Mosquito larvicidal activity of pyrrolidine-2,4-dione derivatives: An investigation against Culex quinquefasciatus and molecular docking studies

The pyrrolidine-2,4-dione derivatives were used to conduct a larvicidal test on Culex quinquefasciatus larvae of the second instar. Mannich base condensation method was used to synthesis the pyrrolidine-2,4-dione derivatives by grindstone method. The reaction conditions were mild, resulting in high yields. An analysis of the synthesized compounds was carried out using FTIR, ¹H NMR, ¹³C NMR, mass spectrometry, and elemental analysis. Synthesized compounds (1a-h) were evaluated for larvicidal activities. Compound 1e (LD₅₀: 26.06 µg/mL), and 1f (LD₅₀: 26.89 µg/mL), and were notably more active against Culex quinquefasciatus than permethrin (LD₅₀: 26.14 µg/mL). The docking studies also demonstrated that 1e, and 1f are potent larvicides with higher binding energy (?12.6 kcal/mol) than the control in the mosquito odorant binding protein (PDB ID: 3OGN). The larvicidal properties of lead molecules have made them important for use as insecticides.     

Permethrin and malathion LD90 values for Culex quinquefasciatus vary with topical application site

Prior research in multiple insect species has demonstrated that insecticide‐induced mortality varies according to the body region exposed on the insect. This variation has been demonstrated in Culex quinquefasciatus Say (Diptera: Culicidae), but has not been quantified using dose–response curves. Applications of technical permethrin or malathion to one of three body regions on Cx. quinquefasciatus resulted in dose–response curves that were not equivalent to one another. The generated LD₉₀ values and curves for each body region were compared with previously reported LD values for analogous sites in several mosquito species, specifically the mesothorax. Based on the present results, the permethrin and malathion LD₅₀ and LD₉₀ concentrations required for droplets impinging on the abdomen and mesothorax of Cx. quinquefasciatus when applied through ground‐based spray systems utilized by mosquito control programmes were calculated.     

Efficacy of leaves extract of Calotropis procera Ait. (Asclepiadaceae) in controlling Anopheles arabiensis and Culex quinquefasciatus mosquitoes

The present study aimed to investigate, the larvicidal, adult emergence inhibition and oviposition deterrent activity of aqueous leaves extract of Calotropis procera against Anopheles arabiensis and Culex quinquefasciatus as natural mosquito larvicide. The larvicidal activity was monitored against 2nd, 3rd and 4th instar larvae of each mosquito species 24 h post-treatment. Adult emergence inhibition activity was tested by exposing 3rd instar larvae of each mosquito species to different concentrations of extracts (200, 400, 600, 800 and 1000 ppm for An. arabiensis and 100, 200, 300, 400, 500 and 600 ppm for Cx. quinquefasciatus). Probit analysis was used to analyze data from bioassay experiments. The oviposition deterrent activity was tested by using three different concentrations of extracts (1000, 500 and 200 for An. arabiensis, and 1000, 500 and 100 for Cx. quinquefasciatus) that caused high, moderate and low larval mortality in the larvicidal experiment against 3rd instar larvae. It was found that, LC50–LC90 values calculated were 273.53–783.43, 366.44–1018.59 and 454.99–1224.62 ppm for 2nd, 3rd and 4th larval instars, respectively, of An. arabiensis and 187.93–433.51, 218.27–538.27 and 264.85–769.13 ppm for 2nd, 3rd and 4th larval instars, respectively, of Cx. quinquefasciatus. Fifty percent of adult emergence inhibition (EI50) was shown at 277.90 and 183.65 ppm for An. arabiensis and Cx. quinquefasciatus, respectively. The pupal stage was not affected till a concentration of 5000 ppm. The extract showed oviposition deterrence and effective repellence against both mosquito species at different concentrations, with the observation on that maximal eggs were laid in low concentration of extract. These results suggest that the leaves extract of C. procera possess remarkable larvicidal, adult emergence inhibitor, repellent and oviposition deterrent effect against both An. arabiensis and Cx. quinquefasciatus, and might be used as natural biocides for mosquito control.     

