacitors, relies on the fundamental concept of rapid switching between two or more discrete forms in the solid state. Herein, we report a breakthrough in the "speed limit" of photochromic molecules on the example of sterically-demanding spiropyran derivatives through their integration within solvent-free confined space, allowing for engineering of the photoresponsive moiety environment and tailoring their photoisomerization rates. The presented conceptual approach realized through construction of the spiropyran environment results in ~1000 times switching enhancement even in the solid state compared to its behavior in solution, setting a record in the field of photochromic compounds. Moreover, integration of two distinct photochromic moieties in the same framework provided access to a dynamic range of rates as well as complementary switching in the material's optical profile, uncovering a previously inaccessible pathway for interstate rapid photoisomerization.

Cytochrome P450 Family 46 Subfamily A Member 1 Promotes the Progression of Colorectal Cancer by Inducing Tumor Cell Proliferation and Angiogenesis.

BACKGROUND/AIM: Cytochrome P450 family 46 subfamily A member 1 (CYP46A1) has been implicated in the development and progression of various cancers. This study aimed to analyze the expression of CYP46A1, examining its relationship with oncogenic behaviors, and determining its prognostic implications in colorectal cancer (CRC). MATERIALS AND METHODS: A total of 225 patients with CRC who underwent curative surgical resection were examined using paraffin-embedded tissue blocks and subjected to tumor-specific survival analysis. The expression of CYP46A1 was assessed in CRC tissues through reverse transcription-polymerase chain reaction, western blotting, and immunohistochemistry. The CRC cells' apoptosis, proliferation, angiogenesis, and lymphangiogenesis were analyzed using terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assays, alongside immunohistochemical staining for Ki-67, CD34, and D2-40 antibodies. RESULTS: CYP46A1 expression was found to be up-regulated in CRC tissues compared to normal colorectal mucosa. Such expression was significantly associated with advanced stage, deeper tumor invasion, lymph node metastasis, distant metastasis, and decreased survival. Furthermore, the mean Ki-67 labeling index and microvessel density values in CYP46A1-positive tumors were significantly elevated compared to CYP46A1-negative tumors. However, there was no discernible correlation between CYP46A1 expression and either the apoptotic index or lymphatic vessel density value. CONCLUSION: CYP46A1 promotes CRC progression, specifically through the induction of tumor cell proliferation and angiogenesis. The insights provided may hold potential implications for future therapeutic interventions targeting CYP46A1.

Symmetry-Mismatched SBU Transformation in MOFs: Postsynthetic Metal Exchange from Zn to Fe and Its Effects on Gas Adsorption and Dye Selectivity.

This research explores the alteration of metal-organic frameworks (MOFs) using a method called postsynthetic metal exchange. We focus on the shift from a Zn-based MOF containing a [Zn4O(COO)6] secondary building unit (SBU) of octahedral site symmetry (ANT-1(Zn)) to a Fe-based one with a [Fe3IIIO(COO)6]+ SBU of trigonal prismatic site symmetry (ANT-1(Fe)). The symmetry-mismatched SBU transformation cleverly maintains the MOF's overall structure by adjusting the conformation of the flexible 1,3,5-benzenetribenzoate linker to alleviate the framework strain. The process triggers a decrease in the framework volume and pore size alongside a change in the framework's charge. These alterations influence the MOF's ability to adsorb gas and dye. During the transformation, core-shell MOFs (ANT-1(Zn@Fe)) are formed as intermediate products, demonstrating unique gas sorption traits and adjusted dye adsorption preferences due to the structural modifications at the core-shell interface. Heteronuclear clusters, located at the framework interfaces, enhance the heat of CO2 adsorption. Furthermore, they also influence the selectivity of the dye size. This research provides valuable insights into fabricating novel MOFs with unique properties by modifying the SBU of a MOF with flexible organic linkers from one site symmetry to another.

An antiviral optimized extract from Sanguisorba officinalis L. roots using response surface methodology, and its efficacy in controlling viral hemorrhagic septicemia of olive flounder (Paralichthys olivaceus).

