Method development and validation for the estimation of gedatolisib in mouse plasma by tandem mass spectrometry and its application to pharmacokinetics studies.

A simple and a sensitive liquid chromatography-tandem mass spectrometry method was developed and validated for the quantification of gedatolisib in mouse plasma. The extraction technique involved a simple precipitation method to extract gedatolisib and idelalisib (internal standard) from mouse plasma. A clean chromatographic separation of gedatolisib and the internal standard was achieved on an Atlantis dC18 column using an isocratic mobile phase (10 mm ammonium formate and acetonitrile; 30:70% v/v, both supplemented with 0.1% formic acid) delivered at a flow rate of 0.7 ml/min. The total run time was 2.0 min, and gedatolisib and idelalisib were eluted at 0.80 and 0.95 min, respectively. Gedatolisib was monitored at m/z 616.40 → 488.20 and idelalisib at 416.05 → 176.10. All the required parameters for the method validation were performed as per US Food and Drug Administration guidelines, and the results were within the acceptance criteria. The method was accurate and proved to be precise at a linearity range of 1.33-2667 ng/ml. The accuracy for gedatolisib in mouse plasma was in the ranges 0.99-1.06% (intra-day) and 0.96-1.04% (inter-day). Gedatolisib appeared to be stable in a series of stability conditions. Gedatolisib showed a good intravenous profile when administered through a solution formulation.

Validated liquid chromatography with tandem mass spectrometry method for determination of paxalisib in mouse plasma: An application to pharmacokinetic study in mice.

A sensitive and rapid LC-MS/MS method has been developed and validated for the quantification of paxalisib in mouse plasma. A liquid-liquid extraction method was used for the extraction of paxalisib and filgotinib (internal standard, IS) from mouse plasma. A clean chromatographic separation of paxalisib and the IS was achieved on an Atlantis dC18 column using an isocratic mobile phase (10 mM ammonium formate and acetonitrile, 30:70%, v/v) delivered at a flow rate of 0.7 mL/min. The total run time was 2.5 min. Paxalisib and filgotinib were eluted at 1.21 and 0.94 min, respectively. The MS/MS transitions monitored were m/z 383.25 → 309.20 and 426.30 → 291.20 for paxalisib and filgotinib, respectively. Method validation was performed as per US Food and Drug Administration guidelines and the results met the acceptance criteria. The method was proved to be accurate and precise at a linearity range of 1.39-2287 ng/mL. The intra- and inter-day precisions for paxalisib in mouse plasma were in the ranges 1.42-9.61 and 4.70-9.63%, respectively. Paxalisib was stable under a series of stability conditions. Post-oral administration to mice, paxalisib maximum plasma concentrations were attained at 2.0 h. Paxalisib's half-life ranged between 3.2 and 4.2 h. Paxalisib exhibited low clearance and moderate volume of distribution. Oral bioavailability was 71%.

Application of waste eggshell as a source of calcium in bacterial bio-cementation to enhance the engineering characteristics of sand.

A technique to produce bio-cementation in sandy soil using the microbially induced calcium carbonate precipitation (MICP) process and calcium ions generated from eggshell is presented in this research. This research also focused on the application of S. pasteurii bacteria and L. fusiformis bacteria along with eggshell and calcium chloride cementing chemicals on the strength properties of sand. The experimental variables maintained in this research are bacteria type (S. pasteurii and L. fusiformis), cementing chemical type (eggshell and calcium chloride) and molarity of the cementing chemical (0.25, 0.50, 0.75 and 1.0). The engineering behaviour of bacteria treated sand was estimated by executing the unconfined compression test and permeability test in the laboratory. From the experimental findings, it is identified that the unconfined compressive strength of sand is enhanced and the value is in the range of 650 kPa. In addition to that, the permeability of sand is minimized in the order of two from 6.3 × E-3 to 3.2 × E-5 cm/s. The best improvement of Young's modulus and calcium carbonate content estimated in this research are 28.9 MPa and 17.9% when the sand is treated with S. pasteurii along 0.50 molarity of eggshell cementing chemical. The experimental findings are validated with the help of microstructural studies of scanning electron microscopy (SEM) and energy-dispersive X-ray analysis (EDX). This research showed that bio-cementation technology in the form of S. pasteurii and eggshell can be effectively adopted to enhance the engineering characteristics of sand.