Cytotoxic,larvicidal, nematicidal,and antifeedant activities of piperidin-connected 2-thioxoimidazolidin-4-one derivatives

The objective of this study was to investigate brine shrimp cytotoxicity, larvicidal, nematicidal, and antifeedant activities of novel piperidin-connected 2-thioxo-imidazolidin-4-one derivatives. The activities of target compounds were compared with some naturally occurring (?)-pinidinol, hydantocidin, and positive controls. Target compounds were synthesized via cyclocondensation method. The compounds were synthesized and then characterized by infrared spectroscopy, ¹H NMR, ¹³C NMR, mass spectral, and elemental analyses. Brine shrimp cytotoxicity assay was investigated using freshly hatched, free-swimming nauplii of Artemiasalina. Larvicidal screening was performed against urban mosquito larvae (Culex quinquefasciatus). Nematicidal activity was evaluated using juvenile nematodes of Meloidogyne javanica. Regarding antifeedant activity, marine-acclimated Oreochromis mossambicus fingerlings were used. Compounds 3a-c (piperidin-connected 2-thioxoimidazolidin-4-one) were found to be lethal to the second instar larvae of mosquito, which produced LD₅₀ values of 1.37, 6.66, 6.51 μg/mL, compared to compounds (?) pinidinol and hyantocidin LD₅₀ values of 18.28 and 22.11 μg/mL respectively. Compound 3a-c was found to kill 100% of fish fingerlings within 6 h at 20 µg/mL, with LD₅₀ values of 1.54, 1.79, 1.52 µg/mL, compared to compounds (?) pinidinol and hyantocidin with LD₅₀ values of 10.21 and 21.05 μg/mL respectively. Compound 3c with LD₅₀ value of 1.57 μg/mL demonstrated high nematicidal activity compared to compound 3a, 3b, (?) Pinidinol and Hyantocidin LD₅₀ values of 6.45, 2.42, 14.25, 26.30 μg/mL respectively. Therefore, the 2-thioxoimidazolidin-4-one with piperidin ring showed high potential cytotoxic, larvicidal, nematicidal, and antifeedent activities.     

Repellent,larvicidal and adulticidal activities of essential oil from Dai medicinal plant Zingiber cassumunar against Aedes albopictus

Zingiber cassumunar is an important plant used in traditional medicine and as a natural mosquito repellent. However, the compounds responsible for the repellent activity of the plant are still unknown. The aim of the study is to identify the components of Z. cassumunar essential oil that show repellent activity against Aedes albopictus. We also evaluated the larvicidal and adulticidal activities of Z. cassumunar essential oil against Ae. albopictus. In-cage mosquito repellent experiments showed that Z. cassumunar essential oil possessed moderate repellent activity with a minimum effective dose (MED) of 0.16 ± 0.01 mg/cm², compared to reference standard N,N-diethyl-3-methylbenzamide (DEET, 0.03 ± 0.01 mg/cm²). Bioassay-guided fractionation identified the major active compound of Z. cassumunar essential oil as (−)-terpinen-4-ol (1) (MED: 0.19 ± 0 mg/cm²). We also found that Z. cassumunar essential oil showed moderate larvicidal activity against first instar larvae of Ae. albopictus with a LC₅₀ (50% lethal concentration) of 44.9 μg/L after 24 h. Fumigation bioassays showed that Z. cassumunar essential oil exhibits moderate adulticidal activity against Ae. albopictus with a LC₅₀ of 5.44%, while (−)-terpinen-4-ol showed significant adulticidal activity with a LC₅₀ of 2.10% after 24 h. This study verifies that the Z. cassumunar essential oil has mosquito repellent activity, and that (−)-terpinen-4-ol is mainly responsible for this activity. Furthermore, this study provides scientific support for the folk usage of Z. cassumunar essential oil as mosquito repellent and indicates that Z. cassumunar essential oil and (−)-terpinen-4-ol can be used as plant-derived repellents and insecticides for mosquito control.     