Viral hemorrhagic septicemia causes considerable economic losses for Korea's olive flounder (Paralichthys olivaceus) aquaculture farms; therefore, effective antiviral agents for controlling viral hemorrhagic septicemia virus (VHSV) infection are imperative. The present study implemented a Box-Behnken design and cytopathic reduction assay to derive an optimized extract of Sanguisorba officinalis L. roots (OE-SOR) with maximum antiviral activity against VHSV. OE-SOR prepared under optimized extraction conditions (55% ethanol concentration at 50 °C for 5 h) exhibited potent antiviral activity against VHSV, with a 50% effective 0.21 µg/mL concentration and a 340 selective index. OE-SOR also showed direct virucidal activity in the plaque reduction assay. Administering OE-SOR to olive flounder exhibited substantial efficacies against VHSV infection. Fish receiving 100 mg/kg body weight/day of OE-SOR as a preventive (40.0%; p < 0.05) or therapeutic (44.4%; p < 0.05) exhibited a higher relative survival than the untreated VHSV-infected control group (mortalities of 100% and 90%, respectively). In addition, fish fed with OE-SOR (100 mg/kg body weight/day) for two weeks conveyed a significantly higher inflammatory cytokine expression (nuclear factor kappa-light-chain-enhancer of activated B cells [NF-κB], interleukin-1 beta [IL-1ß], and tumor necrosis factor-alpha [TNF-α]) than the control group one to two days post-administration. Moreover, no hematological or histological changes were observed in olive flounder treated with OE-SOR over four weeks. Liquid chromatography-quadrupole-time of flight tandem mass spectrometry and -triple quadrupole tandem mass spectrometry analyses identified ziyuglycoside I as a prominent OE-SOR constituent and marker compound (content: 14.5%). This study verifies that OE-SOR is an effective alternative for controlling viral hemorrhagic septicemia in olive flounder farms as it exhibits efficient in vivo anti-VHSV activity and increases innate immune responses.

In-vitro immunomodulatory responses and antiviral activities of antimicrobial peptide octominin against fish pathogenic viruses.

Antimicrobial peptides (AMPs) are considered a novel approach to stimulate fish antiviral mechanisms for defense against a broad range of viral infections by enhancing immunomodulatory activities. Octominin is an AMP derived from the defense proteins of Octopus minor. In this study, preliminary screening of octominin against viral hemorrhagic septicemia virus (VHSV), infectious hematopoietic necrosis virus (IHNV), and infectious pancreatic necrosis virus (IPNV) was carried out. Moreover, immune responses upon octominin treatment and IHNV challenge were investigated using fathead minnow (FHM) cells. The CC50s of octominin for FHM and Chinook salmon embryo-214 (CHSE-214) cells were 2146.2 and 1865.2 µg/mL, respectively. With octominin treatment, EC50 resulted in 732.8, 435.1, and 925.9 µg/mL for VHSV, IHNV, and IPNV, respectively. The selectivity indices were 2.9, 4.9, and 2.0, respectively. The transcriptional analysis results demonstrated the induced transcription factors (Irf3; 143-fold, Irf7; 105-fold, and NF-κB; 8-fold), stress response gene (HspB8; 2-fold), and apoptosis functional gene (p53; 3-fold) in octominin treated (500 µg/mL) FHM cells for 48 h. Moreover, IHNV viral copy number was slightly decreased with the octominin treatment (500 µg/mL) in FHM cells. Overall results suggest that octominin could be a potential antiviral agent, although further studies are necessary to understand its mode of action and the mechanism of its antiviral activity.

Cooperative and Orthogonal Switching in the Solid State Enabled by Metal-Organic Framework Confinement Leading to a Thermo-Photochromic Platform.

Cooperative behavior and orthogonal responses of two classes of coordinatively integrated photochromic molecules towards distinct external stimuli were demonstrated on the first example of a photo-thermo-responsive hierarchical platform. Synergetic and orthogonal responses to temperature and excitation wavelength are achieved by confining the stimuli-responsive moieties within a metal-organic framework (MOF), leading to the preparation of a novel photo-thermo-responsive spiropyran-diarylethene based material. Synergistic behavior of two photoswitches enables the study of stimuli-responsive resonance energy transfer as well as control of the photoinduced charge transfer processes, milestones required to advance optoelectronics development. Spectroscopic studies in combination with theoretical modeling revealed a nonlinear effect on the material electronic structure arising from the coordinative integration of photoresponsive molecules with distinct photoisomerization mechanisms. Thus, the reported work covers multivariable facets of not only fundamental aspects of photoswitch cooperativity, but also provides a pathway to modulate photophysics and electronics of multidimensional functional materials exhibiting thermo-photochromism.

Superprotonic Conductivity of MOFs Confining Zwitterionic Sulfamic Acid as Proton Source and Conducting Medium.

A few metal-organic frameworks (MOFs), which typically use strong acids as proton sources, display superprotonic conductivity (≈10-1  S cm-1 ); however, they are rare due to the instability of MOFs in highly acidic conditions. For the first time, we report superprotonic conductivity using a moderately acidic guest, zwitterionic sulfamic acid (HSA), which is encapsulated in MOF-808 and MIL-101. HSA acts not only as a proton source but also as a proton-conducting medium due to its extensive hydrogen bonding ability and zwitterion effect. A new sustained concentration gradient method results in higher HSA encapsulation compared to conventional methods, producing 10HSA@MOF-808-(bSA)2 and 8HSA@MIL-101. These MOFs show impressive superprotonic conductivity of 2.47×10-1 and 3.06×10-1  S cm-1 , respectively, at 85 °C and 98 % relative humidity, and maintain stability for 7 days.

f-block MOFs: A Pathway to Heterometallic Transuranics.

