In vitro antibiofilm and anti‐adhesion effects of magnesium oxide nanoparticles against antibiotic resistant bacteria

The aim of the current investigation was to determine the antibacterial and antibiofilm potential of MgO nanoparticles (NPs) against antibiotic‐resistant clinical strains of bacteria. MgO NPs were synthesized by a wet chemical method and further characterized by scanning electron microscopy and energy dispersive X‐ray. Antibacterial activity was determined by broth microdilution and agar diffusion methods. The Bradford method was used to assess cellular protein leakage as a result of loss of membrane integrity. Microtiter plate assay following crystal violet staining was employed to determine the effect of MgO NPs on biofilm formation and removal of established biofilms. MIC values ranged between 125 and 500 μg/mL. Moreover, treatment with MgO NPs accelerated rate of membrane disruption, measured as a function of leakage of cellular proteins. Leakage of cellular protein content was greater among gram‐negative bacteria. Cell adherence assay indicated 25.3–49.8% inhibition of bacterial attachment to plastic surfaces. According to a static biofilm method, MgO NPs reduced biofilm formation potential from 31% to 82.9% in a time‐dependent manner. Moreover, NPs also significantly reduced the biomass of 48, 72, 96 and 120 hr old biofilms (P < 0.05). Cytotoxicity experiments using a neutral red assay revealed that MgO NPs are non‐toxic to HeLa cells at concentrations of 15–120 μg/mL. These data provide in vitro scientific evidence that MgO NPs are effective and safe antibiofilm agents that inhibit adhesion, biofilm formation and removal of established biofilms of multidrug‐resistant bacteria.

     

Industrial wastewater treatment in internal circulation bioreactor followed by wetlands containing emergent plants and algae

Wastewater treatment based on ecological principles is a low cost and highly desirable solution for the developing countries like Pakistan. The present study evaluated the effectiveness of biological treatment systems including Internal Circulation (IC) anaerobic bioreactor and constructed wetlands (CWs) containing macrophytes and mixed algal cultures for industrial wastewater treatment. The IC bioreactor reduced COD (52%), turbidity (89%), EC (24%) of the industrial wastewater. However, the effluents of IC bioreactor did not comply with National Environmental Quality Standards (NEQS) of Pakistan. Post-treatment of IC bioreactor effluents was accomplished in CW containing macrophytes (Arundo donax and Eichhornia crassipes) and mixed algal culture. The CWs planted with macrophytes lowered the concentrations of COD (89%) and turbidity (99%). CWs with algal biomass were not effective in further polishing the effluent. Inhibition of algal biomass growth was observed due to physicochemical characteristics of wastewater. The integrated treatment system consisting of IC bioreactor and macrophytes was found more suitable option for industrial wastewater treatment.     

Physiological and biochemical mechanisms of silicon-induced copper stress tolerance in cotton (<Emphasis Type="Italic">Gossypium hirsutum</Emphasis> L.)

Accumulation of excess copper (Cu) in agricultural soils can decrease growth and quality of crops grown on these soils and a little information is available on the role of silicon (Si) in reducing Cu toxicity in plants. A hydroponic study was conducted to investigate the effects of Si (1.0 mM) on growth and physiology of cotton seedlings grown on different Cu (0, 25, and 50 µM) concentrations. Elevated levels of Cu decreased growth, biomass, photosynthetic pigments, and gas exchange characteristics, and increased the electrolyte leakage (EL), hydrogen peroxide (H₂O₂), and thiobarbituric acid reactive substances (TBARS) contents in leaf, stem, and roots of cotton seedlings. Cu stress alone decreased the activities of key antioxidant enzymes in cotton seedlings. Exogenous application of Si alleviated the toxic effects of Cu on cotton seedlings by improving growth, photosynthetic pigments, and gas exchange characteristics under Cu stress. The Si application decreased Cu concentrations in leaves, stem, and roots as compared with the control plants. Furthermore, Si decreased oxidative stress as evidenced by decreased EL, H₂O₂, and TBARS contents, and increased the antioxidant enzyme activities in cotton seedlings. This study provides evidences of Si-mediated reduction of Cu toxicity in cotton seedlings at physiological and biochemical levels.     