Olive leaf extract modulates permethrin induced genetic and oxidative damage in rats

Permethrin is a common synthetic chemical, widely used as an insecticide in agriculture and other domestic applications. The previous reports indicated that permethrin is a highly toxic synthetic pyrethroid pesticide to human and environmental health. Therefore, the present experiment was undertaken to determine the effectiveness of olive leaf extract in modulating the permethrin induced genotoxic and oxidative damage in rats. The animals used were broadly divided into four (A, B, C and D) experimental groups. Group A rats served as control animals and received distilled water intraperitoneally (n = 5). Groups B and C rats received intraperitoneal injections of permethrin (60 mg kg⁻¹ b.w) and olive leaf extract (500 mg kg⁻¹ b.w), respectively. Group D rats received permethrin (60 mg kg⁻¹ b.w) plus olive leaf extract (500 mg kg⁻¹ b.w). Rats were orally administered their respective feed daily for 21 days. At the end of the experiment rats were anesthetized and serum and bone marrow cell samples were obtained. Genotoxic damage was assessed by micronucleus and chromosomal aberration assays. Total antioxidant capacity and total oxidant status were also measured in serum samples to assess oxidative status. Treatment of Group B with permethrin resulted in genotoxic damage and increased total oxidant status levels. Permethrin treatment also significantly decreased (P < 0.05) total antioxidant capacity level when compared to Group A rats. Group C rats showed significant increases (P < 0.05) in total antioxidant capacity level and no alterations in cytogenetic parameters. Moreover, simultaneous treatments with olive leaf extract significantly modulated the toxic effects of permethrin in Group D rats. It can be concluded that olive leaf extract has beneficial influences and could be able to antagonize permethrin toxicity. As a result, this investigation clearly revealed the protective role of olive leaf extract against the genetic and oxidative damage by permethrin in vivo for the first time.     

球形芽孢杆菌C3-41对致倦库蚊的毒效及在蚊体内的再循环

球形芽孢杆菌Csub3-41菌株(Bacillus sphaericus C3-41)对致倦库蚊(Culex puinquefa-seiatus)幼虫有很高毒效,对2龄和3-4龄幼虫的半致死剂量(LD₅₀)分别为63.1 和89.7芽孢/蚊幼虫。处理浓度越高,取食时间越长,蚊幼虫取食到的杀蚊活性物质量越多,死亡率越高。当蚊幼虫取食亚致死剂量杀蚊活性物质后,球形芽孢杆菌在感染的活幼虫体内不增殖;但当蚊幼虫取食致死剂量杀蚊活性物质后,蚊幼死亡,球形芽孢杆菌在死蚊幼虫体内增殖明显,6天内芽孢从感染初期的1.86X10²蚊幼虫增加到1.59X10⁶/蚊幼虫。芽孢在死蚊幼虫体内能正常萌发、生长、产孢和形成毒素。增殖的芽孢同样对致倦库蚊幼虫有较高毒力。     