Designing Z-scheme AgIO4 nanorod embedded with Bi2S3 nanoflakes for expeditious visible light photodegradation of congo red and rhodamine B.

The present study describes the enhanced photodegradation of organic pollutant dyes, congo red (CR) and rhodamine B (RhB) dyes under visible light irradiation. AgIO4 nanorods decorated on Bi2S3 nanoflakes in various proportions were synthesized via sono-chemical route wherein the deposition of varying amounts of AgIO4 on Bi2S3 plays a pivotal role in improving the photodegradation ability. The characterization of the as-synthesized nanohybrids was assessed by XRD, UV-vis DRS, PL, EIS, ESR, FT-IR, XPS, HR-TEM, FE-SEM, N2 adsorption and desorption techniques. The effect of initial CR and RhB dye concentration, reaction pH and usage of nanohybrid concentration were investigated where 30%-AgIO4/Bi2S3 exhibited excellent visible light photodegradation of 95.58% for CR and 96.11% for RhB dyes at 140 min and 100 min respectively. The superoxide (•O2-) and hydroxyl radicals (•OH) played predominant role in the photodegradation of CR and RhB which is experimentally confirmed by radical trapping experiments. Also, the photocatalysts exhibited good photo stability and excellent reusability. The TOC analysis confirmed the complete mineralization of CR and RhB dyes by the nanohybrid and the formation of possible intermediate and degradation pathway was delineated based on GC/MS analysis. The outstanding photodegradation performance were ascribed to the Z-scheme charge transfer path, which effectively promotes the separation and transfer of e-/h+ pairs, resulting in a strong redox activity of the accumulated charge to decompose organic dyes during the degradation reaction. The study suggested that the nanohybrid can be utilized for the removal of organic pollutants from the contaminated water bodies.

A novel SPR based Fe@Ag core-shell nanosphere entrapped on starch matrix an optical probe for sensing of mercury(II) ion: A nanomolar detection, wide pH range and real water sample application.

Recent trends in nanotechnology paved a way for the development of detection systems for heavy metals, toxins and environmental pollutants. The present study focused on Hg2+ detection by a core shell Fe@Ag-starch nanosphere phenylalanine conjugate. The characterization of core shell Fe@Ag-starch nanosphere was performed by using TEM, zetasizer, particlesize analyzer, UV-visible absorption spectrophotometer, EDAX, FTIR and TGA. The NPs showed λmax at 408 nm. The effective diameter of synthesized nanosphere was 37 ± 2 nm and it possessed the surfaces charge of -36.12 ± 2.5 mV. The Fe@Ag-starch-phenylalanine conjugate reacted with Hg2+, the yellow colour of the nanosphere phenylalanine conjugate became colourless. The real water sample was collected and the amount of Hg2+ was calculated by using the prepared nanosphere. The detection of Hg2+ at different conditions including various saline concentrations, temperature and pH were also studied and the detection was found to be effective at 40 °C, pH 5 and 0.1% of saline concentration. The LOD of Hg2+ ions by Fe@Ag-starch nanosphere were calculated to be 1.84 nM. The influence of other metal ions present in the analyte did not show any interference on Hg2+ detection. In addition, the photocatalytic and antibacterial activities of Fe@Ag-starch nanosphere were also studied. The study confirmed that the core shell nanosphere can also be used for environmental cleanup and disinfection.

Cytotoxicological evaluation of copper oxide nanoparticles on green algae, bacteria and crustacean systems.

