Ultrasound mediated efficient synthesis of new 4-oxoquinazolin-3(4H)-yl)furan-2-carboxamides as potent tyrosinase inhibitors: Mechanistic approach through chemoinformatics and molecular docking studies

We have carried out the synthesis of new 4-oxoquinazolin-3(4H)-yl)furan-2-carboxamide derivatives by the reaction between isatoic anhydride, 2-furoic hydrazide and substituted salicylaldehydes in ethanol: water (5:5 v/v) solvent system using p-TSA as a catalyst under ultrasound irradiation at room temperature. The structures of newly synthesized compounds were confirmed through spectral techniques such as IR, ¹H NMR, ¹³C NMR, and LCMS. The important features of this protocol include simple and easy workup procedure, reaction carried out at ambient temperature, use of ultrasound and high yield of oxoquinazolin-3(4H)-yl)furan-2-carboxamides in short reaction time. The synthesized compounds 4a–4j were screened against tyrosinase enzyme and all these compounds found to be potent inhibitors with much lower IC₅₀ value of 0.028 ± 0.016 to 1.775 ± 0.947 µM than the standard kojic acid (16.832 ± 1.162 µM). The kinetics mechanism for compound 4e was analyzed by Lineweaver-Burk plots which revealed that compound inhibited tyrosinase non-competitively by forming an enzyme-inhibitor complex. Along with this all the synthesized compounds (4a–4j) were scanned for their DPPH free radical scavenging ability. The outputs received through in vitro and in silico analysis are coherent to the each other with good binding energy values (kcal/mol) posed by synthesized ligands.     

Morphological specialization of heterantherous Rhynchanthera grandiflora (Melastomataceae) accommodates pollinator diversity

• The tropical Melastomataceae are characterized by poricidal anthers which constitute a floral filter selecting for buzz‐pollinating bees. Stamens are often dimorphic, sometimes with discernible feeding and pollinating functions. Rhynchanthera grandiflora produces nectarless flowers with four short stamens and one long stamen; all anthers feature a narrow elongation with an upwards facing pore.
• We tested pollen transfer by diverse foraging bees and viability of pollen from both stamen types. The impact of anther morphology on pollen release direction and scattering angle was studied to determine the plant's reproductive strategy.
• Medium‐sized to large bees sonicated flowers in a specific position, and the probability of pollen transfer correlated with bee size even among these legitimate visitors. Small bees acted as pollen thieves or robbers. Anther rostrum and pore morphology serve to direct and focus the pollen jet released by floral sonication towards the pollinator's body. Resulting from the ventral and dorsal positioning of the short and long stamens, respectively, the pollinator's body was widely covered with pollen. This improves the plant's chances of outcrossing, irrespective of which bee body part contacts the stigma. Consequently, R. grandiflora is also able to employ bee species of various sizes as pollen vectors.
• The strategy of spreading pollen all over the pollinator's body is rather cost‐intensive but counterbalanced by ensuring that most of the released pollen is in fact transferred to the bee. Thus, flowers of R. grandiflora illustrate how specialized morphology may serve to improve pollination by a functional group of pollinators.

     

Stain Solubilities Part III

Ethylenediaminetetraacetic acid (EDTA) solution is used to decalcify bone specimens for histological examination. Sodium hydroxide (NaOH) has been used to dissolve EDTA and to bring EDTA solutions to neutral pH. This solution, however, requires several weeks to decalcify bone specimens. We investigated a new de-calcification fluid using concentrated ammonium hydroxide (NH₄OH) to dissolve EDTA and to adjust the pH to neutral. Decalcification was performed using a magnetic stirrer with and without vacuum, or with a sonic cleaner. Decalcification end point was confirmed using both the weight loss and X-ray methods. After decalcification, specimens were processed through paraffin and sections were stained with hematoxylin and eosin. Decalcification employing NH₄OH required an average of six days. Light microscopy indicated good retention of cellular detail.     

A benign approach to the preparation of freshwater bryozoan statoblasts for scanning electron microscopy (SEM) imaging

Abstract

Several different species of freshwater Bryozoa, belonging to the genera Plumatella, Rumarcanella and Fredericella, were detected within the Northern Mallee Pipeline (NMP) system in Victoria, Australia, that required definitive identification. These organisms produce asexual buds called statoblasts, with valves composed of sclerotised chitin that bear minute micro-ornamentations of considerable taxonomical significance. Imaging and analysis of these distinctive micro-ornamentations using scanning electron microscopy (SEM) is often employed for species identification. Meticulous preparation of statoblast samples is therefore required that necessitates the removal of adhering debris, dehydration and drying—whilst mitigating specimen damage and distortion. This technical note describes an approach whereby each of these three steps have been individually designed to be as benign as possible, using mild detergent/sonication to remove debris, a gradual and gentle dehydration procedure using ethanol, and critical point drying. For the overall process, these methods are chosen to optimise control and to minimise the use of harsh and hazardous chemicals.     

Purification and properties of ATPase inhibitor from rat liver mitochondria

(1) The ATPase inhibitor protein has been isolated from rat liver mitochondria in purified form. The molecular weight determined by sodium dodecyl sulfate gel electrophoresis is approximately 9500, and the isoelectric point is 8.9.

