Carboxylic acid derivatives suppress the growth of Aspergillus flavus through the inhibition of fungal alpha-amylase.

Aspergillus favus (A. flavus) is a saprophytic fungus and a pathogen affecting several important foods and crops, including maize. A. flavus produces a toxic secondary metabolite called aflatoxin. Alpha-amylase (α-amylase), a hydrolytic enzyme produced by A. Flavus helps in the production of aflatoxin by hydrolysing the starch molecules in to simple sugars such as glucose and maltose. These simple sugars induce the production of aflatoxin. Inhibition of α-amylase has been proven as a potential way to reduce the production of aflatoxin. In the present study, we investigated the effect of selected carboxylic acid derivatives such as cinnamic acid (CA), 2, 4-dichlorophenoxyacetic acid (2,4-D), and 3-(4-hydroxyphenyl)-propionic acid (3,4-HPPA) on the fungal growth and for the α-amylase inhibitory activity. The binding potentials of these compounds with α-amylase have been confirmed by enzyme kinetics and isothermal titration calorimetry. Molecular docking and MD simulation studies were also performed to deduce the atomic level interaction between the protein and selected ligands. The results indicated that CA, 2,4-D and 3,4-HPPA can inhibit the fungal growth which could be partly due to the inhibition on fungal α-amylase activity.Communicated by Ramaswamy H. Sarma.

Unveiling the molecular basis of lobeline's allosteric regulation of NMDAR: insights from molecular modeling.

Neurological and psychiatric disorders contribute significantly to the global disease burden, adversely affecting the quality of life for both patients and their families. Impaired glutamatergic signaling is considered to be a major cause for most of the neurological and psychiatric disorders. Glutamate receptors are over activated in excitotoxic conditions, leading to dysregulation of Ca2+ homeostasis, triggering the production of free radicals and oxidative stress, mitochondrial dysfunction and eventually cell death. Excitotoxicity primarily results from the overactivity of NMDARs, a subtype of ionotropic glutamate receptors, due to their pronounced Ca2+ permeability and conductance characteristics. NMDAR antagonists are suggested to have therapeutic use as they can prevent excitotoxicity. Our previous studies demonstrated lobeline, an alkaloid, exerts neuroprotective action in excitotoxic conditions by blocking NMDAR. However, the atomic level interactions of lobeline with NMDAR was not characterized yet. Structural comparison of lobeline with a known NMDAR antagonist ifenprodil, followed by molecular docking and dynamics simulations revealed that lobeline could bind to the ifenprodil binding site i.e., in the heterodimer interface of GluN1-GluN2B subunits and exert ifenprodil like activities. By in silico structure guided modifications on lobeline and subsequent free energy calculations, we propose putative NMDAR antagonists derived from lobeline.

A Comparative Study of Solifenacin, Mirabegron, and Their Combination as Bladder Relaxants in the Management of Overactive Bladder.

Introduction  Overactive bladder (OAB) is a medical state that presents as the urgency of urine and increased frequency of micturition and is diagnosed on the basis of the presence of these symptoms in the absence of other explainable diagnoses. The management of this condition includes conservative management, medical management/pharmacotherapy, and surgical management. The overactive bladder has been treated with smooth muscle relaxants, but there are conflicting results. Hence, this study aimed to assess the result of the two smooth muscle relaxants, mirabegron and solifenacin, and their combination to manage an overactive bladder. Methodology  A clinical trial was conducted at Swaroop Rani Nehru Hospital, Motilal Nehru Medical College, Prayagraj, India, over the period from November 2019 to December 2020. Ninety patients with OAB were divided into three groups: G1, G2, and G3. These groups were administered solifenacin, mirabegron, and a combination of mirabegron and solifenacin (S+M), respectively. Follow-ups were conducted at 2, 4, 12, and 18 weeks for evaluation. Data were entered into IBM SPSS Statistics for Windows, Version 23 (Released 2015; IBM Corp., Armonk, New York, United States). Appropriate statistical tests, including the chi-square and ANOVA, were employed in this study. Observation  The combination of mirabegron and solifenacin was significantly more effective in terms of response compared to solifenacin alone. There was no significant difference between solifenacin versus mirabegron, or between mirabegron (M) and the combination of mirabegron (M) and solifenacin (S). Side effects were more severe in patients taking high doses of solifenacin. Conclusion  The S + M combination has higher efficacy than solifenacin and mirabegron when given alone.