PURPOSE: Copper oxide (CuO) nanoparticles (NPs) have been utilized in several industries including textile, consumer products, medical, automobiles etc. The discharge of industrial effluents in environment increased the probability of CuO NPs contamination in the ecosystem. METHODS: The present investigation used CuO NPs to determine the toxic effect on Lyngbya species, fresh water algae isolated from natural pond, bacterial species Pseudomonas aeruginosa and Staphylococcus aureus and a crustacean species Daphnia magna. RESULTS: The NPs average diameter and zeta potential was estimated to be 45 ± 3 nm and 29 ± 1.78 mV respectively. The results showed that 0.1 µg/mL CuO NPs showed the growth inhibition of 47 ± 2% on Lyngbya sp. after 5 days of incubation. The CuO NPs also showed toxic effect to bacterial systems such as P. aeruginosa and S. aureus and crustacean system D. magna. Further, there was an increased lipid peroxidation and generation of reactive oxygen species (ROS) in algal cells observed up on NPs exposure. The exposure of NPs suppressed the antioxidant defense system. The amount of glutathione was reduced after the exposure of NPs. CONCLUSION: The study suggested the role of ROS in toxicity of algal and bacterial systems. The present study pointed out the potent toxicity of CuO NPs to the organisms present in the aquatic environment.

A potent multifunctional Ag/Co-polyvinylpyrrolidone nanocomposite for enhanced detection of Cr(III) from environmental samples and its photocatalytic and antibacterial applications.

Trivalent chromium (Cr(III)) is considered to exhibit hormesis (bi-phasic dose-response) property, where low dose be beneficial and high dose shows toxic effect. The present work describe the development of a bimetallic Ag/Co-polyvinylpyrrolidone nanocomposite (Ag/Co-PVP NPs) probe to detect and quantify Cr(III) ions from aqueous samples. The hydrodynamic size and zeta potential of the particle was determined to be 29 ± 1.3 nm and -37.19 ± 2.4 mV respectively. The interaction of Cr(III) with Ag/Co-PVP probe showed drastic change in colour of NPs from dark brown to pale yellow, with corresponding blue shift, tapering width and increased peak intensity. The probe showed high specificity towards Cr(III) among the tested metal ions. A linearity was observed between various dilutions of Cr(III) ions (10 to 50 nM) and the absorbance of Ag/Co-PVP NPs at 428 nm with R2 value of 0.998. The minimum detectable limit of Cr(III) was calculated to be 0.6 nM. The influence of salinity, temperature and pH on detection was studied. The probe was found to detect Cr(III) at acidic pH effectively. Competitive metal ions did not interfere the detection of Cr(III). The water sample collected from Noyyal river was taken to estimate Cr(III) by using the prepared probe to ensure practical applicability. The sample contains 9.3 nM of Cr(III) that was cross verified with AAS analysis. Hence, it is understood that the reported probe can be used to detect Cr(III) selectively with high accuracy from aqueous samples. In addition, the particles also exhibited excellent photocatalytic activity under visible light. Ag/Co-PVP nanocomposites exhibited excellent antibacterial activity against both gram +ve (B. subtilis) and gram -ve (E. coli) bacteria.

Preparation of Ag-cellulose nanocomposite for the selective detection and quantification of mercury at nanomolar level and the evaluation of its photocatalytic performance.

Mercury is a toxic heavy metal that reaches to the water bodies mainly by coal burning, mining and petrol refining. The study was focused to investigate the application of Ag-cellulose nanocomposite to detect and quantify mercury colorimetrically. The Ag-cellulose nanocomposite was characterized by X-ray diffraction, Transmission electron microscopy, Fourier transform infrared spectroscopy, UV-visible spectroscopy, particle size analyzer and zetasizer. The study identified that the presence of other metal ions did not interfere with the detection of Hg2+ ion by the probe. The prepared Ag-cellulose nanocomposite-phenylalanine conjugate incorporated paper strip showed an excellent result in Hg2+ detection. The Ag-cellulose nanocomposite was used to quantify the unknown concentration of mercury on real sample (environmental sample) and it was found to be highly accurate by confirming with atomic absorption spectrophotometric analysis. The Ag-cellulose nanocomposite showed effective detection at 45 °C, pH 9 and 0.1% of salinity. The Ag-cellulose nanocomposite showed efficient photocatalytic performance under visible light irradiation. The half-life period of MB by Ag-cellulose nanocomposite under visible light was determined to be 90 min. The study suggests the application of prepared probe in photocatalysis and the detection of Hg2+ from various environmental samples.

