Biochemical and thermodynamic properties of de novo synthesized urease from Vicia sativa seeds with enhanced industrial applications.

Urease is one of the most significant enzymes in the industry. The objective of this research was to isolate and partially purify urease from Vicia sativa seeds with urease characterization. With a 6.4 % yield, the purification fold was 9.0. By using chromatography, it was determined that the isolated urease had a molecular weight of 55 kDa. The maximum urease activity was found following a 60-s incubation period at 40 °C and pH 8. The activity of urease was significantly boosted by a mean of calcium, barium, DL-dithiothreitol, Na2EDTA, and citrate (16.9, 26.6, 18.6, 13.6, and 31 %), respectively. But nickel and mercury caused inhibitory effects and completely inhibited urease activity, indicating the presence of a thiol (-SH) group in the enzyme active site. The Arrhenius plot was used to analyze the thermodynamic constants of activation, Ea, ΔH*, ΔG*, and ΔS*. The results showed that the values were 30 kJ/mol, 93.14 kJ/mol, 107.17 kJ/mol/K, and -40.80 J/mol/K, respectively. The significance of urease extraction from various sources may contribute to our understanding of the metabolism of urea in plants. The current report has novelty as it explained for the first time the kinetics and thermodynamics of hydrolysis of urea and inactivation of urease from V. sativa seeds.

Development of Nanosuspension of Artemisia absinthium Extract as Novel Drug Delivery System to Enhance Its Bioavailability and Hepatoprotective Potential.

A nanosuspension of Artemisia absinthium extract was formulated and characterized for the enhancement of bioavailability and better hepatoprotective efficacy. The nanosuspension of A. absinthium extract was formulated using an antisolvent precipitation technique, and various formulation parameters were optimized using response surface methodology (RSM). The optimized nanosuspension was characterized using AFM and FT-IR spectroscopy. The drug-release profile and oral bioavailability of the optimized nanosuspension were assessed with reference to coarse suspension. The DPPH radical scavenging method was used to measure the nanosuspension's antioxidant activity, and its in vivo hepatoprotective potential was assessed against CCl4-induced hepatic injury in rats. The developed optimized nanosuspension had suitable zeta potential of -11.9 mV, PDI of 0.285, and mean particle size of 253.8 nm. AFM study demonstrated a homogeneous population of nanoparticles with average size of 25 nm. The formulated nanosuspension of A. absinthium showed faster dissolution rate and 1.13-fold enhanced bioavailability as compared to the coarse suspension (plant extract). Furthermore, the nanoformulation had stronger antioxidant and hepatoprotective potential as compared to the unprocessed coarse extract. These results demonstrated that nanosuspension is a promising strategy for improving the oral bioavailability and bioactivities of A. absinthium extract.

Aspergillus nomiae and fumigatus Ameliorating the Hypoxic Stress Induced by Waterlogging through Ethylene Metabolism in Zea mays L.

Transient and prolonged waterlogging stress (WS) stimulates ethylene (ET) generation in plants, but their reprogramming is critical in determining the plants' fate under WS, which can be combated by the application of symbiotically associated beneficial microbes that induce resistance to WS. The present research was rationalized to explore the potential of the newly isolated 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase-producing fungal endophytic consortium of Aspergillus nomiae (MA1) and Aspergillus fumigatus (MA4) on maize growth promotion under WS. MA1 and MA4 were isolated from the seeds of Moringa oleifera L., which ably produced a sufficient amount of IAA, proline, phenols, and flavonoids. MA1 and MA4 proficiently colonized the root zone of maize (Zea mays L.). The symbiotic association of MA1 and MA4 promoted the growth response of maize compared with the non-inoculated plants under WS stress. Moreover, MA1- and MA4-inoculated maize plants enhanced the production of total soluble protein, sugar, lipids, phenolics, and flavonoids, with a reduction in proline content and H2O2 production. MA1- and MA4-inoculated maize plants showed an increase in the DPPH activity and antioxidant enzyme activities of CAT and POD, along with an increased level of hormonal content (GA3 and IAA) and decreased ABA and ACC contents. Optimal stomatal activity in leaf tissue and adventitious root formation at the root/stem junction was increased in MA1- and MA4-inoculated maize plants, with reduced lysigenous aerenchyma formation, ratio of cortex-to-stele, water-filled cells, and cell gaps within roots; increased tight and round cells; and intact cortical cells without damage. MA1 and MA4 induced a reduction in deformed mesophyll cells, and deteriorated epidermal and vascular bundle cells, as well as swollen metaxylem, phloem, pith, and cortical area, in maize plants under WS compared with control. Moreover, the transcript abundance of ethylene-responsive gene ZmEREB180, responsible for the induction of the WS tolerance in maize, showed optimally reduced expression sufficient for induction in WS tolerance, in MA1- and MA4-inoculated maize plants under WS compared with the non-inoculated control. The existing research supported the use of MA1 and MA4 isolates for establishing the bipartite mutualistic symbiosis in maize to assuage the adverse effects of WS by optimizing ethylene production.

