Periodic emission of droplets from an oscillating electrified meniscus of a low-viscosity, highly conductive liquid

The generation of identical droplets of controllable size in the micrometer range is a problem of much interest owing to the numerous technological applications of such droplets. This work reports an investigation of the regime of periodic emission of droplets from an electrified oscillating meniscus of a liquid of low viscosity and high electrical conductivity attached to the end of a capillary tube, which may be used to produce droplets more than ten times smaller than the diameter of the tube. To attain this periodic microdripping regime, termed axial spray mode II by Juraschek and Röllgen [R. Juraschek and F. W. Röllgen, Int. J. Mass Spectrom. 177, 1 (1998)], liquid is continuously supplied through the tube at a given constant flow rate, while a dc voltage is applied between the tube and a nearby counter electrode. The resulting electric field induces a stress at the surface of the liquid that stretches the meniscus until, in certain ranges of voltage and flow rate, it develops a ligament that eventually detaches, forming a single droplet, in a process that repeats itself periodically. While it is being stretched, the ligament develops a conical tip that emits ultrafine droplets, but the total mass emitted is practically contained in the main droplet. In the parametrical domain studied, we find that the process depends on two main dimensionless parameters, the flow rate nondimensionalized with the diameter of the tube and the capillary time, q, and the electric Bond number BE, which is a nondimensional measure of the square of the applied voltage. The meniscus oscillation frequency made nondimensional with the capillary time, f, is of order unity for very small flow rates and tends to decrease as the inverse of the square root of q for larger values of this parameter. The product of the meniscus mean volume times the oscillation frequency is nearly constant. The characteristic length and width of the liquid ligament immediately before its detachment approximately scale as powers of the flow rate and depend only weakly on the applied voltage. The diameter of the main droplets nondimensionalized with the diameter of the tube satisfies dd≈(6/π)1/3(q/f)1/3, from mass conservation, while the electric charge of these droplets is about 1/4 of the Rayleigh charge. At the minimum flow rate compatible with the periodic regimen, the dimensionless diameter of the droplets is smaller than one-tenth, which presents a way to use electrohydrodynamic atomization to generate droplets of highly conducting liquids in the micron-size range, in marked contrast with the cone-jet electrospray whose typical droplet size is in the nanometric regime for these liquids. In contrast with other microdripping regimes where the mass is emitted upon the periodic formation of a narrow capillary jet, the present regime gives one single droplet per oscillation, except for the almost massless fine aerosol emitted in the form of an electrospray.

Harnessing bacterial signals for suppression of biofilm formation in the nosocomial fungal pathogen Aspergillus fumigatus

Faced with the continued emergence of antibiotic resistance to all known classes of antibiotics, a paradigm shift in approaches toward antifungal therapeutics is required. Well characterized in a broad spectrum of bacterial and fungal pathogens, biofilms are a key factor in limiting the effectiveness of conventional antibiotics. Therefore, therapeutics such as small molecules that prevent or disrupt biofilm formation would render pathogens susceptible to clearance by existing drugs. This is the first report describing the effect of the Pseudomonas aeruginosa alkylhydroxyquinolone interkingdom signal molecules 2-heptyl-3-hydroxy-4-quinolone and 2-heptyl-4-quinolone on biofilm formation in the important fungal pathogen Aspergillus fumigatus. Decoration of the anthranilate ring on the quinolone framework resulted in significant changes in the capacity of these chemical messages to suppress biofilm formation. Addition of methoxy or methyl groups at the C5–C7 positions led to retention of anti-biofilm activity, in some cases dependent on the alkyl chain length at position C2. In contrast, halogenation at either the C3 or C6 positions led to loss of activity, with one notable exception. Microscopic staining provided key insights into the structural impact of the parent and modified molecules, identifying lead compounds for further development.

Physicochemical complexity in complex chemical systems

Self-replication and compartmentalization are two central properties thought to be essential for minimal life, and understanding how such processes interact in the emergence of complex reaction networks is crucial to exploring the development of complexity in chemistry and biology. Autocatalysis can emerge from multiple different mechanisms such as formation of an initiator, template self-replication and physical autocatalysis (where micelles formed from the reaction product solubilize the reactants, leading to higher local concentrations and therefore higher rates). Amphiphiles are also used in artificial life studies to create protocell models such as micelles, vesicles and oil-in-water droplets, and can increase reaction rates by encapsulation of reactants. So far, no template self-replicator exists which is capable of compartmentalization, or transferring this molecular scale phenomenon to micro or macro-scale assemblies. Here a system is demonstrated where an amphiphilic imine catalyses its own formation by joining a non-polar alkyl tail group with a polar carboxylic acid head group to form a template, which was shown to form reverse micelles by Dynamic Light Scattering (DLS). The kinetics of this system were investigated by 1H NMR spectroscopy, showing clearly that a template self-replication mechanism operates, though there was no evidence that the reverse micelles participated in physical autocatalysis. Active oil droplets, composed from a mixture of insoluble organic compounds in an aqueous sub-phase, can undergo processes such as division, self-propulsion and chemotaxis, and are studied as models for minimal cells, or protocells. Although in most cases the Marangoni effect is responsible for the forces on the droplet, the behaviour of the droplet depends heavily on the exact composition. Though theoretical models are able to calculate the forces on a droplet, to model a mixture of oils on an aqueous surface where compounds from the oil phase are dissolving and diffusing through the aqueous phase is beyond current computational capability. The behaviour of a droplet in an aqueous phase can only be discovered through experiment, though it is determined by the droplet's composition. By using an evolutionary algorithm and a liquid handling robot to conduct droplet experiments and decide which compositions to test next, entirely autonomously, the composition of the droplet becomes a chemical genome capable of evolution. The selection is carried out according to a fitness function, which ranks the formulation based on how well it conforms to the chosen fitness criteria (e.g. movement or division). Over successive generations, significant increases in fitness are achieved, and this increase is higher with more components (i.e. greater complexity). Other chemical processes such as chemiluminescence and gelation were investigated in active oil droplets, demonstrating the possibility of controlling chemical reactions by selective droplet fusion. Potential future applications for this might include combinatorial chemistry, or additional fitness goals for the genetic algorithm. Combining the self-replication and the droplet protocells research, it was demonstrated that the presence of the amphiphilic replicator lowers the interfacial tension between droplets of a reaction mixture in organic solution and the alkaline aqueous phase, causing them to divide. Periodic sampling by a liquid handling robot revealed that the extent of droplet fission increased as the reaction progressed, producing more individual protocells with increased self-replication. This demonstrates coupling of the molecular scale phenomenon of template self-replication to a macroscale physicochemical effect.