Pressure-accelerated dissociation of amyloid fibrils in wild-type hen lysozyme

The dynamics of amyloid fibrils, including their formation and dissociation, could be of vital importance in life. We studied the kinetics of dissociation of the amyloid fibrils from wild-type hen lysozyme at 25°C in vitro as a function of pressure using Trp fluorescence as a probe. Upon 100-fold dilution of 8 mg ml⁻¹ fibril solution in 80 mM NaCl, pH 2.2, no immediate change occurred in Trp fluorescence, but at pressures of 50–450 MPa the fluorescence intensity decreased rapidly with time (k_obs = 0.00193 min⁻¹ at 0.1 MPa, 0.0348 min⁻¹ at 400 MPa). This phenomenon is attributable to the pressure-accelerated dissociation of amyloid fibrils into monomeric hen lysozyme. From the pressure dependence of the rates, which reaches a plateau at ∼450 MPa, we determined the activation volume ΔV^0‡ = −32.9 ± 1.7 ml mol(monomer)⁻¹ and the activation compressibility Δκ^‡ = −0.0075 ± 0.0006 ml mol(monomer)⁻¹ bar⁻¹ for the dissociation reaction. The negative ΔV^0‡ and Δκ^‡ values are consistent with the notion that the amyloid fibril from wild-type hen lysozyme is in a high-volume and high-compressibility state, and the transition state for dissociation is coupled with a partial hydration of the fibril.     

Relative Neurotoxicity of Ivermectin and Moxidectin in Mdr1ab (?/?) Mice and Effects on Mammalian GABA(A) Channel Activity

The anthelmintics ivermectin (IVM) and moxidectin (MOX) display differences in toxicity in several host species. Entrance into the brain is restricted by the P-glycoprotein (P-gp) efflux transporter, while toxicity is mediated through the brain GABA(A) receptors. This study compared the toxicity of IVM and MOX in vivo and their interaction with GABA(A) receptors in vitro. Drug toxicity was assessed in Mdr1ab(−/−) mice P-gp-deficient after subcutaneous administration of increasing doses (0.11–2.0 and 0.23–12.9 µmol/kg for IVM and MOX in P-gp-deficient mice and half lethal doses (LD₅₀) in wild-type mice). Survival was evaluated over 14-days. In Mdr1ab(−/−) mice, LD₅₀ was 0.46 and 2.3 µmol/kg for IVM and MOX, respectively, demonstrating that MOX was less toxic than IVM. In P-gp-deficient mice, MOX had a lower brain-to-plasma concentration ratio and entered into the brain more slowly than IVM. The brain sublethal drug concentrations determined after administration of doses close to LD₅₀ were, in Mdr1ab(−/−) and wild-type mice, respectively, 270 and 210 pmol/g for IVM and 830 and 740–1380 pmol/g for MOX, indicating that higher brain concentrations are required for MOX toxicity than IVM. In rat α1β2γ2 GABA channels expressed in Xenopus oocytes, IVM and MOX were both allosteric activators of the GABA-induced response. The Hill coefficient was 1.52±0.45 for IVM and 0.34±0.56 for MOX (p<0.001), while the maximum potentiation caused by IVM and MOX relative to GABA alone was 413.7±66.1 and 257.4±40.6%, respectively (p<0.05), showing that IVM causes a greater potentiation of GABA action on this receptor. Differences in the accumulation of IVM and MOX in the brain and in the interaction of IVM and MOX with GABA(A) receptors account for differences in neurotoxicity seen in intact and Mdr1-deficient animals. These differences in neurotoxicity of IVM and MOX are important in considering their use in humans.     

The comparative persistence of toxicity of Bacillus sphaericus strain 1593 and Bacillus thuringiensis serotype H-14 against mosquito larvae in different kinds of environments

Bacillus sphaericus strain 1593 and B. thuringiensis serotype H-14 were evaluated for persistence of toxicity against two species of mosquito larvae, Culex quinquefasciatus and Aedes aegypti, in a selected simulating plot in Bangkok. Both strains of bacteria demonstrated larvicidal activity towards both species of mosquito larvae. In tap water, the toxicity of B. sphaericus strain 1593 was found to be greater towards C. quinquefasciatus larvae than A. aegypti larvae, whereas the toxicity of B. thuringiensis serotype H-14 was found to be greater towards A. aegypti larvae than C. quinquefasciatus larvae. The persistence of toxicity of these two bacteria was found to be different. The lethal concentration of B. thuriengiensis H-14 against A. aegypti decreased from LC₉₀ to below LC₅₀ in about 15 weeks when tested in tap water. The decrease was faster in polluted water. The toxicity of B. sphaericus 1593 towards C. quinquefasciatus larvae persisted for at least 9 months in tap water and 6 months in polluted water. The multiplication of bacteria was indicated only in populations of B. sphaericus 1593 tested with C. quinquefasciatus larvae.     