Green inspired synthesis of zinc oxide nanoparticles using Silybum marianum (milk thistle) extract and evaluation of their potential pesticidal and phytopathogens activities.

Background: The green approaches for the synthesis of nanoparticles are gaining significant importance because of their high productivity, purity, low cost, biocompatibility, and environmental friendliness. Methods: The aim of the current study is the green synthesis of zinc oxide nanoparticles (ZnO-NPs) using seed extracts of Silybum marianum, which acts as a reducing and stabilizing agent. central composite design (CCD) of response surface methodology (RSM) optimized synthesis parameters (temperature, pH, reaction time, plant extract, and salt concentration) for controlled size, stability, and maximum yields of ZnO-NPs. Green synthesized ZnO-NPs was characterized using UV-visible spectroscopy and Zetasizer analyses. Results: The Zetasizer confirmed that green synthesized ZnO-NPs were 51.80 nm in size and monodispersed in nature. The UV-visible results revealed a large band gap energy in the visible region at 360.5 nm wavelength. The bioactivities of green synthesized ZnO-NPs, including antifungal, antibacterial, and pesticidal, were also evaluated. Data analysis confirmed that these activities were concentration dependent. Bio-synthesized ZnO-NPs showed higher mortality towards Tribolium castaneum of about 78 ± 0.57% after 72 h observation as compared to Sitophilus oryzae, which only displayed 74 ± 0.57% at the same concentration and time intervals. Plant-mediated ZnO-NPs also showed high potential against pathogenic gram-positive bacteria (Clavibacter michiganensis), gram-negative bacteria (Pseudomonas syringae), and two fungal strains such as Fusarium oxysporum, and Aspergillums niger with inhibition zones of 18 ± 0.4, 25 ± 0.4, 21 ± 0.57, and 19 ± 0.4 mm, respectively. Conclusion: The results of this study showed that Silybum marianum-based ZnO-NPs are cost-effective and efficient against crop pests.

Investigation of phytotherapeutic potential of herbal mixtures and their effects on salbutamol induced cardiotoxicity and hyperlipidemia in rabbits.

BACKGROUND: Cardiovascular diseases (CVDs) are the major cause of deaths all over the world. The high level of blood cholesterol and oxidative stress are major risk factors for heart diseases. The phytotherapeutics have attracted attention as potential agents for preventing and treating oxidative stress associated diseases. The objective of present study was to evaluate the synergetic cardio-protective and antilipidemic potential of medicinal plants viz. Coriandrum sativum, Piper nigrum and Cactus grandiflorus. Cardio-protective and anti-lipidemic potential of herbal mixture was evaluated against salbutamol induced cardiotoxicity in rabbits. For this purpose, rabbits were divided into six groups as normal control, salbutamol control, curative and standard drug curative. RESULTS: Salbutamol significantly (p < 0.05) increased the level of serum cardiac biomarkers (ALT, CK-MB, AST and LDH) and lipids (LDL, triglycerides, cholesterol) in rabbits. The prior and post administration of herbal mixture significantly (p < 0.05) lowered the elevated level of serum cardiac biomarkers and lipids equal to normal control. Gross pathological examination revealed that heart of salbutamol control animals became hardened, congested and were enlarged than preventive and curative groups. The phytotherapeutic analysis of medicinal plants revealed the presence of phenols, tannins, alkaloids and steroids. CONCLUSION: The results showed that this herbal mixture has strong cardio-protective and anti-lipidemic potential.