Colorimetric detection of mercury ions from environmental water sample by using 3-(Trimethoxysilyl)propyl methacrylate functionalized Ag NPs-tryptophan nanoconjugate.

Recent trend in the nanotechnology made an interest to make nano based system to detect the environmental pollutant including heavy metals, pesticides and antibiotics. Mercury is toxic heavy metals which causes hazardous effect to human and environmental organisms. They usually reach to the environment by mining, petrol refining and coal burning, which can change to its ionic forms according to the environmental condition. The present study was concentrated on the effective detection of Hg (II) ion from environmental sample colorimetrically by developing Ag NPs - tryptophan nanoconjugate functionalized with 3-(Trimethoxysilyl)propyl methacrylate (TMPM). The characterization of prepared particles was performed by UV-visible spectrophotometer, transmission electron microscopy (HRTEM), Fourier-transform infrared spectroscopy (FTIR), X-ray diffractometer (XRD), particle size analyzer and zeta sizer. The size of prepared NPs was 9 ± 1.10 nm and it possess the surface charge of -37.41 ± 4 mV. Upon the interaction of Ag NPs - tryptophan nanoconjugate and Hg (II) ion, the colour of the conjugate disappeared. The effect of environmental factors (Temperature, pH and saline concentration) on Hg (II) ion detection was also investigated. The probe indicated that Ag NPs - tryptophan nanoconjugate functionalized with TMPM was found to be an efficient tool for mercury detection from various environmental water samples.

Anthraquinone from Edible Fungi Pleurotus ostreatus Protects Human SH-SY5Y Neuroblastoma Cells Against 6-Hydroxydopamine-Induced Cell Death-Preclinical Validation of Gene Knockout Possibilities of PARK7, PINK1, and SNCA1 Using CRISPR SpCas9.

Parkinson's disease (PD) results from the degeneration of the nervous tissue brought about by ecological and hereditary components which affects nerve cells in the brain. It is the world's second most normal neurodegenerative issue, which can essentially weaken the personal satisfaction, make reliance, and trigger untimely mortality of affected people. The commonness pace of PD is 0.5-1% among individuals in the age group of 65-69 years and 1-3% among those 80 or more. Clinical appearances incorporate bradykinesia, tremors, unbending nature, and postural unsteadiness; spectrums of non-motor symptoms include psychological hindrance and passionate and behavioral brokenness. In this study, 6-OHDA-induced neurotoxicity was analyzed for various cytotoxicity analyses. The genes identified were PINK1 (PTEN-induced kinase 1), PARK7 (Parkinsonism-associated deglycase) and SNCA 1 (alpha synuclein1) validated using CRISPR spcas9 genome editing tool. In this study, Anthraquinone isolated from Pleurotus ostreatus was treated against a dopaminergic neurotoxin, 6-hydroxydopamine (6-OHDA), which induced neurotoxicity in SH-SY5Y cells. Experimental groups in SH-SY5Y neuroblastoma cells were treated with anthraquinone (50 nM) and 6-OHDA (100 nM). MTT and ROS assays were performed to assess the cell viability and oxidative stress within the cells, followed by mixed-member proportional (Mitochondrial membrane potential), dual staining, and immunoblotting. 6-OHDA-induced cell death in SH-SY5Y cells was dose-dependently attenuated by treatment with anthraquinone. The genes responsible for mutation were studied and the mutated RNAs knockout possibilities was studied using CRISPR spcas9 genome editing tool. Treatment with anthraquinone attenuated the level of oxidative stress and reduced the mitochondrial dysfunction associated with 6-OHDA treatment. Immunoblot analysis carried out with apoptotic markers showed that cytochrome C and caspase-3 expression increased significantly in anthraquinone-treated cells, whereas 6-OHDA-treated group showed a significant decrease when compared with an experimental control group. The mutated genes PARK7, PINK1, and SNCA1 were analyzed and found to exhibit four gene knock possibilities to treat PD. Reports demonstrate that other than following up on the biosynthesis of dopamine and its metabolites, these mixes counteract D2 receptors' extreme touchiness. It is proposed that further examinations need be directed to better understand the activity of the bioactive mixes circulated in these edible fungi Pleurotus ostreatus. The gene knockout possibilities identified by CRISPR SpCas9 will pave a way for better research for PD treatment.