Cyclopaldic Acid,Seiridin, and Sphaeropsidin A as Fungal Phytotoxins,and Larvicidal and Biting Deterrents against Aedes aegypti (Diptera: Culicidae): Structure?Activity Relationships

Aedes aegypti L. is the major vector of the arboviruses responsible for dengue fever, one of the most devastating human diseases. From a preliminary screening of fungal phytotoxins, cyclopaldic acid ( 1 ), seiridin ( 2 ), sphaeropsidin A ( 4 ), and papyracillic acid ( 5 ) were evaluated for their biting deterrent and larvicidal activities against Ae. aegypti L. Because compounds 1, 2, 4 , and 5 exhibited mosquito biting deterrent activities and 1 and 4 demonstrated larvicidal activities, further structure? activity relationship studies were initiated on these toxins. In biting‐deterrence bioassays, 1, 2, 4 , and 5 , 3,8‐didansylhydrazone of cyclopaldic acid, 1F , 5‐azidopentanoate of cyclopaldic acid A, 1G , the reduced derivative of cyclopaldic acid, 1 H , isoseiridin ( 3 ), 2′‐O‐acetylseiridin ( 2A ), 2′‐oxoseiridin ( 2C ), 6‐O‐acetylsphaeropsidin A ( 4A ), 8,14‐methylensphaeropsidin A methyl ester ( 4B ), and sphaeropsidin B ( 4C ) showed activities higher than the solvent control. Sphaeropsidin B ( 4C ) was the most active compound followed by 2A , while the other compounds were less active. Biting‐deterrence activity of compound 4C was statistically similar to DEET. In the larvicidal screening bioassays, only compounds 1 and 4 demonstrated larvicidal activities. Based on LD₅₀ values, compound 4 (LD₅₀ 36.8 ppm) was significantly more active than compound 1 (LD₅₀ 58.2 ppm). However, the activity of these compounds was significantly lower than permethrin.     

Evaluation of the toxicity of different phytoextracts of Ocimum basilicum against Anopheles stephensi and Culex quinquefasciatus

The larvicidal effect of the crude carbon tetrachloride, methanol and petroleum ether leaf extracts of a widely grown medicinal plant, Ocimum basilicum, against Anopheles stephensi and Culex quinquefasciatus was evaluated. Petroleum ether extract was found to be the most effective against the larvae of both mosquitoes, with LC₅₀ values of 8.29, 4.57; 87.68, 47.25 ppm and LC₉₀ values of 10.06, 6.06; 129.32, 65.58 ppm against A. stephensi and C. quinquefasciatus being observed after 24 and 48 h of treatment, respectively. The efficacy of petroleum ether was followed by that of the carbon tetrachloride and methanol extracts, which had LC₅₀ values of 268.61, 143.85; 446.61, 384.84 ppm and LC₉₀ values of 641.23, 507.80; 923.60, 887.00 ppm against A. stephensi after 24 and 48 h, respectively, and LC₅₀ values of 24.14, 17.02; 63.48, 53.77 ppm and LC₉₀ values of 295.38, 204.23; 689.71, 388.87 ppm against C. quinquefasciatus after 24 and 48 h of treatment, respectively. These extracts are highly toxic against mosquito larvae from a range of species; therefore, they may be useful for the management of mosquito larvae to control vector borne diseases.     