Mir-153-3p Modulates the Breast Cancer Cells' Chemosensitivity to Doxorubicin by Targeting KIF20A.

Breast cancer is considered the solid tumor most sensitive to chemotherapy. However, it can become resistant to various chemotherapeutic drugs, including doxorubicin, which triggers cell death by intercalation between DNA bases, free radical formation, and topoisomerase II inhibition. When drug resistance develops, several miRNAs are dysregulated, suggesting that miRNAs may play a significant role in resistance formation. In the current study, we investigated how doxorubicin sensitivity of breast cancer cells is affected by miR-153-3p and its target gene. The MTT method was used to determine the chemo-sensitizing effect of miR-153-3p on doxorubicin in MCF-7 and MDA-MB-231 cell lines. Results of Western blot and dual luciferase confirmed that miR-153-3p targets KIF20A and decreases its expression. Transwell and flow cytometry experiments showed that miR-153-3p and doxorubicin together had higher effects on MCF-7 and MDA-MB-231 cell proliferation, migration, and invasion, as well as increasing apoptosis and arresting cells in the G1 phase. Proteins related to apoptosis and the cell cycle exhibited the same tendency. Intracellular vesicle formation was inhibited and RAB26 was also downregulated by treatment with miR-153-3p alone or in combination with doxorubicin. Doxorubicin's ability to suppress tumors may be enhanced by miR-153-3p, according to in vivo studies. According to our findings, miR-153-3p has a direct effect on KIF20A and may regulate the formation of intracellular vesicles, which in turn makes breast cancer cells more susceptible to doxorubicin.

Evaluation and optimization of nanosuspensions of Chrysanthemum coronarium and Azadirachta indica using Response Surface Methodology for pest management.

The rapidly emerging field of nanotechnology is considered an important achievement in the agriculture sector to increase the pest mortality rate and improve the crop production. The present study evaluates the novel pesticidal and anti-microbial activities of Chrysanthemum coronarium and Azadirachta indica in the nano-suspensions form. The anti-solvent precipitation method was used to formulate nano-suspensions proposed by Response Surface Methodology (RSM). Physicochemical nature of plant extracts and nano-suspensions was characterized through analysis of Zeta-sizer, FT-IR, and HPLC. Characterization results revealed a minimum particle size of 121.1 and 170.1 nm for Chrysanthemum coronarium and Azadirachta indica, respectively. The pesticidal activity of nano-suspension was performed against red flour beetle (RFB) and lesser grain borer (LGB) pests, which showed the maximum mortality rate of 100% with 100% concentration of plant extracts and nano-suspensions of Chrysanthemum coronarium and Azadirachta indica against both insects. In comparison, the combination of these both plant extracts revealed the maximum 100% mortality with a 50% concentration of nano-suspensions (mixing ratio 1:1) after 72 h. The antibacterial activity showed the maximum zone inhibition of 9.96 ± 0.17 and 14.17 ± 0.50 mm against S.aureus and E. coli with nano-suspension of Chrysanthemum coronarium, and 12.09 ± 0.11 and 14.10 ± 0.49 mm with nano-suspension of Azadirachta indica, respectively. It is concluded that individual nano-suspensions showed better pesticidal as well as antimicrobial activities than combinations. However, the constructed nanosuspension can be applied to control the plant pests and diseases simultaneously.

Natural Polyphyllins (I, II, D, VI, VII) Reverses Cancer Through Apoptosis, Autophagy, Mitophagy, Inflammation, and Necroptosis.