Sub-pocket-focused designing of tacrine derivatives as potential acetylcholinesterase inhibitors.

Human acetylcholinesterase (hAChE) has a potential role in the management of acetylcholine, one of the neurotransmitters that modulate the overall activity of cholinergic system, AChE inhibitors have a greater impact in the therapeutics. Though the atomic structure of hAChE has been extensively studied, the precise active site geometry upon binding to different ligands are yet to be explored. In the present study, an extensive structural analysis of our recently reported hAChE-tacrine complex has carried out and revealed the presence of two prominent sub-pockets located at the vicinity of the hAChE active site. Structural bioinformatics assisted studies designed 132 putative sub-pockets focused tacrine derivatives (SPFTDs), their molecular docking, free energy estimations revealed that they are stronger than tacrine in terms of binding affinity. Our in vitro studies also supported the in silico findings, all these SPFTDs are having better potencies than tacrine. Cytotoxic nature of these SPFTDs on HepG2 and Neuro-2a cell lines, diminishes the possibilities for future in vivo studies. However, the identification of these sub pockets and the SPFTDs paved a new way to the future drug discovery especially since AChE is one of the promising and approved drug targets in treatment of AD drug discovery.

Structural analysis and ensemble docking revealed the binding modes of selected progesterone receptor modulators.

Uterine fibroids (UF) are benign smooth muscle neoplasm of uterus that have a significant impact on a woman's quality of life as they perturb hormonal homeostasis resulting in heavy menstrual bleeding, impaired fertility, pregnancy complications and loss. UF can be surgically removed through invasive procedures, but their recurrence rate is often high. Progesterone receptor (PR) has an imperative role in UF management. Mifepristone, ulipristal acetate (UPA) and asoprisnil (ASO) are some selective progesterone receptor modulators (SPRMs), acts on PR, but due to their side effects in long term use, they were withdrawn from the market. Hence, there is a dire need for novel, highly efficient with least side effects, therapeutics for the treatment of UF. To contribute toward the drug discovery for UF, we made an extensive structural comparison of reported PR crystal structures, also elucidated the binding modes of four existing SPRMs against human PR through ensemble docking approach. Our studies revealed the presence of 5 highly repeating water molecules that has an important role in ligand binding and structural stability. Our ensemble docking and MD simulation revealed that studied ligands have preferential selectivity toward the specific conformation of PR. It is anticipated that our study will be a useful resource to all the drug discovery scientists who are engaged in the identification of lead molecules against UF.

Growth Velocity in South Indian Children Between Three and 18 Years of Age.

Objective Height velocity is a crucial anthropometric parameter for the evaluation of mild- or recent-onset short stature; however, there is no data on height velocity in South Indian children. We undertook this study to establish the normative data. Methods This prospective longitudinal study included 3327 apparently healthy children aged three to 18 years from government and private schools of Krishna district, Andhra Pradesh. Height and weight were measured at baseline and three-monthly intervals for one year (October 2018 to October 2019). Results Age- and sex-specific height velocity percentiles were generated. The data was available in 1627 boys and 1700 girls. The mean peak height velocity (PHV) was 7.18±2.56 cm in boys observed at 12-12.9 years and 5.8±2.56 cm in girls at 10-10.9 years. Conclusion Normative height velocity data for South Indian children has been presented.

AMX - the highly automated macromolecular crystallography (17-ID-1) beamline at the NSLS-II.

The highly automated macromolecular crystallography beamline AMX/17-ID-1 is an undulator-based high-intensity (>5 × 1012 photons s-1), micro-focus (7 µm × 5 µm), low-divergence (1 mrad × 0.35 mrad) energy-tunable (5-18 keV) beamline at the NSLS-II, Brookhaven National Laboratory, Upton, NY, USA. It is one of the three life science beamlines constructed by the NIH under the ABBIX project and it shares sector 17-ID with the FMX beamline, the frontier micro-focus macromolecular crystallography beamline. AMX saw first light in March 2016 and started general user operation in February 2017. At AMX, emphasis has been placed on high throughput, high capacity, and automation to enable data collection from the most challenging projects using an intense micro-focus beam. Here, the current state and capabilities of the beamline are reported, and the different macromolecular crystallography experiments that are routinely performed at AMX/17-ID-1 as well as some plans for the near future are presented.