Theoretical insights into thermal cyclophanediene to dihydropyrene electrocyclic reactions; a comparative study of Woodward Hoffmann allowed and forbidden reactions

The thermally allowed electrocyclic reaction syn-cyclophanediene (CPD) to dihydropyrene (DHP) was compared with the disallowed thermal electrocyclic reaction in anti CPD through density functional theory (DFT) calculations at the B3LYP/6-31?+?G(d) level. Moreover, the results were also compared with the electrocyclization of 1,3,5 hexatriene to 1,3-cyclohexadiene . The Woodward-Hoffmann (W-H) allowed thermal reaction in syn CPD 11 has a calculated activation barrier of 6.23 kcal mol^?1, compared with 29 kcal mol^?1 for the electrocyclization of 1,3,5 hexatriene to 1,3-cyclohexadiene. The enhanced acceleration of electrocyclization is believed to arise from geometrically enforced spatially aligned termini of the hexatriene. Substituents at the electrocyclic terminus of cyclophanediene significantly affected (up to three fold) the activation barriers. Mono-substitution of CPD has substituent dependent acceleration or deceleration whereas di-substitution always increased the activation barrier. The activation barrier for electrocyclization in 33 is 4.44 kcal mol^?1, which is the lowest activation barrier for any thermal electrocyclic reaction. Cyclophanedienes (CPDs) substituted with electron-rich substituents cyclized with high activation barriers and vice versa, a phenomenon significantly different from electrocyclic reaction of 1,3,5-hexatriene where no such trend is traceable. Comparison of W-H allowed and forbidden electrocyclization in syn and anti CPDs, respectively, revealed quite similar electronic demand, although the transition states are different in nature. The transition state for a W-H forbidden reaction is biradicaloid, with most of the spin density at the electrocyclic termini; however, the transition state for a W-H allowed reaction has no such contribution. We also believe that this is the first study of its type, where W-H allowed and forbidden reactions are compared on a similar set of molecules, and compared for electronic effect through substituents.     

A Comprehensive Spectroscopic and Computational Investigation to Probe the Interaction of Antineoplastic Drug Nordihydroguaiaretic Acid with Serum Albumins

Exogenous drugs that are used as antidote against chemotheray, inflammation or viral infection, gets absorbed and interacts reversibly to the major serum transport protein i.e. albumins, upon entering the circulatory system. To have a structural guideline in the rational drug designing and in the synthesis of drugs with greater efficacy, the binding mechanism of an antineoplastic and anti-inflammatory drug Nordihydroguaiaretic acid (NDGA) with human and bovine serum albumins (HSA & BSA) were examined by spectroscopic and computational methods. NDGA binds to site II of HSA with binding constant (K_b) ~10⁵ M^-1 and free energy (ΔG) ~ -7.5 kcal.mol^-1. It also binds at site II of BSA but with lesser binding affinity (K_b) ~10⁵ M^-1 and ΔG ~ -6.5 kcal.mol^-1. The negative value of ΔG, ΔH and ΔS for both the albumins at three different temperatures confirmed that the complex formation process between albumins and NDGA is spontaneous and exothermic. Furthermore, hydrogen bonds and hydrophobic interactions are the main forces involved in complex formation of NDGA with both the albumins as evaluated from fluorescence and molecular docking results. Binding of NDGA to both the albumins alter the conformation and causes minor change in the secondary structure of proteins as indicated by the CD spectra.     

Inactivation of (Na+ + K+)-stimulated ATPase by a cytotoxic protein from cobra venom in relation to its lytic effects on cells