Cancer is the second leading cause of mortality worldwide. Conventional therapies, including surgery, radiation, and chemotherapy, have limited success because of secondary resistance. Therefore, safe, non-resistant, less toxic, and convenient drugs are urgently required. Natural products (NPs), primarily sourced from medicinal plants, are ideal for cancer treatment because of their low toxicity and high success. NPs cure cancer by regulating different pathways, such as PI3K/AKT/mTOR, ER stress, JNK, Wnt, STAT3, MAPKs, NF-kB, MEK-ERK, inflammation, oxidative stress, apoptosis, autophagy, mitophagy, and necroptosis. Among the NPs, steroid saponins, including polyphyllins (I, II, D, VI, and VII), have potent pharmacological, analgesic, and anticancer activities for the induction of cytotoxicity. Recent research has demonstrated that polyphyllins (PPs) possess potent effects against different cancers through apoptosis, autophagy, inflammation, and necroptosis. This review summarizes the available studies on PPs against cancer to provide a basis for future research.

Molecular mechanisms of anticancer activities of polyphyllin VII.

Cancer is the leading cause of mortality in the world. The major therapies for cancer treatment are chemotherapy, surgery, and radiation therapy. All these therapies expensive, toxic and show resistance. The plant-derived compounds are considered safe, cost-effective and target cancer through different pathways. In these pathways include oxidative stress, mitochondrial dependent and independent, STAT3, NF-kB, MAPKs, cell cycle, and autophagy pathways. One of the new plants derived compounds is Polyphyllin VII (PPVII), which target cancer through different molecular mechanisms. In literature, there is a review gap of studies on PPVII; therefore in the current review, we summarized the available studies on PPVII to provide a base for future research.

Identification of Hypotensive Biofunctional Compounds of Coriandrum sativum and Evaluation of Their Angiotensin-Converting Enzyme (ACE) Inhibition Potential.

The aim of this study was to identify and characterize the bioactive compounds of Coriandrum sativum responsible for the treatment of hypertension and to explore their mechanism of action as angiotensin-converting enzyme (ACE) inhibitors. Bioactive fractions like alkaloids, flavonoids, steroids, and tannins were extracted and evaluated for their ACE inhibition potential. Among them, only flavonoid-rich fraction showed high ACE inhibition potential with IC50 value of 28.91 ± 13.42 µg/mL. The flavonoids were characterized through LC-ESI-MS/MS. Seventeen flavonoids were identified in this fraction of Coriandrum sativum in negative ionization mode which includes pinocembrin, apigenin, pseudobaptigenin, galangin-5-methyl ether, quercetin, baicalein trimethyl ether, kaempferol dimethyl ether, pinobanksin-5-methylether-3-O-acetate, pinobanksin-3-O-pentenoate, pinobanksin-3-O-phenylpropionate, pinobanksin-3-O-pentanoate, apigenin-7-O-glucuronoide, quercetin-3-O-glucoside, apigenin-3-O-rutinoside, rutin, isorhamnetin-3-O-rutinoside, and quercetin dimethyl ether-3-O-rutinoside, while six flavonoids including daidzein, luteolin, pectolinarigenin, apigenin-C-glucoside, kaempferol-3-7-dimethyl ether-3-O-glucoside, and apigenin-7-O-(6-methyl-beta-D-glucoside) were identified in positive ionization mode. The results of this study revealed that Coriandrum sativum is a valuable functional food that possesses a number of therapeutic flavonoids with ACE inhibition potential that can manage blood pressure very efficiently.

Hypoglycemic potential of herbal product dolabi compared with pioglitazone in streptozotocin-induced diabetic rats.