A micro-extraction technique using a new digitally controlled syringe combined with UHPLC for assessment of urinary biomarkers of oxidatively damaged DNA

The formation of reactive oxygen species (ROS) within cells causes damage to biomolecules, including membrane lipids, DNA, proteins and sugars. An important type of oxidative damage is DNA base hydroxylation which leads to the formation of 8-oxo-7,8-dihydro-29-deoxyguanosine (8-oxodG) and 5-hydroxymethyluracil (5-HMUra). Measurement of these biomarkers in urine is challenging, due to the low levels of the analytes and the matrix complexity. In order to simultaneously quantify 8-oxodG and 5-HMUra in human urine, a new, reliable and powerful strategy was optimised and validated. It is based on a semi-automatic microextraction by packed sorbent (MEPS) technique, using a new digitally controlled syringe (eVolH), to enhance the extraction efficiency of the target metabolites, followed by a fast and sensitive ultrahigh pressure liquid chromatography (UHPLC). The optimal methodological conditions involve loading of 250 mL urine sample (1:10 dilution) through a C8 sorbent in a MEPS syringe placed in the semi-automatic eVolH syringe followed by elution using 90 mL of 20% methanol in 0.01% formic acid solution. The obtained extract is directly analysed in the UHPLC system using a binary mobile phase composed of aqueous 0.1% formic acid and methanol in the isocratic elution mode (3.5 min total analysis time). The method was validated in terms of selectivity, linearity, limit of detection (LOD), limit of quantification (LOQ), extraction yield, accuracy, precision and matrix effect. Satisfactory results were obtained in terms of linearity (r2 . 0.991) within the established concentration range. The LOD varied from 0.00005 to 0.04 mg mL21 and the LOQ from 0.00023 to 0.13 mg mL21. The extraction yields were between 80.1 and 82.2 %, while inter-day precision (n=3 days) varied between 4.9 and 7.7 % and intra-day precision between 1.0 and 8.3 %. This approach presents as main advantages the ability to easily collect and store urine samples for further processing and the high sensitivity, reproducibility, and robustness of eVolHMEPS combined with UHPLC analysis, thus retrieving a fast and reliable assessment of oxidatively damaged DNA.

Tracking biochemical changes during adventitious root formation in olive (Olea europaea L.)

The activity of oxidative enzymes and the levels of free auxins were determined during adventitious root formation in olive explants. Rooting trials were performed both with in vitro-cultured micro shoots of the cultivar ‘Galega Vulgar’, treated with indole-3-butyric acid (IBA) and with salicylhydroxamic acid(SHAM) + IBA, as well as with semi-hardwood cuttings of the cultivars ‘Galega Vulgar’ (difficult-to-root)and ‘Cobrançosa’ (easy-to-root), treated with IBA. The auxin (IBA) was used in all experiments as a rooting promoter, while SHAM was used in micropropagation trials as rooting inhibitor, providing a negative control. Free indole-3-acetic acid (IAA) and IBA concentrations were determined in microshoots, as well as in semi-hardwood cuttings, throughout the rooting period at pre-established time-points. At the sametime-points, the enzymatic activity of polyphenol oxidases (PPO), peroxidases (POX), and IAA oxidase(IAAox) was evaluated in the microshoots. Microshoots treated with SHAM + IBA revealed higher POX and IAAox activity, as well as lower PPO activity, than those treated only with IBA. IAA levels were higher in IBA-treated microshoots during induction phase, but lower during early initiation phase. Incontrast, free IBA levels were higher in microshoots treated with SHAM + IBA during induction, but lower during initiation. A similar pattern of free auxin levels was observed in semi-hardwood cuttings of the two contrasting cultivars under evaluation. The similarities found on the auxin patterns of microshoots treated with SHAM and those of semi-hardwood cuttings of the difficult-to-root olive cultivar allow considering SHAM a reliable control for when simulation of a difficult-to-root behavior is necessary. The inhibitory effect of SHAM in root formation could be related with 1) the inhibition of alternative oxidase(AOX), leading to a down regulation of phenylpropanoid biosynthetic pathways, which would decrease the concentration of phenolic substrates for PPO; 2) an increase in IAAox activity resulting in lower free IAA levels or; 3) a defective conversion of IBA into IAA.