Binding of rosmarinic acid curcumin and capsaicin with PLA2: A comparative study.

Phospholipase A2 (PLA2) is a key enzyme involved in the formation of pro-inflammatory mediators like eicosanoids. Inhibition of PLA2 is regarded as one of the effective methods of controlling inflammation. The present study investigated the binding potentials of three natural compounds, rosmarinic acid (RA), capsaicin (CAP), and curcumin (CUR) by means of in silico and in vitro methods. Our study revealed that RA has relatively better binding affinity and inhibition potentials when compared to the other two molecules. Our ITC experiments were also suggested a slightly better binding energy for the RA. The stoichiometry of the protein ligand complex obtained from one of the ITC experiments suggested the possibilities of binding of a small molecule MCW (degraded product of CUR) on PLA2. Overall study demonstrated that the anti-inflammatory activity of RA, CUR and CAP may be partly due to the inhibition of PLA2.

Crystal structure of human acetylcholinesterase in complex with tacrine: Implications for drug discovery.

Alzheimer's disease (AD) is one of the most common, progressive neurodegenerative disorders affecting the aged populations. Though various disease pathologies have been suggested for AD, the impairment of the cholinergic system is one of the critical factors for the disease progression. Restoration of the cholinergic transmission through acetylcholinesterase (AChE) inhibitors is a promising disease modifying therapy. Being the first marketed drug for AD, tacrine reversibly inhibits AChE and thereby slows the breakdown of the chemical messenger acetylcholine (ACh) in the brain. However, the atomic level of interactions of tacrine towards human AChE (hAChE) is unknown for years. Hence, in the current study, we report the X-ray structure of hAChE-tacrine complex at 2.85 Å resolution. The conformational heterogeneity of tacrine within the electron density was addressed with the help of molecular mechanics assisted methods and the low-energy ligand configuration is reported, which provides a mechanistic explanation for the high binding affinity of tacrine towards AChE. Additionally, structural comparison of reported hAChE structures sheds light on the conformational selection and induced fit effects of various active site residues upon binding to different ligands and provides insight for future drug design campaigns against AD where AChE is a drug target.

Tumor Derived Extracellular Vesicles Drive T Cell Exhaustion in Tumor Microenvironment through Sphingosine Mediated Signaling and Impacting Immunotherapy Outcomes in Ovarian Cancer.

SPHK1 (sphingosine kinase-1) catalyzes the phosphorylation of sphingosine to sphingosine-1-phosphate (S1P), is found to be highly expressed in solid tumors. Here, extracellular vesicles (EVs) are identified as the key transporters of SPHK1 to the tumor microenvironment. Consequently, SPHK1-packaged EVs elevate S1P levels in the tumor microenvironment, where S1P appears as an immunosuppressive agent. However, the exact mechanism of how S1P mediates its immunosuppressive effects in cancer is not understood. It is investigated that S1P can induce T cell exhaustion. S1P can also upregulate programmed death ligand-1 (PDL-1) expression through E2F1-mediated transcription. Notably, an SPHK1 inhibitor PF543 improves T cell-mediated cytotoxicity. Furthermore, combining PF543 with an anti-PD-1 antibody reduces tumor burden and metastasis more effectively than PF543 alone in vivo. These data demonstrate a previously unrecognized mechanism of how SPHK1-packaged EVs contribute to the progression of ovarian cancer and thus present the potential clinical application of inhibiting SPHK1/S1P signaling to improve immune checkpoint blockage (anti-PD-1 antibody) therapy in ovarian cancer.

Neuroprotective derivatives of tacrine that target NMDA receptor and acetyl cholinesterase - Design, synthesis and biological evaluation.