The mechanism of action of the cytotoxic protein P₆ isolated from cobra venom (Naja naja) which shows preferential cytotoxicity particularly to Yoshida sarcoma cells has been studied by its effects on the membrane-bound enzyme (Na⁺ + K⁺)-ATPase (ATP phosphohydrolase, EC 3.6.1.3) of a variety of cell systems. Evidence obtained with Yoshida sarcoma cells, dog and human erythrocytes and three tissue culture cell lines KB (human oral carcinoma), Hela (human cervix carcinoma) and L-132 (human lung embryonic) shows that inhibition of (Na⁺ + K⁺)-ATPase by the P₆ protein can be correlated with its lytic activity. (Na⁺ + K⁺)-ATPase of Yoshida sarcoma membrane fragments inactivated by P₆ protein could be reconstituted by the addition of phosphatidylserine and phosphatidic acid. It is conceivable that lysis of cells by the P₆ protein may be due to an imbalance of K⁺ and Na⁺ in the cell which leads to swelling and disintegration of the membrane structure. Observations indicate that the P₆ protein combines with membrane constituents of susceptible cells. The overall evidence suggests that both the specificity of its protein structure and the highly basic nature of the P₆ protein are factors which enable it to compete with the lipid moiety maintaining the (Na⁺ + K⁺)-ATPase of the susceptible cells in proper conformation for activity.     

METABOLIC PARAMETERS OF ONTOGENESIS OF ELECTRICAL ACTIVITY IN THE BRAIN. SODIUM-POTASSIUM ACTIVATED ADENOSINE TRIPHOSPHATASE IN DEVELOPING CHICK EMBRYO BRAIN

—(1) The activity of the Na-K ATPase in the particulate fraction of the chick embryo brain has been assayed at different stages of development with the objective of finding whether or not changes in the activity of this enzyme bear any relation to the maturation of spontaneous and evoked electrical activity of the growing chick brain. (2) The specific activity of the enzyme is low on day 6 and it rises rapidly between days 10 and 12, at which time it attains a plateau and remains essentially unchanged from day 12 until day 20. Experimental evidence rules out the possible presence of an inhibitor of the enzyme in 8-day-old brain homogenates, suggesting that these developmental changes in the activity of the enzyme may represent new synthesis of enzyme rather than its activation. The period between days 10 and 12 does not represent a unique stage of general protein synthesis. (3) The chick brain particulate enzyme has an optimum activity at pH 7·4 and at 37°. It is optimally activated by a Na⁺ concentration of 100mm and K⁺ concentration of 20 mm . The enzyme is inhibited by ouabain and Ca²⁺. (4) The results have been discussed.     

The Chemical Fractionation of Rabbit and Swine Thymus

A chemical fractionation procedure, previously found applicable to bovine thymus and bovine and ovine palatine tonsils, was used to fractionate rabbit and hog thymus. With respect to the chemical fractionation steps, yields of fractions, and optical and electrophoretic properties, extracts from hog and rabbit thymus were indistinguishable from similar extracts prepared from calf thymus. The study provides composition and yield data applicable to the thymus of a small mammal readily available in most laboratories.     

Surfactant-mediated amyloidogenesis behavior of stem bromelain; a biophysical insight

Neurodegenerative disorders are mainly associated with amyloid fibril formation of different proteins. Stem bromelain (SB), a cysteine protease, is known to exist as a molten globule state at pH 10.0. It passes through the identical surrounding (pH 10.0) in the gut epithelium of intestine upon oral administration. Protein–surfactant complexes are widely employed as drug carriers, so the nature of surfactant toward protein is of great interest. The present work describes the effect of cationic surfactants (CTAB & DTAB) and their hydrophobic behavior toward amyloidogenesis behavior of SB at pH 10.0. Multiple approaches including light scattering, far UV-CD, turbidity measurements, and dye binding assay (ThT, Congo red and ANS) were performed to measure the aggregation propensity of SB. Further, we monitored the hydrodynamic radii of aggregates formed using dynamic light scattering technique. Structure of fibrils was also visualized through fluorescence microscopy as well as TEM. At pH 10.0, low concentration of CTAB (0–200 μM) induced amyloid formation in SB as evident from a prominent increase in turbidity and light scattering, gain in β-sheet content, and enhanced ThT fluorescence intensity. However, further increase in CTAB concentration suppressed the fibrillation phenomenon. In contrast, DTAB did not induce fibril formation at any concentration used (0–500 μM) due to lower hydrophobicity. Net negative charge developed on protein at high pH (10.0) might have facilitated amyloid formation at low concentration of cationic surfactant (CTAB) due to electrostatic and hydrophobic interactions.