Spatial distribution and source identification of metal contaminants in soil and rice grain samples: a study on exploration of soil quality and risk assessment.

The present study aims to assess soil quality and potential health risks associated with soil pollution of the Batala region of Punjab, India. Physico-chemical parameters such as pH (6.69-7.43), electrical conductivity (0.17-0.33 mS/cm), and total organic carbon (1.01-5.94%) were observed to be within permissible limits. The maximum mean content (mg/kg) of heavy metals in soil was found as Fe (4060.93), Zn (444.33), Mn (278.5), Pb (23.16), Cu (21.78), Ni (20.16), Co (7.14), and Cd (1.85) which were below the prescribed limits but beyond the geochemical background limits of world soil. For rice grain samples, metal content (mg/kg) was seen as Fe (307.01) > Zn (12.41) > Mn (7.43) > Cu (4.57) and was below the permissible limits. The mean bioaccumulation factor for various metals was in the order as Zn > Cu > Fe > Mn. Single and integrated soil pollution indices revealed that among 18 sites, six were highly contaminated. The ecological risk index (Er) has shown that contamination of soil with Cd, Zn, and Ni was higher than that of other metals studied. The estimated daily intake of metal (EDI), hazard quotient (HQ), and hazard index (HI) were higher for children than those for adults. Spatial variability based on metal pollution load and soil quality was also determined using cluster analysis (CA) and principal component analysis (PCA). During CA, soil samples from 18 sites formed three statistically significant clusters based on the level of metal pollution at the specific site. PCA showed that all variables were reduced into two main components 1 and 2 with eigenvalues as 3.82 (47% variance) and 1.53 (19.7% variance), respectively.

Highly Integrated µGC Based on a Multisensing Progressive Cellular Architecture with a Valveless Sample Inlet.

Microscale gas chromatographs (µGCs) promise in-field analysis of volatile organic compounds (VOCs) in environmental and industrial monitoring, healthcare, and homeland security applications. As a step toward addressing challenges with performance and manufacturability, this study reports a highly integrated monolithic chip implementing a multisensing progressive cellular architecture. This architecture incorporates three µGC cells that are customized for different ranges of analyte volatility; each cell includes a preconcentrator and separation column, two complementary capacitive detectors, and a photoionization detector (PID). An on-chip carrier gas filter scrubs ambient air for the analysis. The monolithic chip, with all 16 components, is 40.3 × 55.7 mm2 in footprint. To accommodate surface adsorptive and low-volatility analytes, the architecture eliminates the commonly used inlet valve, eliminating the need for chemically inactive surfaces in the valves and pumps, allowing the use of standard parts. Representative analysis is demonstrated from a nonpolar 14-analyte mixture, a polar 12-analyte mixture, and a 3-phosphonate ester mixture, covering a wide vapor pressure range (0.005-68.5 kPa) and dielectric constant range (1.8-23.2). The three types of detectors show highly complementary responses. Quantitative analysis is shown in the tens to hundreds ppb range. With 200 mL samples, the projected detection limits reach 0.12-4.7 ppb. Limited tests performed at 80% humidity showed that the analytes with vapor pressures <12 kPa were unaffected. A typical full run takes 28 min and consumes 2.3 kJ energy for the fluidic elements (excluding electronics). By eliminating chip-to-chip fluidic interconnections and requiring just one custom-fabricated element, this work presents a path toward high-performance and highly manufacturable µGCs.

A multi-plex protein expression system for production of complex enzyme formulations in Trichoderma reesei.