The complex and multifactorial nature of neuropsychiatric diseases demands multi-target drugs that can intervene with various sub-pathologies underlying disease progression. Targeting the impairments in cholinergic and glutamatergic neurotransmissions with small molecules has been suggested as one of the potential disease-modifying approaches for Alzheimer's disease (AD). Tacrine, a potent inhibitor of acetylcholinesterase (AChE) is the first FDA approved drug for the treatment of AD. Tacrine is also a low affinity antagonist of N-methyl-D-aspartate receptor (NMDAR). However, tacrine was withdrawn from its clinical use later due to its hepatotoxicity. With an aim to develop novel high affinity multi-target directed ligands (MTDLs) against AChE and NMDAR, with reduced hepatotoxicity, we performed in silico structure-based modifications on tacrine, chemical synthesis of the derivatives and in vitro validation of their activities. Nineteen such derivatives showed inhibition with IC50 values in the range of 18.53 ± 2.09 - 184.09 ± 19.23 nM against AChE and 0.27 ± 0.05 - 38.84 ± 9.64 µM against NMDAR. Some of the selected compounds also protected rat primary cortical neurons from glutamate induced excitotoxicity. Two of the tacrine derived MTDLs, 201 and 208 exhibited in vivo efficacy in rats by protecting against behavioral impairment induced by administration of the excitotoxic agent, monosodium glutamate. Additionally, several of these synthesized compounds also exhibited promising inhibitory activitiy against butyrylcholinesterase. MTDL-201 was also devoid of hepatotoxicity in vivo. Given the therapeutic potential of MTDLs in disease-modifying therapy, our studies revealed several promising MTDLs among which 201 appears to be a potential candidate for immediate preclinical evaluations.

Scanning electron microscopy of Cysticercus fasciolaris (larval Taenia taeniaeformis Batsch, 1786) from the wild rat, Rattus rattus Linnaeus, 1758, morpho-physiology and risk to human health.

Autopsy of Rattus rattus (n=65) revealed 1-7 creamish to white cysts, 2-7 mm in diameter embedded in the liver parenchyma of 12 rats. Dissection yielded 45-65 mm×5 mm sized segmented strobilocercii of Cysticercus fasciolaris (larval Taenia taeniaeformis) coiled inside. Light microscopy indicated large scolices, the rostellum armed with four suckers and one row of taenoid type hooks having long blunt handle with sharp pointed blade, other row of hooks was in the developing stage. The strobila lacked genital organs and ended in a tail-bulb suggesting it was juvenile. SEM showed anterior crown of 19 hooks and poorly developed posterior crown. A collar (66.0-86.0 µm in width) armed with papillae (30.2-35.4 µm base and 12.5-14.5 µm tip) and pores (17.0-22.4 µm in diameter) surrounded the hooks. The body segments were 125-145 µm in diameter, at the tail bud, 380-410 µm in diameter. The segments were armed with pores, 11.5-14.5 µm in diameter. Netted (2.5-4.5 µm wide and of varying lengths) body segments provide flexibility to the worm during attachment. The tail bud was 2.4-2.7 mm in length and 1.5-1.71 mm in width. The SEM data presents a significant advancement over light microscopy and the morphological features generated herein can safely be utilized to correlate with the parasite's physiological functions. This is the first report of R. rattus as a natural intermediate host of T. taeniaeformis, and may pose serious risk to human health in urban areas of Bareilly, India and merits attention.

FMX - the Frontier Microfocusing Macromolecular Crystallography Beamline at the National Synchrotron Light Source II.

Two new macromolecular crystallography (MX) beamlines at the National Synchrotron Light Source II, FMX and AMX, opened for general user operation in February 2017 [Schneider et al. (2013). J. Phys. Conf. Ser. 425, 012003; Fuchs et al. (2014). J. Phys. Conf. Ser. 493, 012021; Fuchs et al. (2016). AIP Conf. Proc. SRI2015, 1741, 030006]. FMX, the micro-focusing Frontier MX beamline in sector 17-ID-2 at NSLS-II, covers a 5-30 keV photon energy range and delivers a flux of 4.0 × 1012 photons s-1 at 1 Šinto a 1 µm × 1.5 µm to 10 µm × 10 µm (V × H) variable focus, expected to reach 5 × 1012 photons s-1 at final storage-ring current. This flux density surpasses most MX beamlines by nearly two orders of magnitude. The high brightness and microbeam capability of FMX are focused on solving difficult crystallographic challenges. The beamline's flexible design supports a wide range of structure determination methods - serial crystallography on micrometre-sized crystals, raster optimization of diffraction from inhomogeneous crystals, high-resolution data collection from large-unit-cell crystals, room-temperature data collection for crystals that are difficult to freeze and for studying conformational dynamics, and fully automated data collection for sample-screening and ligand-binding studies. FMX's high dose rate reduces data collection times for applications like serial crystallography to minutes rather than hours. With associated sample lifetimes as short as a few milliseconds, new rapid sample-delivery methods have been implemented, such as an ultra-high-speed high-precision piezo scanner goniometer [Gao et al. (2018). J. Synchrotron Rad. 25, 1362-1370], new microcrystal-optimized micromesh well sample holders [Guo et al. (2018). IUCrJ, 5, 238-246] and highly viscous media injectors [Weierstall et al. (2014). Nat. Commun. 5, 3309]. The new beamline pushes the frontier of synchrotron crystallography and enables users to determine structures from difficult-to-crystallize targets like membrane proteins, using previously intractable crystals of a few micrometres in size, and to obtain quality structures from irregular larger crystals.

