Oxidative Skin Research Articles

Targets of Red Grapes: Oxidative Damage of DNA and Leukaemia Cells

Vitis vinifera is a widespread crop all over the world. The biophenols present in grapes have a remarkable influence on wine quality and also confer potential health-protecting properties to this fruit. The aim of the present work was to assess the beneficial properties of skin, seeds and pulp of red table grapes (RTG) (Vitis vinifera, Palieri Cadiz variety). Two potential beneficial activities of red table grapes (RTG) were analyzed: (i) The maintenance of genomic stability studying their genotoxic/antigenotoxic effects, and (ii) the in vitro cytotoxic effect against tumor cells of RTG components. The genotoxic and/or antigenotoxic effect was measured applying the somatic mutation and recombination test on wing imaginal discs of Drosophila melanogaster. The cytotoxic effect was monitored using the HL60 human leukemia model to evaluate the antiproliferative potential of the different parts of RTG. The three major parts (skin, seeds and pulp) are not genotoxic. When antigenotoxicity assays were performed using hydrogen peroxide as the oxidative genotoxin, skin, seed and pulp exerted a desmutagenic effect, with seeds and skin showing the most potent effect. The cytotoxicity tests using HL60 cells indicated that only skin and pulp fractions are able to inhibit the tumor growth, with skin having the lowest IC50 (1.8 mg/mL versus 8 mg/mL of pulp). These results suggest that RTG are potent anti-mutagens that protect DNA from oxidative damage as well as being cytotoxic toward the HL60 tumor cell line.

Inherently aligned microfluidic electrodes composed of liquid metal

This paper describes the fabrication and characterization of microelectrodes that are inherently aligned with microfluidic channels and in direct contact with the fluid in the channels. Injecting low melting point alloys, such as eutectic gallium indium (EGaIn), into microchannels at room temperature (or just above room temperature) offers a simple way to fabricate microelectrodes. The channels that define the shape and position of the microelectrodes are fabricated simultaneously with other microfluidic channels (i.e., those used to manipulate fluids) in a single step; consequently, all of the components are inherently aligned. In contrast, conventional techniques require multiple fabrication steps and registration (i.e., alignment of the electrodes with the microfluidic channels), which are technically challenging. The distinguishing characteristic of this work is that the electrodes are in direct contact with the fluid in the microfluidic channel, which is useful for a number of applications such as electrophoresis. Periodic posts between the microelectrodes and the microfluidic channel prevent the liquid metal from entering the microfluidic channel during injection. A thin oxide skin that forms rapidly and spontaneously on the surface of the metal stabilizes mechanically the otherwise low viscosity, high surface tension fluid within the channel. Moreover, the injected electrodes vertically span the sidewalls of the channel, which allows for the application of uniform electric field lines throughout the height of the channel and perpendicular to the direction of flow. The electrodes are mechanically stable over operating conditions commonly used in microfluidic applications; the mechanical stability depends on the magnitude of the applied bias, the nature of the bias (DC vs. AC), and the conductivity of the solutions in the microfluidic channel. Electrodes formed using alloys with melting points above room temperature ensure mechanical stability over all of the conditions explored. As a demonstration of their utility, the fluidic electrodes are used for electrohydrodynamic mixing, which requires extremely high electric fields (~10(5) V m(-1)).

A Study of the Method of Manufacturing Bimaterial Composite Parts through Semisolid Metal Processing

This work evaluated the method of manufacturing bimaterial composite parts by semisolid metal processing (SSP) through strain-induced melt-activated thixo-forging. Sn-15 pct Pb and Pb-30 pct Sn semisolid alloys were chosen as model alloys. Bimetal composite parts were manufactured successfully by forging the semisolid alloys into the same die simultaneously. Optical photography, scanning electron microscopy, energy dispersive X-ray spectroscopy, and Vickers hardness were employed to characterize the samples. The results showed that the composite semisolid fluid flowed in a laminar way. Globular primary grains in the two semisolid alloys maintained their respective geometry and constitutions. The mixture of two liquid phases was limited in a thin layer beneath the interface between the two semisolid alloys. The absence of an oxide-enriched layer at the interface suggests that the oxide skins of the feedstock were torn during the processing, leading to the formation of metallurgical bonding at the composite interface. This work showed that SSP is a promising technology for bimaterial/multimaterial composite manufacturing. The bimaterial composite parts achieved by SSP have a good composite interface and well-located material distribution.