Heterologous protein production has been challenging in the hyper-cellulolytic fungus, Trichoderma reesei as the species is known for poor transformation efficiency, low homologous recombination frequency, and marginal screening systems for the identification of successful transformants. We have applied the 2A-peptide multi-gene expression system to co-express four proteins, which include three cellulases: a cellobiohydrolase (CBH1), an endoglucanase (EG1), and a ß-D-glucosidase (BGL1), as well as the enhanced green fluorescent protein (eGFP) marker protein. We designed a new chassis vector, pTrEno-4X-2A, for this work. Expression of these cellulase enzymes was confirmed by real-time quantitative reverse transcription PCR and immunoblot analysis. The activity of each cellulase was assessed using chromogenic substrates, which confirmed the functionality of the enzymes. Expression and activity of these enzymes were proportional to the level of eGFP fluorescence, thereby validating the reliability of this screening technique. An 18-fold differencein protein expression was observed between the first and third genes within the 2A-peptide construct. The availability of this new multi-gene expression and screening tool is expected to greatly impact multi-enzyme applications, such as the production of complex commercial enzyme formulations and metabolic pathway enzymes, especially those destined for cell-free applications.

Passive Wireless Pressure Gradient Measurement System for Fluid Flow Analysis.

Using distributed MEMS pressure sensors to measure small flow rates in high resistance fluidic channels is fraught with challenges far beyond the performance of the pressure sensing element. In a typical core-flood experiment, which may last several months, flow-induced pressure gradients are generated in porous rock core samples wrapped in a polymer sheath. Measuring these pressure gradients along the flow path requires high resolution pressure measurement while contending with difficult test conditions such as large bias pressures (up to 20 bar) and temperatures (up to 125 °C), as well as the presence of corrosive fluids. This work is directed at a system for using passive wireless inductive-capacitive (LC) pressure sensors that are distributed along the flow path to measure the pressure gradient. The sensors are wirelessly interrogated with readout electronics placed exterior to the polymer sheath for continuous monitoring of experiments. Using microfabricated pressure sensors that are smaller than ø15 × 3.0 mm3, an LC sensor design model for minimizing pressure resolution, accounting for sensor packaging and environmental artifacts is investigated and experimentally validated. A test setup, built to provide fluid-flow pressure differentials to LC sensors with conditions that mimic placement of the sensors within the wall of the sheath, is used to test the system. Experimental results show the microsystem operating over full-scale pressure range of 20,700 mbar and temperatures up to 125 °C, while achieving pressure resolution of <1 mbar, and resolving gradients of 10-30 mL/min, which are typical in core-flood experiments.

Post-Translational Modification, Phase Separation, and Robust Gene Transcription.

A few recent reports reveal fundamental new insights into the intricate regulatory mechanisms that govern RNA polymerase II (Pol II)-mediated gene transcription. Whereas a histidine-rich domain (HRD) triggers phase separation, promoting transcription elongation, a phosphatase switch promotes transcription termination. A paradigm that might govern the underlying mechanisms leading to robust gene transcription is now starting to emerge.

Lipase-mediated multicomponent synthesis of 1H-Pyrazolo[1,2-b]phthalazine-5,10-dione derivatives in a binary solvent medium.

A new method for the synthesis of 1H-pyrazolo[1,2-b]phthalazine-5,10-dione derivatives via lipase from the Aspergillus niger-catalyzed multicomponent reaction of aldehydes, malononitrile, and phthalhydrazide is reported herein for the first time. This novel method holds several advantages, including its efficiency, environmental friendliness, simple workup procedure, and good yield (70-86%). The effects of temperature, organic solvents, and water content were investigated. This protocol has the potential to replace traditional chemical synthesis routes for the synthesis of nitrogen-containing heterocyclic compounds.

Effect of Pd-Sensitization on Poisonous Chlorine Gas Detection Ability of TiO2: Green Synthesis and Low-Temperature Operation.