Microbial, environmental and anthropogenic factors influencing the indoor microbiome of the built environment.

A built environment is a human-made environment providing surroundings for human occupancy, activities, and settlement. It is supposed to safeguard humans from all undesirable and harmful pollutants; however, indoor concentrations of some pollutants are much greater than that of the outdoors. Bioaerosols infiltrate from the outdoors in addition to many indoor sources of bioaerosols including the use of various chemicals as well as activities like cooking, smoking, cleaning, or even normal movement. They are also associated with a number of serious health concerns. Various ecological factors associated with the generation, the persistence as well as the dispersal of these microbial components of indoor bioaerosols, are discussed in this review, that have not been considered all together till now. The factors like microbial taxa, environmental factors, and anthropogenic activities (human occupancy, activities, and impact of urbanization) are addressed in the review. Effects of both indoor environmental factors like architectural design, lighting, ventilation, temperature, humidity, indoor/outdoor ratio, particulate matter, indoor chemistry as well as outdoor environmental factors like geography, seasons, and meteorology on the microbial concentrations have been discussed. Efforts are underway to design selective pressures for microbes to create a healthy symbiotic built microbiome as the "right" indoor microbiome is a "healthy" indoor microbiome.

Chemical similarity assisted search for acetylcholinesterase inhibitors: Molecular modeling and evaluation of their neuroprotective properties.

Alzheimer's disease (AD) is an obstinate and progressive neurodegenerative disorder, mainly characterized by cognitive decline. Increasing number of AD patients and the lack of promising treatment strategies demands novel therapeutic agents to combat various disease pathologies in AD. Recent progresses in understanding molecular mechanisms in AD helped researchers to streamline the various therapeutic approaches. Inhibiting acetylcholinesterase (AChE) activity has emerged as one of the potential treatment strategies. The present study discusses the identification of two potent AChE inhibitors (ZINC11709541 and ZINC11996936) from ZINC database through conventional in silico approaches and their in vitro validations. These inhibitors have strong preferences towards AChE than butyrylcholinesterase (BChE) and didn't evoke any significant reduction in the cell viability of HEK-293 cells and primary cortical neurons. Furthermore, promising neuroprotective properties has also been displayed against glutamate induced excitotoxicity in primary cortical neurons. The present study proposes two potential drug lead compounds for the treatment of AD, that can be used for further studies and preclinical evaluation.

Piperidine-4-carboxamide as a new scaffold for designing secretory glutaminyl cyclase inhibitors.

Alzheimer's disease (AD), a common chronic neurodegenerative disease, has become a major public health concern. Despite years of research, therapeutics for AD are limited. Overexpression of secretory glutaminyl cyclase (sQC) in AD brain leads to the formation of a highly neurotoxic pyroglutamate variant of amyloid beta, pGlu-Aß, which acts as a potential seed for the aggregation of full length Aß. Preventing the formation of pGlu-Aß through inhibition of sQC has become an attractive disease-modifying therapy in AD. In this current study, through a pharmacophore assisted high throughput virtual screening, we report a novel sQC inhibitor (Cpd-41) with a piperidine-4-carboxamide moiety (IC50 = 34 µM). Systematic molecular docking, MD simulations and X-ray crystallographic analysis provided atomistic details of the binding of Cpd-41 in the active site of sQC. The unique mode of binding and moderate toxicity of Cpd-41 make this molecule an attractive candidate for designing high affinity sQC inhibitors.

ATP7A Clinical Genetics Resource - A comprehensive clinically annotated database and resource for genetic variants in ATP7A gene.