OBJECTIVES: To investigate and compare the hypoglycemic potential of commonly used polyherbal formulation dolabi with pioglitazone in streptozotocin-induced diabetic rats. METHODS: A total of 24 adult male Wistar rats were randomized into diabetic control group, diabetic group receiving dolabi (17.6 mg kg-1 day-1), diabetic group receiving dolabi (35.2 mg kg-1 day-1) and diabetic group receiving pioglitazone (2.7 mg kg-1 day-1), with 6 rats in each group. The study was conducted for 4 weeks. Blood samples were collected for fructosamine estimation, fasting plasma glucose (FPG) and oral glucose tolerance test (OGTT) were measured at different time points. RESULTS: Compared with baseline, the groups receiving dolabi (35.2 mg kg-1 day-1) and pioglitazone showed signififi cant reductions (P<0.05) in fructosamine levels at the end point and same was the case for FPG and OGTT with pioglitazone showing greater hypoglycemic potential. However, the group receiving dolabi (17.6 mg kg-1 day-1) showed significant reduction (P<0.05) only in FPG level and failed to achieve significant changes either in fructosamine level or OGTT. Moreover, signififi cant differences (P<0.05) in FPG and fructosamine levels were observed between groups receiving dolabi (17.6 mg kg-1 day-1) and pioglitazone at the end point. CONCLUSIONS: These results showed sluggish hypoglycemic effects of dolabi at manufacturer's recommended doses. At a higher dose however, good glycemic control was achieved with dolabi and the results were comparable to pioglitazone. The shorter duration of study (4 weeks) might be the reason of poor glycemic control associated with dolabi at a normal dose.

Benefits of Zubex beyond glycemic control: Evidence of the antiatherogenic effect.

OBJECTIVE: To determine the effect of Zubex (aqueous extract of Curcuma longa linn, Iron murakab, Eugenia jambolana, Lin seed or flflax seed, processed egg shell calcium and Asphaltum) on plasma sialic acid (PSA) along with cardiovascular (CV) risk factors in streptozotocin-induced diabetic rats. METHODS: Thirty male albino rats were divided into groups (1-6) with 5 rats in each group. Group 1 and 2 served as normal control (NC) and diabetic control (DC), respectively, and were given normal saline only. Groups 3-5 were given Zubex in different doses 100-400 mg/(kg day). Group 6 received glibenclamide 600 µg/(kg day) orally as a reference drug. Fasting plasma glucose (FPG) and PSA levels were determined at baseline after every 2 weeks for 10 weeks. The other parameters including blood lipids and hepatic enzymes were determined at baseline and at the end of the study. Finally, the liver was subjected to histological examination. RESULTS: Compared with DC group, Zubex treated groups showed signifificant decline in FPG levels (P<0.05). At the endpoint, the decrease in PSA concentration was significant (P<0.05) from baseline at the doses of 200 and 400 mg/(kg day) only and insignifificant at the dose of 100 mg/ (kg day). Statistically signifificant improvements were observed in blood lipids at the doses of 200 and 400 mg/kg (P<0.05) compared with DC; but, the improvement was insignifificant in low density lipoprotein-cholesterol and high density lipoprotein-cholesterol at the dose of 100 mg/ (kg day). Signifificant decreases were also found in hepatic enzyme levels at all the doses of Zubex (P<0.05). Histological examination showed marked improvement in streptozotocin induced liver injury after treatment of all the 3 doses of Zubex. CONCLUSION: Zubex could ameliorate PSA and other diabetic complications effectively and may be a useful alternative/adjuvant in diabetes treatment.

Production of rabbit antibodies against purified Glucose oxidase

Glucose oxidase is an active oxygen species generating enzyme produced from Aspergillus niger grown in submerged fermentation. Disintegration of the mycelium resulted in high glucose oxidase activity that was subjected to ammonium sulfate precipitation at 60-85% saturation rates that resulted to 6.14 U mg -1 specific activity. Purification of enzyme by anion exchange column (DEAE-Cellulose) resulted into 22.53 U mg-1 specific activity and 10.27 fold purification. This was applied to sephadex G-200 column for gel filtration chromatography. It was observed that enzyme achieved 59.37 U mg-1of specific activity with 27.08 fold purity and 64.36% recovery. Purified glucose oxidase was injected into rabbits through intravenous route, to raise the glucose oxidase antibodies. After 30 days incubation period, the rabbits were slaughtered and serum was separated from blood. The antibodies were isolated by ammonium sulfate precipitation and confirmed by agar gel precipitation test. This could be a convenient and low cost alternate assay for the estimation of glucose oxidase in biological fluids. Moreover, such antibodies against the said enzyme could be used in various therapeutic and diagnostic applications.