Ganoderma lucidum mushroom-mediated green synthesis of nanocrystalline titanium dioxide (TiO2) is explored via a low-temperature (≤70 °C) wet chemical method. The role of Ganoderma lucidum mushroom extract in the reaction is to release the ganoderic acid molecules that tend to bind to the Ti4+ metal ions to form a titanium-ganoderic acid intermediate complex for obtaining TiO2 nanocrystallites (NCs), which is quite novel, considering the recent advances in fabricated gas sensing materials. The X-ray powder diffraction, field emission scanning electron microscopy, Raman spectroscopy, and Brunauer-Emmett-Teller measurements etc., are used to characterize the crystal structure, surface morphology, and surface area of as-synthesized TiO2 and Pd-TiO2 sensors, respectively. The chlorine (Cl2) gas sensing properties are investigated from a lower range of 5 ppm to a higher range of 400 ppm. In addition to excellent response-recovery time, good selectivity, constant repeatability, as well as chemical stability, the gas sensor efficiency of the as-synthesized Pd-TiO2 NC sensor is better (136% response at 150 °C operating temperature) than the TiO2 NC sensor (57% at 250 °C operating temperature) measured at 100 ppm (Cl2) gas concentration, suggesting that the green synthesized Pd-TiO2 sensor demonstrates efficient Cl2 gas sensing properties at low operating temperatures over pristine ones.

Progressive Cellular Architecture in Microscale Gas Chromatography for Broad Chemical Analyses.

Gas chromatography is widely used to identify and quantify volatile organic compounds for applications ranging from environmental monitoring to homeland security. We investigate a new architecture for microfabricated gas chromatography systems that can significantly improve the range, speed, and efficiency of such systems. By using a cellular approach, it performs a partial separation of analytes even as the sampling is being performed. The subsequent separation step is then rapidly performed within each cell. The cells, each of which contains a preconcentrator and separation column, are arranged in progression of retentiveness. While accommodating a wide range of analytes, this progressive cellular architecture (PCA) also provides a pathway to improving energy efficiency and lifetime by reducing the need for heating the separation columns. As a proof of concept, a three-cell subsystem (PCA3mv) has been built; it incorporates a number of microfabricated components, including preconcentrators, separation columns, valves, connectors, and a carrier gas filter. The preconcentrator and separation column of each cell are monolithically implemented as a single chip that has a footprint of 1.8 × 5.2 cm2. This subsystem also incorporates two manifold arrays of microfabricated valves, each of which has a footprint of 1.3 × 1.4 cm2. Operated together with a commercial flame ionization detector, the subsystem has been tested against polar and nonpolar analytes (including alkanes, alcohols, aromatics, and phosphonate esters) over a molecular weight range of 32-212 g/mol and a vapor pressure range of 0.005-231 mmHg. The separations require an average column temperature of 63-68 °C within a duration of 12 min, and provide separation resolutions >2 for any two homologues that differ by one methyl group.

A study on water quality monitoring of Buddha Nullah, Ludhiana, Punjab (India).

Diverse genotoxic agents, entering the aquatic environment through natural and anthropogenic events, pose serious threats to its biotic components. The present study involves the monitoring of water quality by assessing the genotoxic effects and physico-chemical parameters including heavy metals of 10 surface water samples collected from different locations of Buddha Nullah, a tributary of Sutlej flowing through Ludhiana, Punjab (India). Genotoxicity was evaluated following Allium cepa root chromosomal aberration assay and DNA nicking assay using plasmid (pBR322) whilst the metal (cadmium, chromium, cobalt, copper, lead, nickel and zinc) analysis was conducted using atomic absorption spectrophotometer. All water samples collected from the study area had cobalt and lead content more than the permissible limits (0.04 and 0.01, respectively) recommended by the Bureau of Indian Standards and the World Health Organization. The samples also induced genotoxicity following both bioassays. The water samples collected from Gaunspur (GP), a site approx. 75.53 km upstream of the Sutlej-Buddha Nullah joining point, has shown the maximum genotoxic effect, i.e. 38.62% in terms of per cent total aberrant cells during A. cepa assay and 100% DNA damage during DNA nicking assay. The Pearson correlation indicated that genotoxicity had a significant positive correlation with the content of cobalt (at p ≤ 0.5). During cluster analysis, the samples from 10 sites formed four statistically significant clusters based on the level of pollution that was dependent on two factors like similarity in physico-chemical characteristics and source of pollution at a specific site.

Allosteric Motions of the CRISPR-Cas9 HNH Nuclease Probed by NMR and Molecular Dynamics.