A novel series of heterometallic f-block-frameworks including the first examples of transuranic heterometallic 238 U/239 Pu-metal-organic frameworks (MOFs) and a novel monometallic 239 Pu-analog are reported. In combination with theoretical calculations, we probed the kinetics and thermodynamics of heterometallic actinide(An)-MOF formation and reported the first value of a U-to-Th transmetallation rate. We concluded that formation of uranyl species could be a driving force for solid-state metathesis. Density of states near the Fermi edge, enthalpy of formation, band gap, proton affinity, and thermal/chemical stability were probed as a function of metal ratios. Furthermore, we achieved 97 % of the theoretical maximum capacity for An-integration. These studies shed light on fundamental aspects of actinide chemistry and also foreshadow avenues for the development of emerging classes of An-containing materials, including radioisotope thermoelectric generators or metalloradiopharmaceuticals.

Confinement-Driven Photophysics in Hydrazone-Based Hierarchical Materials.

Confinement-imposed photophysics was probed for novel stimuli-responsive hydrazone-based compounds demonstrating a conceptual difference in their behavior within 2D versus 3D porous matrices for the first time. The challenges associated with photoswitch isomerization arising from host interactions with photochromic compounds in 2D scaffolds could be overcome in 3D materials. Solution-like photoisomerization rate constants were realized for sterically demanding hydrazone derivatives in the solid state through their coordinative immobilization in 3D scaffolds. According to steady-state and time-resolved photophysical measurements and theoretical modeling, this approach provides access to hydrazone-based materials with fast photoisomerization kinetics in the solid state. Fast isomerization of integrated hydrazone derivatives allows for probing and tailoring resonance energy transfer (ET) processes as a function of excitation wavelength, providing a novel pathway for ET modulation.

Metal-Photoswitch Friendship: From Photochromic Complexes to Functional Materials.

Cooperative metal-photoswitch interfaces comprise an application-driven field which is based on strategic coupling of metal cations and organic photochromic molecules to advance the behavior of both components, resulting in dynamic molecular and material properties controlled through external stimuli. In this Perspective, we highlight the ways in which metal-photoswitch interplay can be utilized as a tool to modulate a system's physicochemical properties and performance in a variety of structural motifs, including discrete molecular complexes or cages, as well as periodic structures such as metal-organic frameworks. This Perspective starts with photochromic molecular complexes as the smallest subunit in which metal-photoswitch interactions can occur, and progresses toward functional materials. In particular, we explore the role of the metal-photoswitch relationship for gaining fundamental knowledge of switchable electronic and magnetic properties, as well as in the design of stimuli-responsive sensors, optically gated memory devices, catalysts, and photodynamic therapeutic agents. The abundance of stimuli-responsive systems in the natural world only foreshadows the creative directions that will uncover the full potential of metal-photoswitch interactions in the coming years.

Ligand functionalization of defect-engineered Ni-MOF-74.

Incorporating functionality into the framework of metal-organic frameworks (MOFs) has attracted substantial interest because the physical and chemical properties of MOFs can be tuned by functionalizing pores. The ligand functionalization of MOF-74 is challenging because of its pristine organic ligand and framework structure. Herein, we report a series of ligand-functionalized Ni-MOF-74 derivatives synthesized by defect engineering using a mixed-ligand approach. Defect generation and ligand functionalization of Ni-MOF-74 were simultaneously achieved by incorporation of fragmented organic ligands such as 5-formylsalicylic acid, 3-hydroxysalicylic acid, 2-hydroxynicotinic acid and 5-hydroxy-1H-benzimidazole-4-carboxylic acid. The resulting defect-engineered Ni-MOF-74 derivatives maintained relatively good crystallinity up to fragment incorporation levels of ∼20% and exhibited modified permanent porosity and CO2 adsorption properties depending on the functional groups and defect concentrations in the framework.

Capture Instead of Release: Defect-Modulated Radionuclide Leaching Kinetics in Metal-Organic Frameworks.

Comparison of defect-controlled leaching-kinetics modulation of metal-organic frameworks (MOFs) and porous functionalized silica-based materials was performed on the example of a radionuclide and radionuclide surrogate for the first time, revealing an unprecedented readsorption phenomenon. On a series of zirconium-based MOFs as model systems, we demonstrated the ability to capture and retain >99% of the transuranic 241Am radionuclide after 1 week of storage. We report the possibility of tailoring radionuclide release kinetics in MOFs through framework defects as a function of postsynthetically installed organic ligands including cation-chelating crown ether-based linkers. Based on comprehensive analysis using spectroscopy (EXAFS, UV-vis, FTIR, and NMR), X-ray crystallography (single crystal and powder), and theoretical calculations (nine kinetics models and structure simulations), we demonstrated the synergy of radionuclide integration methods, topological restrictions, postsynthetic scaffold modification, and defect engineering. This combination is inaccessible in any other material and highlights the advantages of using well-defined frameworks for gaining fundamental knowledge necessary for the advancement of actinide-based material development, providing a pathway for addressing upcoming challenges in the nuclear waste administration sector.