ATP7A is a critical copper transporter involved in Menkes Disease, Occipital horn Syndrome and X-linked distal spinal muscular atrophy type 3 which are X linked genetic disorders. These are rare diseases and their genetic epidemiology of the diseases is unknown. A number of genetic variants in the genes have been reported in published literature as well as databases, however, understanding the pathogenicity of variants and genetic epidemiology requires the data to be compiled in a unified format. To this end, we systematically compiled genetic variants from published literature and datasets. Each of the variants were systematically evaluated for evidences with respect to their pathogenicity and classified as per the American College of Medical Genetics and the Association of Molecular Pathologists (ACMG-AMP) guidelines into Pathogenic, Likely Pathogenic, Benign, Likely Benign and Variants of Uncertain Significance. Additional integrative analysis of population genomic datasets provides insights into the genetic epidemiology of the disease through estimation of carrier frequencies in global populations. To deliver a mechanistic explanation for the pathogenicity of selected variants, we also performed molecular modeling studies. Our modeling studies concluded that the small structural distortions observed in the local structures of the protein may lead to the destabilization of the global structure. To the best of our knowledge, ATP7A Clinical Genetics Resource is one of the most comprehensive compendium of variants in the gene providing clinically relevant annotations in gene.

Rapid detection and quantification of sucrose adulteration in cow milk using Attenuated total reflectance-Fourier transform infrared spectroscopy coupled with multivariate analysis.

Adulteration of milk to gain economic benefit has become a common practice in recent years. Sucrose is illegally added in milk to reconstitute its compositional requirement by improving the total solid contents. The present study is aimed to use FTIR spectroscopy in combination with multivariate chemometric modelling for the differentiation and quantification of sucrose in cow milk. Pure milk and adulterated milk spectra (0.5-7.5% w/v) were observed in the spectral region 4000-400 cm-1. Principal component analysis (PCA) was used for the discrimination of pure milk and adulterated milk. Soft independent modelling of class analogy (SIMCA) was able to classify test samples with a classification efficiency of 100%. Partial least square regression (PLS-R) and principle component regression (PCR) models were established for normal spectra, 1st derivative and 2nd derivative for the quantification of sucrose in milk. PLS-R model (normal spectra) in the combined wavenumber range of 1070-980 cm-1 showed the best prediction based on parameters like coefficient of determination (R2) (Cal: 0.996; Val: 0.993), RMSE (Cal: 0.15% w/v; Val: 0.20% w/v), RE% (Cal: 4.9% w/v; Val: 5.1% w/v) and RPD (13.40). This method has a detection level of 0.5% w/v sucrose adulteration.

Development of an FTIR based chemometric model for the qualitative and quantitative evaluation of cane sugar as an added sugar adulterant in apple fruit juices.

A Fourier Transform Infrared Spectroscopy based chemometric model was evaluated for the rapid identification and estimation of cane sugar as an added sugar adulterant in apple fruit juices. For all the ninety samples, spectra were acquired in the mid-infrared range (4000 cm-1-400 cm-1). The spectral analysis provided information regarding the distinctive variable region, which lies in the range of 1200cm-1 to 900cm-1, designated as fingerprint region for the carbohydrates. A specific peak in the fingerprint region was observed at 997cm-1 in all the adulterated samples and was undetectable in pure samples. Based on different levels of cane sugar adulteration (5, 10, 15, and 20%), principal component analysis showed the clustering of samples and further helped us in compression of data by selecting wavenumbers with maximum variability based on the loading line plot. Supervised classification methods (SIMCA and LDA) were evaluated based on their classification efficiencies for a test set. Though SIMCA showed 100% classification efficiency (Raw data set), LDA was able to classify the test set with an accuracy of only 96.67% (Raw as well as Transformed data set) between pure and 5% adulterated samples. For the quantitative estimation, calibration models were developed using partial least square regression (PLS-R) and principal component regression method (PCR) methods. PLS-1st derivative showed a maximum coefficient of determination (R2) with a value of 0.991 for calibration and 0.992 for prediction. The RMSECV, RMSEP, LOD and LOQ observed for PLS-1st derivative model were 0.75% w/v, 0.61% w/v, 1.28%w/v and 3.88%w/v, respectively. The coefficient of variation as a measure of precision (repeatability) was also determined for all models, and it ranged from 0.23% to 1.83% (interday), and 0.25% to 1.43% (intraday).