Disruption of NAD(P)H:quinone oxidoreductase 1 gene in mice leads to 20S proteasomal degradation of p63 resulting in thinning of epithelium and chemical-induced skin cancer

NAD(P)H:quinone oxidoreductase 1 (NQO1) is a cytosolic enzyme that protects cells against chemical and radiation-induced oxidative stress and skin cancer. Disruption of NQO1 gene in mice showed thinning of skin epithelium and loss of cytokeratin 14, an early marker of skin differentiation. Immunohistochemistry and western analysis demonstrated downregulation of p63 in NQO1-/- mouse skin, as compared with wild-type (WT) mouse. Further analysis including modulation of NQO1 expression revealed a direct correlation between the levels of NQO1 and p63 in skin-derived keratinocytes and dermal fibroblasts. Modulation of proteasomal activity revealed that p63 is degraded by 20S proteasome and that this degradation is significantly rescued by NQO1. Coimmunoprecipitation studies showed that NQO1 interacts directly with p63 but not 20S to protect against this degradation. In addition, benzo[a]pyrene treatment led to induction of NQO1 and stabilization of p63 in WT but not in NQO1-/- mouse skin and keratinocytes. These data suggest that NQO1 controls stabilization of p63 and progression towards keratinocyte differentiation leading to normal skin development and presumably skin carcinogenesis.

Efficacy of Glutathione in Ameliorating Sulfur Mustard Analog-Induced Toxicity in Cultured Skin Epidermal Cells and in SKH-1 Mouse Skin In Vivo

Exposure to chemical warfare agent sulfur mustard (HD) is reported to cause GSH depletion, which plays an important role in HD-linked oxidative stress and skin injury. Using the HD analog 2-chloroethyl ethyl sulfide (CEES), we evaluated the role of GSH and its efficacy in ameliorating CEES-caused skin injury. Using mouse JB6 and human HaCaT epidermal keratinocytes, we observed both protective and therapeutic effects of exogenous GSH (1 or 10 mM) in attenuating a CEES-caused decrease in cell viability and DNA synthesis, as well as S and G(2)M phase arrest in cell cycle progression. However, the protective effect of GSH was stronger than its ability to reverse CEES-induced cytotoxic effect. The observed effect of GSH could be associated with an increase in intracellular GSH levels after its treatment before or after CEES exposure, which strongly depleted cellular GSH levels. N-Acetyl cysteine, a GSH precursor, also showed both protective and therapeutic effects against CEES-caused cytotoxicity. Buthionine sulfoximine, which reduces cellular GSH levels, caused an increased CEES cytotoxicity in both JB6 and HaCaT cells. In further studies translating GSH effects in cell culture, pretreatment of mice with 300 mg/kg GSH via oral gavage 1 h before topical application of CEES resulted in significant protection against CEES-caused increase in skin bifold and epidermal thickness, apoptotic cell death, and myeloperoxidase activity, which could be associated with increased skin GSH levels. Together, these results highlight GSH efficacy in ameliorating CEES-caused skin injury and further support the need for effective antioxidant countermeasures against skin injury by HD exposure.

High-temperature strength of compacted sub-micrometer aluminium powder

Aluminium powders with a mean particle size of around 1 μm were compacted by cold isostatic pressing (CIP) and additional forging. The specimens are characterized by hot compression tests, dilatometry and metallography. A 3D interconnected structure of alumina films <5 nm in thickness is observed by transmission electron microscopy and field emission gun scanning electron microscopy; it is associated with the natural oxide skin which covers every aluminium powder and occupies around 3 vol.%. The compression tests are carried out in the range of 350–520 °C at strain rates of 0.003–3 s −1. The compressive strength was 100–150 and 130–180 MPa for the CIPed and forged samples, respectively. The low strain rate sensitivity m (<0.08) suggests that the alumina network forms a barrier, which suppresses any restoration mechanism across the grain boundaries as well as grain boundary sliding during hot deformation. The high strength of such compacted sub-micron Al powder is attributed to the conservation of a 3D alumina closed cell network filled with elastoplastic aluminium.