CRISPR-Cas9 is a widely employed genome-editing tool with functionality reliant on the ability of the Cas9 endonuclease to introduce site-specific breaks in double-stranded DNA. In this system, an intriguing allosteric communication has been suggested to control its DNA cleavage activity through flexibility of the catalytic HNH domain. Here, solution NMR experiments and a novel Gaussian-accelerated molecular dynamics (GaMD) simulation method are used to capture the structural and dynamic determinants of allosteric signaling within the HNH domain. We reveal the existence of a millisecond time scale dynamic pathway that spans HNH from the region interfacing the adjacent RuvC nuclease and propagates up to the DNA recognition lobe in full-length CRISPR-Cas9. These findings reveal a potential route of signal transduction within the CRISPR-Cas9 HNH nuclease, advancing our understanding of the allosteric pathway of activation. Further, considering the role of allosteric signaling in the specificity of CRISPR-Cas9, this work poses the mechanistic basis for novel engineering efforts aimed at improving its genome-editing capability.

Molecular Dynamics Reveals a DNA-Induced Dynamic Switch Triggering Activation of CRISPR-Cas12a.

CRISPR-Cas12a is a genome-editing system, recently also harnessed for nucleic acid detection, which is promising for the diagnosis of the SARS-CoV-2 coronavirus through the DETECTR technology. Here, a collective ensemble of multimicrosecond molecular dynamics characterizes the key dynamic determinants allowing nucleic acid processing in CRISPR-Cas12a. We show that DNA binding induces a switch in the conformational dynamics of Cas12a, which results in the activation of the peripheral REC2 and Nuc domains to enable cleavage of nucleic acids. The simulations reveal that large-amplitude motions of the Nuc domain could favor the conformational activation of the system toward DNA cleavages. In this process, the REC lobe plays a critical role. Accordingly, the joint dynamics of REC and Nuc shows the tendency to prime the conformational transition of the DNA target strand toward the catalytic site. Most notably, the highly coupled dynamics of the REC2 region and Nuc domain suggests that REC2 could act as a regulator of the Nuc function, similar to what was observed previously for the HNH domain in the CRISPR-associated nuclease Cas9. These mutual domain dynamics could be critical for the nonspecific binding of DNA and thereby for the underlying mechanistic functioning of the DETECTR technology. Considering that REC is a key determinant in the system's specificity, our findings provide a rational basis for future biophysical studies aimed at characterizing its function in CRISPR-Cas12a. Overall, our outcomes advance our mechanistic understanding of CRISPR-Cas12a and provide grounds for novel engineering efforts to improve genome editing and viral detection.

Exploration of highly selective fluorogenic 'on-off' chemosensor for H2 PO4 - ions: ICT-based sensing and ATPase activity profiling.

In this study, the recognition contour of Chemosensor 1 was investigated using semiaqueous methanol (XH , mole fraction = 0.31) for a range of anions and bioactive species. Host-receptor signalling based on the internal charge transfer mechanism for Chemosensor 1 was explored and reported. Structure of Chemosensor 1 and its plausible anion coordination based on hydrogen bonding is complemented with density functional theory. Consequently, we investigated the applicability of the synthesized probe in blood plasma, urine, tap water samples, and for monitoring of ATP in lysosomes by apyrase enzyme.

Structural insights into GDP-mediated regulation of a bacterial acyl-CoA thioesterase.