Effects of Bacillus sphaericus 1593 and 2362 spore/crystal toxin on cultured mosquito cells

An in vitro assay system for the toxin of Bacillus sphaericus strains 1593 and 2362 has been developed utilizing cultured Culex quinquefasciatus cells. The cytotoxic activity of extracts of B. sphaericus strain 1593 did not necessarily correlate with insecticidal activity. Cytotoxicity and larvicidal activity were neutralized by immune rabbit serum prepared against crude toxin extracts as well as by serum prepared against purified toxin from strain 2362. This purified toxin was also found to be cytotoxic. Activation with mosquito larval gut homogenates enhanced cytotoxicity of both 1593 extracts and purified toxin from 2362. The activity of cytotoxic preparations against three mosquito cell lines paralleled the activity of B. sphaericus spores against larvae of these mosquito species. The results suggest the presence of a protoxin and one or more cytotoxic proteins derived from it.     

Water-relation Parameters of Individual Mesophyll Cells of the Crassulacean Acid Metabolism Plant Kalanchoë daigremontiana

Water-relation parameters of leaf mesophyll cells of the CAM plant Kalanchoë daigremontiana have been determined directly in cells of tissue slices using the pressure-probe technique. Turgor pressures measured in cells of the second to fourth layer from the cut surface showed an average of 1.82 ± 0.62 bar (mean ± sd; n = 157 cells). This was lower than expected from measurements of the osmotic pressure of the cell sap. The half-time (T_1/2) for water-flux equilibration of individual cells was 2.5 to 8.8 seconds. This is the fastest T_1/2 found so far for higher-plant cells. The calculated values of the hydraulic conductivity were in the range of 0.20 to 1.6 × 10⁻⁵ centimeters second⁻¹ bar⁻¹, with an average of (0.69 ± 0.46) × 10⁻⁵ centimeters second⁻¹ bar⁻¹ (mean ± sd; n = 8 cells). The T_1/2 values of water exchange of individual cells are consistent with the overall rates of water-flux equilibration measured for tissue slices.The volumetric elastic moduli (∈) of individual cells were in the range 13 to 128 bar for turgor pressures between 0.0 and 3.4 bar; the average ∈ value was 42.4 ± 27.7 bar (mean ± sd; n = 21 cells). This ∈ value is similar to that observed for other higher-plant cells.The water-storage capacity of individual cells, calculated as C_c = V/(∈ + πⁱ) (where V = cell volume and πⁱ = internal osmotic pressure) was 9.1 × 10⁻⁹ cubic centimeters bar⁻¹ per cell, and the capacity for the tissue was 2.2 × 10⁻² cubic centimeters bar⁻¹ gram⁻¹ fresh weight. The significance of the water-relation parameters determined at the cellular level is discussed in terms of the water relations of whole leaves and the high water-use efficiency characteristic of CAM plants.     

Specificity of RSG-1.2 peptide binding to RRE-IIB RNA element of HIV-1 over Rev peptide is mainly enthalpic in origin

Rev is an essential HIV-1 regulatory protein which binds to the Rev responsive element (RRE) present within the env gene of HIV-1 RNA genome. This binding facilitates the transport of the RNA to the cytoplasm, which in turn triggers the switch between viral latency and active viral replication. Essential components of this complex have been localized to a minimal arginine rich Rev peptide and stem IIB region of RRE. A synthetic peptide known as RSG-1.2 binds with high binding affinity and specificity to the RRE-IIB than the Rev peptide, however the thermodynamic basis of this specificity has not yet been addressed. The present study aims to probe the thermodynamic origin of this specificity of RSG-1.2 over Rev Peptide for RRE-IIB. The temperature dependent melting studies show that RSG-1.2 binding stabilizes the RRE structure significantly (ΔT _m = 4.3°C), in contrast to Rev binding. Interestingly the thermodynamic signatures of the binding have also been found to be different for both the peptides. At pH 7.5, RSG-1.2 binds RRE-IIB with a K_a = 16.2±0.6×10⁷ M⁻¹ where enthalpic change ΔH = −13.9±0.1 kcal/mol is the main driving force with limited unfavorable contribution from entropic change TΔS = −2.8±0.1 kcal/mol. A large part of ΔH may be due to specific stacking between U72 and Arg15. In contrast binding of Rev (K_a = 3.1±0.4×10⁷ M⁻¹) is driven mainly by entropy (ΔH = 0 kcal/mol and TΔS = 10.2±0.2 kcal/mol) which arises from major conformational changes in the RNA upon binding.     