Controlling Reactive Oxygen Species in Skin at Their Source to Reduce Skin Aging

Activity of an age-related, superoxide-forming, cell-surface oxidase (arNOX) comparing dermis, epidermis, serum, and saliva from female and male subjects ages 28-72 years measured spectrophotometrically using reduction of ferricytochrome c correlated with oxidative skin damage as estimated from autofluoresence of skin using an Advanced Glycation End products Reader (AGE-Reader; DiagnOptics B.V., Netherlands). By reducing arNOX activity in skin with arNOX-inhibitory ingredients (NuSkin's ageLOC technology), skin appearance was improved through decreased protein cross-linking and an accelerated increase in collagen.

Molecular Rectification in Metal−SAM−Metal Oxide−Metal Junctions

This Article compares the ability of self-assembled monolayers (SAMs) of alkanethiolates with ferrocene (Fc) head groups (SC(11)Fc), and SAMs of alkanethiolates lacking the Fc moiety (SC(10)CH(3) and SC(14)CH(3)), to conduct charge. Ultraflat surfaces of template-stripped silver (Ag(TS)) supported these SAMs, and a eutectic alloy of gallium and indium (EGaIn), covered with a skin of gallium oxide (presumably Ga(2)O(3)), formed electrical top-contacts with them. EGaIn is a liquid at room temperature, but its spontaneously formed surface oxide skin gives it apparent non-Newtonian properties and allows it to be molded into conically shaped tips; these tips formed soft electrical contacts with SAMs and formed stable SAM-based tunneling junctions in high (70-90%) yields. Measurements of current density, J, versus applied voltage, V, showed that tunneling junctions composed of SAMs of SC(11)Fc rectify current with a rectification ratio R approximately 1.0 x 10(2) (R = |J(-V)|/|J(V)| at +/-1 V and with a log-standard deviation of 3.0). In contrast, junctions lacking the Fc moiety, that is, junctions composed of SAMs of SC(n-1)CH(3) (with n = 11 or 15 and presenting terminal CH(3) groups), showed only slight rectification (R = 1.5 (1.4) and 2.1 (2.5), respectively). A statistical analysis of large numbers (N = 300-1000) of data gave detailed information about the spread in values and the statistical significance of the rectification ratios and demonstrated the ability of the experimental techniques described here to generate SAM-based junctions in high yield useful in physical-organic studies.

Does hepatitis C increase the accumulation of advanced glycation end products in haemodialysis patients?

Hepatitis C may cause increased levels of oxidative stress that contribute to accumulation of advanced glycation end products (AGEs), which increase the risk of cardiovascular disease (CVD). The aim of this study was to determine the influence of hepatitis C on AGE accumulation in haemodialysis patients. AGE accumulation was measured by means of skin autofluorescence (AF) in 92 haemodialysis (HD) patients and 93 age-matched healthy controls. In the HD patients, CVD-related biochemical variables were also measured. The HD patients were tested for hepatitis C virus (HCV) antibodies and allocated to a HCV+ or HCV- group. Skin AF of the healthy subjects was lower than skin AF in the HD patients (3.13 +/- 0.95 vs 2.2 +/- 0.47; P < 0.001). We calculated the average increase of skin AF in the healthy subjects to be 0.017 arbitrary units per year, being 14 times lower than in HD patients with CVD only and 20 times lower than in HD patients suffering from combined CVD and diabetes mellitus (DM). Multivariate regression analysis showed that AGE accumulation in HD patients can be described by the independent effects of age, DM, CVD and HD vintage. Although inter-cellular adhesion molecule 1 and liver enzymes were elevated in HCV+ HD patients, levels of oxidative stress markers and skin AF were not significantly different between HCV+ and HCV- HD patients. AGE accumulation was higher in the HD patients than in the healthy controls. AGE accumulation did not differ in HCV+ and HCV- HD patients. This might be due to the fact that hepatitis C did not cause oxidative stress in our HD population. Independent markers of AGE accumulation were age, HD vintage, DM and CVD, but not hepatitis C.