(2) The protein inhibits both the soluble ATPase and the particle-bound ATPase from rat liver mitochondria. It also inhibits ATPase activities of soluble F₁, and inhibitor-depleted submitochondrial particles derived from bovine heart mitochondria.

(3) On particle-bound ATPase the inhibitor has its maximal effect if incubated in the presence of Mg²⁺. ATP at slightly acidic pH.

(4) The inhibitor has a minimal effect on P_i-ATP exchange activity in sonicated submitochondrial particles. However, unexpectedly the inhibitor greatly stimulates P_i-ATP exchange activity in whole mitochondria while the low ATPase activity of the mitochondria is not affected. The possible mechanism of action of the inhibitor on intact mitochondria is offered.     

The interaction between the mitochondrial ATPase (F1) and the ATPase inhibitor

1. The naturally occurring mitochondrial ATPase inhibitor inhibits the mitochondrial ATPase (F₁) non-competitively.2. The interaction between inhibitor and inhibitor-depleted F₁ or submitochondrial particles is diminished when the ratio of ATP/ADP is low or when energy is generated by substrate oxidation.3. The dissociation of the inhibitor from coupled Mg-ATP particles is promoted when substrates are being oxidized. This results in the appearance of a large uncoupler-stimulated ATPase activity. Activation of the uncoupler-stimulated ATPase activity is also achieved by incubation of the particles with ADP.4. The ATPase activity of Mg-ATP particles is determined by the turnover capacity of F₁. When endogenous inhibitor is removed, energy dissipation becomes the rate-limiting step. This energy dissipation can be activated by an uncoupler.5. Evidence is presented for the existence of a non-inhibited intermediate F₁-inhibitor complex.     

Optimisation of conditions for the extraction of casearins from Casearia sylvestris using response surface methodology

Optimal conditions for the extraction of casearins from Casearia sylvestris were determined using response surface methodology. The maceration and sonication extraction techniques were performed using a 3 x 3 x 3 full factorial design including three acidity conditions, three solvents of different polarities and three extraction times. The yields and selectivities of the extraction of casearins were significantly influenced by acidity conditions. Taking into account all variables tested, the optimal conditions for maceration extraction were estimated to involve treatment with dichloromethane saturated with ammonium hydroxide for 26 h. Similar yields and selectivities for casearins were determined for sonication extraction using the same solvent but for the much shorter time of 1 h. The best results for stabilisation of the fresh plant material were obtained using leaves that had been oven dried at 40 degrees C for 48 h.     

Sonication: A new method for gene transfer to plants

Sonication is a novel method for gene transfer into plant protoplasts and intact plant cells. The mode of action of ultrasound and its chemical, biochemical and physiological effects are reviewed. The state of the art of acoustic transformation is presented and possible mechanisms are discussed.     

Gel filtration of egg phosphatidylcholine vesicles

The abilities of Sepharose 2b (Pharmacia), Controlled Pore Glass (Electro-Nucleonics) and Bio-Gel A150m (Bio-Rad) to purify small unilamellar vesicles prepared by sonication and the ethanol-injection methods were compared. The Bio-Gel causes complete aggregation of the sonicated vesicles and partial aggregation of the ethanol-injection vesicles. Both Sepharose and Controlled Pore Glass are acceptable for purifying vesicles from multilamellar liposomes; however, neither will separate the vesicles from sonication by-products which might be formed.     

Characterization of a non-detergent PS II-cytochrome b/f preparation (BS)

A non-detergent photosystem II preparation, named BS, has been characterized by countercurrent distribution, light saturation curves, absorption spectra and fluorescence at room and at low temperature (–196°C). The BS fraction is prepared by a sonication-phase partitioning procedure (Svensson P and Albertsson P-Å, Photosynth Res 20: 249–259, 1989) which removes the stroma lamellae and the margins from the grana and leaves the appressed partition region intact in the form of vesicles. These are closed structures of inside-out conformation. They have a chlorophyll a/b ratio of 1.8–2.0, have a high oxygen evolving capacity (295 mol O₂ per mg chl h), are depleted in P700 and enriched in the cytochrome b/f complex. They have about 2 Photosystem II reaction centers per 1 cytochrome b/f complex.The plastoquinone pool available for PS II in the BS vesicles is 6–7 quinones per reaction center, about the same as for the whole thylakoid. It is concluded, therefore, that the plastoquinone of the stroma lamellae is not available to the PS II in the grana and that plastoquinone does not act as a long range electron transport shuttler between the grana and stroma lamellae.Compared with Photosystem II particles prepared by detergent (Triton X-100) treatment, the BS vesicles retain more cytochrome b/f complex and are more homogenous in their surface properties, as revealed by countercurrent distribution, and they have a more efficient energy transfer from the antenna pigments to the reaction center.Abbreviations DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - F_v variable fluorescence - LHC light-harvesting complex - PpBQ phenyl-p-benzoquinone - PQ plastoquinone pool - P700 reaction center of PS I - PS I, PS II Photosystem I, II - Q_A first bound plastoquinone accepter - RC reaction centre