Thioesterases catalyze the cleavage of thioester bonds within many activated fatty acids and acyl-CoA substrates. They are expressed ubiquitously in both prokaryotes and eukaryotes and are subdivided into 25 thioesterase families according to their catalytic active site, protein oligomerization, and substrate specificity. Although many of these enzyme families are well-characterized in terms of function and substrate specificity, regulation across most thioesterase families is poorly understood. Here, we characterized a TE6 thioesterase from the bacterium Neisseria meningitidis Structural analysis with X-ray crystallographic diffraction data to 2.0-Å revealed that each protein subunit harbors a hot dog-fold and that the TE6 enzyme forms a hexamer with D3 symmetry. An assessment of thioesterase activity against a range of acyl-CoA substrates revealed the greatest activity against acetyl-CoA, and structure-guided mutagenesis of putative active site residues identified Asn24 and Asp39 as being essential for activity. Our structural analysis revealed that six GDP nucleotides bound the enzyme in close proximity to an intersubunit disulfide bond interactions that covalently link thioesterase domains in a double hot dog dimer. Structure-guided mutagenesis of residues within the GDP-binding pocket identified Arg93 as playing a key role in the nucleotide interaction and revealed that GDP is required for activity. All mutations were confirmed to be specific and not to have resulted from structural perturbations by X-ray crystallography. This is the first report of a bacterial GDP-regulated thioesterase and of covalent linkage of thioesterase domains through a disulfide bond, revealing structural similarities with ADP regulation in the human ACOT12 thioesterase.

Structural and Functional Characterization of the PaaI Thioesterase from Streptococcus pneumoniae Reveals a Dual Specificity for Phenylacetyl-CoA and Medium-chain Fatty Acyl-CoAs and a Novel CoA-induced Fit Mechanism.

PaaI thioesterases are members of the TE13 thioesterase family that catalyze the hydrolysis of thioester bonds between coenzyme A and phenylacetyl-CoA. In this study we characterize the PaaI thioesterase from Streptococcus pneumoniae (SpPaaI), including structural analysis based on crystal diffraction data to 1.8-Å resolution, to reveal two double hotdog domains arranged in a back to back configuration. Consistent with the crystallography data, both size exclusion chromatography and small angle x-ray scattering data support a tetrameric arrangement of thioesterase domains in solution. Assessment of SpPaaI activity against a range of acyl-CoA substrates showed activity for both phenylacetyl-CoA and medium-chain fatty-acyl CoA substrates. Mutagenesis of putative active site residues reveals Asn(37), Asp(52), and Thr(68) are important for catalysis, and size exclusion chromatography analysis and x-ray crystallography confirm that these mutants retain the same tertiary and quaternary structures, establishing that the reduced activity is not a result of structural perturbations. Interestingly, the structure of SpPaaI in the presence of CoA provides a structural basis for the observed substrate specificity, accommodating a 10-carbon fatty acid chain, and a large conformational change of up to 38 Å in the N terminus, and a loop region involving Tyr(38)-Tyr(39). This is the first time PaaI thioesterases have displayed a dual specificity for medium-chain acyl-CoAs substrates and phenylacetyl-CoA substrates, and we provide a structural basis for this specificity, highlighting a novel induced fit mechanism that is likely to be conserved within members of this enzyme family.

Identification and synthesis of novel inhibitors of mycobacterium ATP synthase.

A non-diaryl quinoline scaffold 6,7-dihydropyrazolo[1,5-a]pyrazin-4-one was identified by screening of diverse set of compounds against M. smegmatis ATP synthase. Herein, we disclose our efforts to develop the structure activity relationship against Mycobacterium tuberculosis (Mtb.H37Rv strain) around the identified hit 1. A scaffold hopping approach was used to identify compounds 14a, 14b and 24a with improved activity against MTb.H37Rv.

Design and synthesis of novel, potent and selective hypoxanthine analogs as adenosine A1 receptor antagonists and their biological evaluation.

Multipronged approach was used to synthesize a library of diverse C-8 cyclopentyl hypoxanthine analogs from a common intermediate III. Several potent and selective compounds were identified and evaluated for pharmacokinetic (PK) properties in Wistar rats. One of the compounds 14 with acceptable PK parameters was selected for testing in in vivo primary acute diuresis model. The compound demonstrated significant diuretic activity in this model.

Autonomous Microsystems for Downhole Applications: Design Challenges, Current State, and Initial Test Results.