Reversibly Deformable and Mechanically Tunable Fluidic Antennas

AbstractThis paper describes the fabrication and characterization of fluidic dipole antennas that are reconfigurable, reversibly deformable, and mechanically tunable. The antennas consist of a fluid metal alloy injected into microfluidic channels comprising a silicone elastomer. By employing soft lithographic, rapid prototyping methods, the fluidic antennas are easier to fabricate than conventional copper antennas. The fluidic dipole radiates with ≈90% efficiency over a broad frequency range (1910–1990 MHz), which is equivalent to the expected efficiency for a similar dipole with solid metallic elements such as copper. The metal, eutectic gallium indium (EGaIn), is a low‐viscosity liquid at room temperature and possesses a thin oxide skin that provides mechanical stability to the fluid within the elastomeric channels. Because the conductive element of the antenna is a fluid, the mechanical properties and shape of the antenna are defined by the elastomeric channels, which are composed of polydimethylsiloxane (PDMS). The antennas can withstand mechanical deformation (stretching, bending, rolling, and twisting) and return to their original state after removal of an applied stress. The ability of the fluid metal to flow during deformation of the PDMS ensures electrical continuity. The shape and thus, the function of the antenna, is reconfigurable. The resonant frequency can be tuned mechanically by elongating the antenna via stretching without any hysteresis during strain relaxation, and the measured resonant frequency as a function of strain shows excellent agreement (±0.1–0.3% error) with that predicted by theoretical finite element modeling. The antennas are therefore sensors of strain. The fluid metal also facilitates self‐healing in response to sharp cuts through the antenna.

The surface structure of gas-atomized metallic glass powders

By means of X-ray photoelectron spectrometry depth profile and transmission electron microscopy analysis, this study reveals that amorphous Cu54Ni6Zr22Ti18 powders are enveloped by a thick (∼30 nm) oxide skin and a subsequent oxidation-induced partially nanocrystallized zone. The thick oxide skin changes the metallic glass surface to oxide ceramic. It is proposed that effective disruption of the oxide skin is essential to the sintering of metallic glass.

Age-related ENOX protein (arNOX) activity correlated with oxidative skin damage in the elderly.

ENOX proteins with an oscillatory pattern of production of superoxide (measured by ferricytochrome c reduction) and with a period length of 26 min increase linearity with age beginning at about 30 y to a maximum of about age 60. The proteins are shed and appear in serum, saliva and urine. Enhanced arNOX activity correlates with age and with oxidative changes contributing to skin aging. Topical cosmetic preparations containing substances that block arNOX activity are under evaluation to reduce visible symptoms of skin aging.

Viscoelastic properties of oxide-coated liquid metals

Many liquid metals exposed to air develop an oxide film on their outer surface. This film is sufficiently solid-like to provide mechanical stability to small liquid metal droplets, yet weak enough to allow the droplets to be malleable. These properties are useful in both micro-electronics and microfluidics; however, little is known about how to characterize them. Here we probe the elastic, yielding, and relaxation properties of oxide-coated gallium and eutectic gallium indium using a rheometer equipped with a parallel-plate geometry. By using parallel plates of different size, we show that surface stresses dominate bulk stresses. These experiments also demonstrate that the apparent elastic properties of the oxide film are highly sensitive to its strain history. Moreover, the apparent elasticity is sensitive to the stresses stored in the oxide skin. We probe these stresses and their time-dependence, with both torque and normal force measurements. We also characterize the time-dependence of the elasticity b...

Definition of the Influence of Pore Size on the Fatigue Limit Using Short Crack Propagation Experiments

Abstract Aluminium high pressure die casting is used to reduce costs and weight of various components in the automotive industry. The main problem of components made by pressure die casting is connected with inherent flaws (porosity, oxide skins, etc.), which can hardly be avoided. Today the fatigue calculation of aluminium high pressure die cast parts is usually performed using two S/N curves—one account for the flawed basic material and one for the pore-free surface layer. This does not provide an accurate estimate for the computation of the lifetime or safety against failure of the component. In a cast component the number and size of pores increases towards the center. As a consequence the fatigue strength decreases. This inhomogeneous distribution throughout the component has to be taken into account for a realistic estimation of the fatigue strength. To do so, two models are required—one to compute the pore size distribution in the component [Oberwinkler, C., Leitner, H., and Eichlseder, W. “Improvement of an Existing Model to Estimate the Pore Distribution for A Fatigue Proof Design of Al HPDC Components,” TMS 2009, Shape Casting: 3rd Int. Symposium, 2009] and a material model [Oberwinkler, C., Leitner, H., and Eichlseder, W. “Computation of Fatigue Safety Factors for High-Pressure Die Cast (HPDC) Aluminium Components Taking into Account the Pore Size Distribution,” SAE World Congress 2009], which describes the influence of the defect size on the fatigue strength. This paper focuses on the definition of the correlation between the fatigue strength and the pore size. The lifetime of a component is defined by the crack growth if the crack initiation period can be neglected. Casting defects usually show high stress concentration factors due to their irregular shapes where cracks initiate. In this case the defect size is a part of the total crack length. Pores can be considered as physically short cracks because of their size of approximately 10–500 μm. The El-Haddad equation [El Haddad, M. H., Smith, K. N., and Topper, T. H., “Fatigue Crack Propagation of Short Cracks,” ASME Transactions, Vol. 101, 1979, pp. 42–46] has been used to describe the influence of the defect size on the fatigue strength. Short crack experiments have been performed to show that due to the shorter cracks the threshold stress intensity factor decreases below the threshold stress intensity factor of long cracks.