Spatial distribution modulation of mixed building blocks in metal-organic frameworks.

The placement of mixed building blocks at precise locations in metal-organic frameworks is critical to creating pore environments suitable for advanced applications. Here we show that the spatial distribution of mixed building blocks in metal-organic frameworks can be modulated by exploiting the different temperature sensitivities of the diffusion coefficients and exchange rate constants of the building blocks. By tuning the reaction temperature of the forward linker exchange from one metal-organic framework to another isoreticular metal-organic framework, core-shell microstructural and uniform microstructural metal-organic frameworks are obtained. The strategy can be extended to the fabrication of inverted core-shell microstructures and multi-shell microstructures and applied for the modulation of the spatial distribution of framework metal ions during the post-synthetic metal exchange process of a Zn-based metal-organic framework to an isostructural Ni-based metal-organic framework.

Creating Tunable Mesoporosity by Temperature-Driven Localized Crystallite Agglomeration.

A new synthetic approach for tunable mesoporous metal-organic frameworks (MeMs) is developed. In this approach, mesopores are created in the process of heat conversion of highly mosaic metal-organic framework (MOF) crystals with non-interpenetrated low-density nanocrystallites into MOF crystals with two-fold interpenetrated high-density nanocrystallites. The two-fold interpenetration reduces the volume of the nanocrystallites in the mosaic crystal, and the accompanying localized agglomeration of the nanocrystallites results in the formation of mesopores among the localized crystallite agglomerates. The pore size can be easily modulated from 7 to 90 nm by controlling the heat treatment conditions, that is, the aging temperature and aging time. Various proteins can be encapsulated in the MeM, and immobilized enzymes show catalyst activity comparable to that of the free native enzymes. Immobilized ß-galactosidase is recyclable and the enzyme activity of the immobilized catalase is maintained after exposure to high temperatures and various organic solvents.

Antibacterial Activities of Prenylated Isoflavones from Maclura tricuspidata against Fish Pathogenic Streptococcus: Their Structure-Activity Relationships and Extraction Optimization.

Streptococcus zoonotic bacteria cause serious problems in aquaculture with clinical effects on humans. A structure-antibacterial activity relationships analysis of 22 isoflavones isolated from M. tricuspidata (leaves, ripe fruits, and unripe fruits) against S. iniae revealed that prenylation of the isoflavone skeleton was an important key for their antibacterial activities (minimum inhibitory concentrations: 1.95-500 µg/mL). Through principal component analysis, characteristic prenylated isoflavones such as 6,8-diprenlygenistein (4) were identified as pivotal compounds that largely determine each part's antibacterial activities. M. tiricuspidata ripe fruits (MTF), which showed the highest antibacterial activity among the parts tested, were optimized for high antibacterial activity and low cytotoxicity on fathead minnow cells using Box-Behnken design. Optimized extraction conditions were deduced to be 50%/80 °C/7.5 h for ethanol concentration/extraction temperature/time, and OE-MTF showed contents of 6,8-diprenlygenistein (4), 2.09% with a MIC of 40 µg/mL. These results suggest that OE-MTF and its active isoflavones have promising potential as eco-friendly antibacterial agents against streptococcosis in aquaculture.

Superprotonic Conductivity of MOF-808 Achieved by Controlling the Binding Mode of Grafted Sulfamate.

A metal-organic framework (MOF) having superprotonic conductivity, MOF-808, is prepared by modulating the binding mode of the sulfamate (SA) moieties grafted onto the metal clusters. The activation of the SA-grafted MOF-808 at 150 °C changes the binding mode of the grafted SA from monodentate to bridging bidentate, thus converting the neutral amido (-S-NH2 ) moiety of the grafted SA to the more acidic cationic sulfiliminium (-S=NH2+ ) moiety. Further, the acidic sulfiliminium moiety of MOF-808-4SA-150 results in more efficient proton conduction than the amido moiety of MOF-808-4SA-60. At 60 °C and 95 % relative humidity, MOF-808-4SA-150 is found to have a proton conductivity of 7.89×10-2  S cm-1 , which is more than 30-times higher than that of MOF-808-4SA-60. Moreover, this superprotonic conductivity is well maintained over 1000 cycles of conductivity measurements and for similar cyclic measurements each day for seven days.