This paper describes two platforms for autonomous sensing microsystems that are intended for deployment in chemically corrosive environments at elevated temperatures and pressures. Following the deployment period, the microsystems are retrieved, recharged, and interrogated wirelessly at close proximity. The first platform is the Michigan Micro Mote for High Temperature (M³HT), a chip stack 2.9 × 1.1 × 1.5 mm³ in size. It uses RF communications to support pre-deployment and post-retrieval functions, and it uses customized electronics to achieve ultralow power consumption, permitting the use of a chip-scale battery. The second platform is the Environmental Logging Microsystem (ELM). This system, which is 6.5 × 6.3 × 4.5 mm³ in size, uses the smallest suitable off-the-shelf electronic and battery components that are compatible with assembly on a flexible printed circuit board. Data are stored in non-volatile memory, permitting retrieval even after total power loss. Pre-deployment and post-retrieval functions are supported by optical communication. Two types of encapsulation methods are used to withstand high pressure and corrosive environments: an epoxy filled volume is used for the M³HT, and a hollow stainless-steel shell with a sapphire lid is used for both the M³HT and ELM. The encapsulated systems were successfully tested at temperature and pressure reaching 150 °C and 10,000 psi, in environments of concentrated brine, oil, and cement slurry. At elevated temperatures, the limited lifetimes of available batteries constrain the active deployment period to several hours.

Neoplastic Fever: All who Shiver are not Infected.

We report the case of previously healthy 14 years old male who presented high grade fever and headache. There was a history of convulsion at age of 7 years, so MRI Brain was done. It was suggestive of a central nervous system neoplasm. Our patient had only two days of fever which is an unusual presentation of a neoplasm. The paper should be of interest to the clinicians as neoplastic fever as cause of acute febrile illness is considered as a remote possibility.

Oral bioavailability and pharmacodynamic activity of hesperetin nanocrystals generated using a novel bottom-up technology.

In the present study, nanocrystalline solid dispersion (NSD) was developed to enhance the release rate and oral bioavailability of hesperetin (HRN). NSD of HRN was prepared using a novel bottom-up technology platform. It is a spray drying based technology to generate solid particles, containing drug nanocrystals dispersed in small molecule excipients. HRN and mannitol were used in a 5:5 ratio, and an average crystallite size of HRN in NSD with mannitol was found to be 137.3 ± 90.0 nm. An in vitro release study revealed a statistically significant release rate enhancement for HRN nanocrystals (46.3 µg/mL/min) as compared to that of the control (29.5 µg/mL/min). Further, a comparative oral bioavailability study of NSD and control in Sprague-Dawley rats established significant improvement in Cmax and oral bioavailability (AUC0-∞) by 1.79- and 2.25-fold, respectively, for HRN nanocrystals. The findings of oral bioavailability were corroborated by intestinal uptake and Caco-2 cell uptake studies, wherein HRN, when administered in nanocrystalline form, showed higher penetration in intestinal mucosa and higher uptake in Caco-2 cells. Finally, the therapeutic efficacy of HRN nanocrystals was tested by a reactive oxygen species (ROS) generation assay and carrageenan induced anti-inflammatory model. HRN nanocrystals markedly inhibited ROS generation in MCF-7 cells, and carrageenan induced inflammation in rats. The process of NSD formation was found to be based on classical nucleation theory wherein mannitol contributed to NSD formation by acting as a plasticizer and crystallization inducer, and by providing sites for heterogeneous nucleation/crystallization.

Hysteretic DC electrowetting by field-induced nano-structurations on polystyrene films.

Electrowetting (EW) offers executive wetting control of conductive liquids on several polymer surfaces. We report a peculiar electrowetting response for aqueous drops on a polystyrene (PS) dielectric surface in the presence of silicone oil. After the first direct current (DC) voltage cycle, the droplet failed to regain Young's angle, yielding contact angle hysteresis, which is close to a value found in ambient air. We conjecture that the hysteretic EW response appears from in situ surface modification using electric field induced water-ion contact with PS surface inducing nano-structuration by electro-hydrodynamic (EHD) instability. Atomic force microscopy confirms the formation of nano-structuration on the electrowetted surface. The effects of molecular weight, applied electric field, water conductivity and pH on nano-structuration are studied. Finally, the EW based nano-structuration on PS surface is used for the enhanced loading of aqueous dyes on hydrophobic surfaces.