The co-application effects of fullerene and ascorbic acid on UV-B irradiated mouse skin

The role of fullerene as a pro-oxidant or anti-oxidant in Ultraviolet B ray (UV-B)-induced disorders in mouse skin was investigated. Fullerene gave no photo-toxic effect to UV-B-irradiated mouse skin. Since erythema was concentrated at the pore circumference in a UV-B irradiation experiment in mouse skin, the sebaceous gland pairs was strongly implicated as a site for the generation of reactive oxygen species (ROS). In a histological evaluation of the skin stained with CH(3)MDFDA (ROS index) and YO-Pro-1 (apoptosis index), the fluorescence intensity of a sebaceous gland significantly increased with UV-B irradiation. With the application of fullerene to UV-irradiated mouse skin, no toxicity was recognized in comparison with the control, and erythema, the ROS index, and the apoptosis index decrease with the application of fullerene. Ascorbyl radical (AA*) increased with the application of ascorbate (AA) to UV-B-irradiated mouse skin, and AA* decreased with the application of fullerene. The co-application of AA and fullerene, which suppressed AA* in vitro, significantly suppressed erythema, and also suppressed both the ROS index and apoptosis index in mouse skin after UV-B irradiation. In both mouse skin at 48 h after UV-B irradiation and in an attempt to reproduce this phenomenon artificially in vitro, a similar high AA* peak (AA*/H*>4) was observed in electron spin resonance (ESR) charts. The binding of fullerene with AA impairs the Fenton reaction between AA and Fe-protein based on the observation of ascorbate-specific UV absorption and a linear equation for the calibration curve. Therefore, fullerene may impair the intercalation of AA to a heme pocket by binding with AA. These results suggest that the co-application of AA and fullerene is effective against oxidative skin damage caused by UV-B irradiation, and the development of an AA* inhibitor such as fullerene should be useful for reducing organ damage associated with Fe-protein oxidation.

Enhanced heterogeneous nucleation in AZ91D alloy by intensive melt shearing

Intensive melt shearing was applied to the commercial AZ91D alloy melt to investigate its effects on grain refinement. Alloy melts with and without melt shearing were also filtered using a pressurized filtration technique to concentrate the potential nucleating particles for electron microscopic examination. The results showed that intensive melt shearing resulted in significant refinement of both the Al 8Mn 5 intermetallics and the primary α-Mg phase in the as-cast AZ91D alloy, and that this grain-refining effect is insensitive to the superheat and can persist even after prolonged isothermal holding. The pressurized filtration experiments showed for the first time that oxide films and skins consist of nano-sized MgO particles populated densely in a liquid matrix. Intensive melt shearing can effectively disperse such MgO particles throughout the alloy melt. The HRTEM investigation and detailed crystallographic analysis confirmed that dispersed MgO particles act as potent heterogeneous nucleation sites for both the Al 8Mn 5 and α-Mg phase.