Chicory (Cichorium intybus) and wormwood (Artemisia absinthium) extracts exhibit strong larvicidal activity against mosquito vectors of malaria,dengue fever,and filariasis

Vector-borne diseases transmitted by mosquitoes cause globally important diseases such as malaria, dengue fever, and filariasis. The incidence of these diseases can be reduced through mosquito control programs but these control programs currently rely on synthetic insecticides that can impact the environment, and has selected widespread mosquito resistance. Environment friendly and biodegradable natural insecticides discovered in plants offer an alternative approach to mosquito control. Here, we investigated extracts from root or aerial parts of Chicory (Cichorium intybus) and wormwood (Artemisia absinthium) against the early 4th instar larvae of Anopheles stephensi (malaria vector), Aedes aegypti (dengue fever vector), and Culex quinquefasciatus (filariasis vector). The root and aerial parts extracts of A. absinthium and C. intybus at 200, 100, 50, 25 and 12.5?ppm caused significant mortality of the tested mosquito species. Root extracts exhibited higher larvicidal activity that aerial part extracts. The highest larvicidal activity was recorded in methanol extract of roots of C. intybus with LC50?=?66.16, 18.88 and LC¬90?=?197.56, 107.16?ppm for An. stephensi; LC50?=?78.51, 40.15 and LC90?=?277.31, 231.28?ppm for Ae. aegypti and LC50?=?103.99, 64.56 and LC¬90?=?314.04, 247.54?ppm for Cx. quinquefasciatus. These results reveal potent mosquito larvicidal activity against vectors of malaria, dengue fever, and filariasis is present in extracts of chicory and wormwood.     

Characterization of the kinetic and thermodynamic landscape of RNA folding using a novel application of isothermal titration calorimetry

A novel isothermal titration calorimetry (ITC) method was applied to investigate RNA helical packing driven by the GAAA tetraloop–receptor interaction in magnesium and potassium solutions. Both the kinetics and thermodynamics were obtained in individual ITC experiments, and analysis of the kinetic data over a range of temperatures provided Arrhenius activation energies (ΔH^‡) and Eyring transition state entropies (ΔS^‡). The resulting rich dataset reveals strongly contrasting kinetic and thermodynamic profiles for this RNA folding system when stabilized by potassium versus magnesium. In potassium, association is highly exothermic (ΔH_25°C = −41.6 ± 1.2 kcal/mol in 150 mM KCl) and the transition state is enthalpically barrierless (ΔH^‡ = −0.6 ± 0.5). These parameters are sigificantly positively shifted in magnesium (ΔH_25°C = −20.5 ± 2.1 kcal/mol, ΔH^‡ = 7.3 ± 2.2 kcal/mol in 0.5 mM MgCl₂). Mixed salt solutions approximating physiological conditions exhibit an intermediate thermodynamic character. The cation-dependent thermodynamic landscape may reflect either a salt-dependent unbound receptor conformation, or alternatively and more generally, it may reflect a small per-cation enthalpic penalty associated with folding-coupled magnesium uptake.     