Rapid colorimetric detection of mercury using silver nanoparticles in the presence of methionine.

Development of potential sensors is inevitable for the detection of environmental pollutants including toxins, organic pollutants and heavy metal which cause hazardous effect to human and other living organisms. The present study is to develop silver nanoparticle (Ag NPs) based sensor for the accurate, sensitive and selective colorimetric detection of Hg2+ ions from aqueous samples at nano molar level. The nanoparticles were synthesized chemically and it was stabilized by polyvinylpyrrolidone (PVP). The prepared particles were characterized by UV-visible spectroscopy, scanning electron microscopy (SEM), particle analysiser and Zetasizer. The UV-visible spectra of Ag NPs showed absorbance maximum at 392 nm. The average diameter of Ag NPs was determined to be 6 ±â€¯0.9 nm by using particle analyzer. The zeta sizer analysis showed that the PVP stabilized Ag NPs possessed a zeta potential of -35.56 ±â€¯3 mV. The Ag NPs-methionine conjugate showed the colour change from the brownish yellow colour to colourless when it was reacted with mercury. The Ag NPs conjugated methionine is sensitive to mercury and detects the mercury at nano molar level. The influence of other metal ion did not interfere with the detection and quantification of Hg2+. The detection of Hg2+ was also performed with paper strip and agarose gel method. The Ag NPs conjugate with methionine can applied for the detection of Hg2+ from various aqueous samples.

An edible fungi Pleurotus ostreatus inhibits adipogenesis via suppressing expression of PPAR γ and C/EBP α in 3T3-L1 cells: In vitro validation of gene knock out of RNAs in PPAR γ using CRISPR spcas9.

OBJECTIVE: Obesity is now well recognized as a disorder, one that is essentially preventable through changes in lifestyle. Obesity is also a main concern associated with expanded morbidity and mortality from many noncommunicable illnesses (NCDs). The study aimed to determine the antiobesity effect of Pleurotus ostreatus (PO) and its bioactive anthraquinone (AQ). The overall promoter genes CEBPα (CCAAT enhancer binding protein α) and PPARγ (Peroxisome proliferator activated receptor γ) in controlling the homeostasis of glucose was analysed using 3T3-L1 cell line. Finally, an insilico study was carried out using CRISPR software to identify the RNA's involved in adipogenesis especially of the control gene PPARγ. MATERIALS AND METHODS: Preliminary screening of the edible fungi and their bio actives led to the marvellous discovery of side effect free agonists for treating obesity (adipogenesis). An edible fungi Pleurotus ostreatus (PO) were analysed in a screening platform with different series of tests for adipocyte differentiation, triglyceride analysis, lipolysis determination, glucose uptake assay, cytotoxicity assay and lipase activity followed by specific gene expression analysis. The gene knockout mechanism was also elucidated by CRISPR spcas 9 tool. RESULTS: The antiadipogenic (antiobesity) activity of DMSO extract of PO were found to stimulate the insulin dependent uptake of glucose. The extract also decreased the levels of triglycerides and glycerol accumulation in differentiated adipocyte cells. The binding FABP4 (Fatty acid binding protein) and transport protein FATP1 (Fatty acid transport protein) along with the fat breaking LPL (lipoprotein lipase) was found to be inhibited after the PO treatment at varying concentration (0-300 µg/ml). CRISPR spcas9 genome editing software was used as an insilico approach in validating the efficiency of mouse embryonic and human adipogenic cell line (3T3-L1). These tool analysed and found 4 RNAs gene knock out possibilities in PPARγ and their efficiency for further treating obesity. CONCLUSION: These novel finding contribute to the confirmation that edible fungi PO and it's bioactive AQ is an adequate supplement for constraining the lipid and triglycerides in differentiated mature adipocytes by reversing the fat deposition. Thereby, forbidding the enzymes linked with fat absorption. Besides, the CRISPR tool identified gene knock out possibilities of control gene PPARγ, will pave a way in further research for treating obesity.