Study on blood and buccal mucous related genes expression in the patients suffering from coal-burning arsenism

To observe the oxidative stress and skin disease related genes expression in cells exfoliated from buccal mucous and blood in patients with arseniasis caused by coal-burning pollution in Guizhou. Buccal and blood cell samples were collected from arsenicosis patients and healthy subjects residing in Guizhou. Total RNA was isolated for related genetic transcript analysis. Real-time PCR (RT-PCR) was employed to detect the change of related gene expression. Serum indicators of liver injury, such as serum alanine aminotransferase (ALT), were detected. Serum levels of ALT in arsenic patients were significantly higher than those in controls (36.9 +/- 17.4 / 25.8 +/- 2.4, t = 0.019, P < 0.05) even when no symptom was found in these patients. Buccal K10 gene was overexpressed (3.6 +/- 0.9 / 1.8 +/- 0.7, t = 0.041, P < 0.05) and MT-1 expression was significantly lower (0.8 +/- 0.5 / 3.9 +/- 0.4, t = 0.028, P < 0.05) in arsenic patients than that in control while the expressions of other genes were the same as those of controls. The expressions of MT-1 and K10 gene in buccal mucous cells may be regarded as sensitive molecular markers for skin and hepatic pathologic changes in arsenic patients. Protection of liver function should be considered as an important step in arseniasis prevention and control.

Mathematical Modeling and Computer Simulation of Molten Aluminum Purification by Flotation in Stirred Reactor

The removal of inclusions by flotation in mechanically agitated vessels is widely used in liquid aluminum treatments. Originating from different sources (oxide skins, refractory, or recycling wastes), inclusions may have disastrous repercussions such as deterioration of the physical properties of the cast products or difficulties during forging processes. With the aim of both a better understanding of the physical processes acting during flotation and the optimization of the refining process, a mathematical modeling of the behavior of the population of inclusions has been set up. Transport phenomena, agglomeration of inclusions, and flotation are considered here. The model combines population balance with convective transport of the inclusions, in order to calculate the time evolution of the inclusion size distribution. An operator-splitting technique is employed to solve the coupled population balance equation (PBE) and the transport equation. The transport equation is solved using a finite volume technique associated with a total variation diminishing scheme, whereas the PBE resolution relies on the fixed pivot technique developed by Kumar and Ramkrishna. A laboratory-scale flotation vessel is modeled and the results of a two-dimensional (2-D) simulation are presented.

Enzyme-Independent NO Stores in Human Skin: Quantification and Influence of UV Radiation

Nitric oxide (NO) has many functions in the skin, including the mediation of inflammation and antimicrobial defense, wound healing, regulation of keratinocyte homeostasis, and regulation of apoptosis following UV radiation. NO is synthesized by a family of NO synthase enzymes, but its rapid release following UV exposure suggests the existence of preformed stores. NO can be converted into nitrite or nitrosothiols that are stable until cleaved by UV to release NO. Using dermal microdialysis, suction blister epidermal samples, and sweat collection, we demonstrated cutaneous concentrations of total NO-related products of 12+/-5.97 microM, 0.03+/-0.03 micromol mg(-1) epidermal protein, and 22+/-9.34 microM, respectively. The predominant oxyanion was nitrate (60-75%) followed by nitrite. S-Nitrosothiols were barely detectable. Serum total NO-related products correlated directly with those of the upper dermis and sweat (R(2)=0.62 and 0.3, respectively). UVA irradiation (10 mW cm(-2)) increased the yield of NO-related products by microdialysis, peaking after 30 minutes. Dialysis with noradrenaline abrogated this rise. Both the skin and the dermal vasculature contain biologically significant stores of NO, particularly nitrite, which can be directly mobilized by UVA irradiation. The level of circulating NO-related products probably determines skin-bound stores.

Dynamic wetting and heat transfer at the initiation of aluminum solidification on copper substrates

Dynamic wetting and heat transfer during the start of solidification were studied with the help of molten aluminum droplets falling from a crucible onto a copper substrate. A high-speed camera captured the change in the spreading droplet’s geometry, while thermocouple, inserted inside the substrate, allowed a heat transfer analysis to be performed. Droplet spreading factors and interfacial heat fluxes were then used to, respectively, characterize dynamic wetting and heat transfer for the various experimental conditions explored. These were: (1) effects of chemical composition of the aluminum alloy, (2) initial temperature of the substrate, (3) surface roughness of the substrate, and (4) composition of the gaseous atmosphere. The experiments were all carried out in gaseous atmospheres containing oxygen in sufficient amount to form oxide skins at the surface of the droplets and the substrates. The results showed instances where an improvement in the dynamic wetting was accompanied by an increase in heat transfer during the early stages of solidification but this was not systematic. In these cases where a positive correlation was not observed, it was postulated this was caused by factors such as variations in the oxidation at the surface of the substrates and the droplets as well as gas trapped at the interface between the droplets and the substrates.