Kinetic and Thermodynamic Characterization of Glucoamylase from Colletotrichum sp. KCP1

Extracellular glucoamylase of Colletotrichum sp. KCP1 produced through solid state fermentation was purified by two steps purification process comprising ammonium sulphate precipitation followed by gel permeation chromatography (GPC). The Recovery of glucoamylase after GPC was 50.40 % with 19.3-fold increase in specific activity. The molecular weight of enzyme was found to be 162.18 kDa by native-PAGE and was dimeric protein of two sub-units with molecular weight of 94.62 and 67.60 kDa as determined by SDS-PAGE. Activation energy for starch hydrolysis was 26.45 kJ mol⁻¹ while temperature quotient (Q₁₀) was found to be 1.9. The enzyme was found to be stable over wide pH range and thermally stable at 40–50 °C up to 120 min while exhibited maximum activity at 50 °C with pH 5.0. The pKa₁ and pKa₂ of ionisable groups of active site controlling V_max were 3.5 and 6.8, respectively. V_max, K_m and K_cat for starch hydrolysis were found to be 58.82 U ml⁻¹, 1.17 mg (starch) ml⁻¹ and 449 s⁻¹, respectively. Activation energy for irreversible inactivation (E_a(d)) of glucoamylase was 74.85 kJ mol⁻¹. Thermodynamic parameters of irreversible inactivation of glucoamylase and starch hydrolysis were also determined.     

Isolation and Characterization of Antidermatophytic Bioactive Molecules from Piper longum L. Leaves

Piper longum L. (Piperaceae) commonly known as “long pepper” is a well known medicinal plant in ayurveda. Different parts of this plant, such as root, seed, fruit, whole plant etc. are used traditionally in various ailments. Here we have investigated the antidermatophytic activity of sequentially extracted petroleum ether, chloroform, methanol and water extracts from P. longum leaf against Trichophytonmentagrophytes, T. rubrum, T. tonsurans, Microsporum fulvum and M. gypseum. Better activity of chloroform and methanol extracts was observed. The chloroform extract was selected for further study and the MIC value was recorded as 5.0 mg ml⁻¹ against the test organisms. In the chloroform extract, tannins and phenolic compounds were detected. Further activity-guided fractionation of chloroform extract by silica gel column chromatography yielded nine major fractions. Among these, fraction-1, 4, 5 and 7 showed higher antidermatophytic activity. Fraction-4 on further purification by repeated column chromatography yielded a potential antidermatophytic fraction showing MIC value of 0.625 mg ml⁻¹ against T. mentagrophytes and T. rubrum as determined by broth microdilution method. The major compounds were identified as 1,2-benzenedicarboxylic acid, bis(2-ethylhexyl) ester (C₂₄H₃₈O₄] (41.45 %), 2,2-dimethoxybutane (C₆H₁₄O₂] (13.6 %) and β-myrcene (C₁₀H₁₆) (6.75 %) based on GC–MS data.     

Enhancement of the rate of pyrophosphate hydrolysis by nonenzymatic catalysts and by inorganic pyrophosphatase

To estimate the proficiency of inorganic pyrophosphatase as a catalyst, ³¹P NMR was used to determine rate constants and thermodynamics of activation for the spontaneous hydrolysis of inorganic pyrophosphate (PP_i) in the presence and absence of Mg²⁺ at elevated temperatures. These values were compared with rate constants and activation parameters determined for the reaction catalyzed by Escherichia coli inorganic pyrophosphatase using isothermal titration calorimetry. At 25 °C and pH 8.5, the hydrolysis of MgPP_i²⁻ proceeds with a rate constant of 2.8 × 10⁻¹⁰ s⁻¹, whereas E. coli pyrophosphatase was found to have a turnover number of 570 s⁻¹ under the same conditions. The resulting rate enhancement (2 × 10¹²-fold) is achieved entirely by reducing the enthalpy of activation (ΔΔH^‡ = −16.6 kcal/mol). The presence of Mg²⁺ ions or the transfer of the substrate from bulk water to dimethyl sulfoxide was found to increase the rate of pyrophosphate hydrolysis by as much as ∼10⁶-fold. Transfer to dimethyl sulfoxide accelerated PP_i hydrolysis by reducing the enthalpy of activation. Mg²⁺ increased the rate of PP_i hydrolysis by both increasing the entropy of activation and reducing the enthalpy of activation.