Evaluation of Therapeutic Potential of Eugenol-A Natural Derivative of Syzygium aromaticum on Cervical Cancer

Background: The intendment of this study is to determine the pursuance in ­ vitro anticancer activity and cytotoxicity of Syzygium aromaticum against the human cervical cancer cell line (HeLa) compared to the normal cell lines. Apoptogenic properties of DCM extract of Eugenol was determined in this entire study. Materials and Methods: HeLa cell lines were cultured in DMEM medium and incubated with different concentration of DCM ­ Eugenol extract. MTT assay brought out the way to determine the cell viability and quantification was done with the optical absorbance at 570 nm and 620 nm as reference. Apoptotic cells were affirmed by dual staining using acridine orange bromide. Besides, the morphology of the nucleus was also confirmed by dual staining. Eugenol inhibited 50% growth (IC50) of HeLa cell lines at 200 mg/ml of extract concentration. Results: Inhibitory efficacy of eugenol isolated from Syzyzgyium aromaticum showed the cell ­ viability in time and dose dependent manner with consistent morphological changes. Flow cytometer determined the apoptosis confirming the cytotoxicity value for MTT at IC50 with 81.85% cell viability. Dual staining firmly enacts the damaged cells due to AO indicating apoptosis confirmation by dual staining. Morphological analysis also clearly states that nil apoptosis has been seen in control and similarly in eugenol treated when compared to cancerous HeLa cell ­ line. Conclusion: Evaluation of cytotoxicity effect of eugenol isolated from Syzygium aromaticum showed it can be unrivalled dormant source of prodigious changes in HeLa cell line indicating (revealing) that chemotherapeutic agent.

Extraction and Characterization of Chick PEA (Cicer arietinum) Extract with Immunostimulant Activity in BALB/C MICE

Traditional plant medicines are used for a range of cancer conditions. The chickpea is highly proteinaceous and consumed as a staple in many parts of the world. An evaluation of chemoprotective and immunomodulatory activities of Cicer arietinum (CE) in cisplatin-induced immunosuppressed mice was here performed. Cisplatin was given at 100mg/kg, intraperitoneally, and after induction of immunosuppression mice were treated with Cicer arietinum (0.5 mg/dose/animal/IP) for a period of 10-day. The influence of the extract on lymphoid organ weight, bone marrow cellularity, alpha esterase activity and on enzyme levels such as (SGOT,SGPT, Urea, Creatinine was assessed to identify any chemoprotective influence. Administration of CE to cisplatin-treated mice alleviated the drastic reduction in bone marrow cellularity and α- esterase positive cells seen with cisplatin. Thus myelosuppression was reversed or inhibited. Cisplatin bids to DNA and causes damage resulting in chromosome breaks, micronucleus formation and cell death. CE is comprised of numerous middle-chain aliphatic alcohols, aldehydes and ketones, in addition to compounds like 7-hydroxy-1-methoxy-6-methylanthraquinone, cyclohexadecane (CAS) and 6-(amino methyl)-2-naphthol. These latter are thought to contribute to the characteristic aroma of C. arietimnum. In conclusion, administration of CE in cisplatin-treated mice, boosted bone marrow cellularity and increased α- esterase positive cells, thus reversing myelosupproession.