ABSTRACT
Electrospun zein nanofibers have attracted interest as drug delivery systems due to their propensity for controlled drug release, flexible structure and low toxicity. However, comparatively little is known regarding the relationship between production method and fiber characteristics, both in terms of fiber architecture and protein structure. Here we use a range of imaging and spectroscopic techniques to elucidate the effects of solvent composition on zein secondary structure, fiber diameter and fiber integrity, plus we utilize the new technique of transition temperature microscopy to examine the thermal properties of the fibers. Zein nanofibers were prepared using ethanol, acetic acid and water mixes as solvents, alone and with plasticizers (polyethylene glycol, glycerol) and casein. Electrospinning was performed under controlled conditions and the products characterized using scanning electron microscopy (SEM), attenuated total reflection Fourier Transform infrared spectrometry (ATR - FTIR) and transition temperature microscopy (TTM). The choice of solvent, concentration and voltage, alongside the presence of additives (plasticizers and casein) were noted to influence both the diameter of the fibers and the tendency for bead formation. A relationship was noted between protein secondary structure and fiber architecture, with an enhanced ß-sheet content, enhanced by the inclusion of casein, being associated with higher beading. In addition, thermal imaging of electrospun zein fiber mats was successfully achieved using TTM via two dimensional mapping of the softening temperatures across the spun samples, in particular demonstrating the plasticizing effects of the polyethylene glycol and glycerol.
Subject(s)
Nanofibers/chemistry , Nanotechnology/methods , Temperature , Zein/chemistry , Electric Conductivity , Microscopy , Nanofibers/ultrastructure , Protein Structure, Secondary , Solutions , Solvents , Spectroscopy, Fourier Transform Infrared , Transition Temperature , ViscosityABSTRACT
The potential disease-carrying mosquito, Aedes japonicus (Theobald) (Diptera: Culicidae), was identified among larvae collected in suburban Vancouver, BC, in July 2014, and over 200 were found at the same site in February 2015 where it presumably had overwintered in the egg stage. In late May 2015, a female was captured taking a bloodmeal 13 km east of the larval site. This population and those in the Washington and Oregon states are clearly disjunct from those in eastern North America, and their origin, probably from one or more different introductions from Asia, is discussed. Key characters of those in British Columbia are examined and match the description of subspecies japonicus, presumably like the others in North America.
Subject(s)
Aedes , Insect Vectors , Animals , British Columbia , FemaleABSTRACT
Addition of a drug to a self-emulsifying drug delivery system (SEDDS) can affect the emulsification process after administration, leading to variation in the emulsion droplet size formed and potentially its clinical behavior (Mercuri et al., Pharm. Res., 2011, 28, 1540-1551). However, the mechanisms involved and, in particular, the location of the drug within the system are poorly understood. Here, we have investigated the location of a model drug, ibuprofen, in the emulsions formed from a simple anhydrous SEDDS (soybean oil, Tween 80 and Span 80), using a range of physical characterization techniques. (1)H NMR studies showed an interaction between the drug and the polyoxyethylene chains of the surfactant Tween 80. Micropolarity assessment of the emulsion droplet interfacial region, using the chemical probes pyrene and Reichardt's dye, confirmed this interaction, and suggested that the drug was altering the microenvironment around the surfactants, and hence the behavior of the SEDDS with water during emulsification. Both dielectric spectroscopy and polarized light microscopy highlighted the differential behavior with water of placebo and drug-loaded SEDDS, also seen in the initial visual observational studies on the emulsification performance of the SEDDS. (1)H NMR studies with three other NSAIDs indicate that this effect is not confined to ibuprofen alone. The study has therefore indicated that the drug's influence on the emulsification process may be related to interactions within the microenvironment of the surfactant layer. Furthermore, such interactions may be usefully identified and characterized using a combination of micropolarity, spectroscopic and microscopic methods.
Subject(s)
Emulsifying Agents/chemistry , Emulsions/chemistry , Ibuprofen/chemistry , Anti-Inflammatory Agents, Non-Steroidal/chemistry , Chemistry, Pharmaceutical/methods , Drug Delivery Systems/methods , Hexoses/chemistry , Magnetic Resonance Spectroscopy/methods , Microscopy, Polarization/methods , Particle Size , Polyethylene Glycols/chemistry , Polysorbates/chemistry , Pyrenes/chemistry , Pyridinium Compounds/chemistry , Soybean Oil/chemistry , Surface-Active Agents/chemistry , Water/chemistryABSTRACT
Sexual communication of nun moth, Lymantria monacha (L.), pink gypsy moth, Lymantria mathura Moore, and fumida tussock moth, Lymantria fumida Butler (all Lepidoptera: Noctuidae: Lymantriinae), is known to be mediated by pheromones. We now show that males are attracted by the sounds of conspecific females over short distances and that wing fanning male and female L. monacha, L. mathura and L. fumida produce species- and sex-specific wing beat and associated click sounds that could contribute to reproductive isolation. Evidence for short-range communication in these lymantriines includes (i) scanning electron micrographs revealing metathoracic tympanate ears, (ii) laser interferometry showing particular sensitivity of tympana tuned to frequency components of sound signals from conspecifics, and (iii) phonotaxis of male L. monacha and L. fumida to speakers playing back sound signals from conspecific females. We conclude that tympanate ears of these moths have evolved in response not only to bat predation, but also for short-range mate finding and possibly recognition.
Subject(s)
Moths/anatomy & histology , Moths/physiology , Animal Communication , Animals , Auditory Perception , Ear/anatomy & histology , Ear/physiology , Female , Hearing , Male , Microscopy, Electron, Scanning , Sexual Behavior, Animal , Species Specificity , Wings, Animal/physiologyABSTRACT
The response to hydration of the internal protein dynamics was studied by the means of solid state NMR relaxation and magic angle spinning exchange techniques. Two proteins, lysozyme from bacteriophage T4 and human alphaB-crystallin were used as exemplars. The relaxation rates R1 and R1rho of 13C and 15N nuclei were measured as a function of a hydration level of the proteins in the range 0-0.6 g of water/g of protein. Both proteins were totally 15N-enriched with natural 13C abundance. The relaxation rates were measured for different spectral bands (peaks) that enabled the characterization of the dynamics separately for the backbone, side chains, and CH3 and NH3+ groups. The data obtained allowed a comparative analysis of the hydration response of the protein dynamics in different frequency ranges and different sites in the protein for two different proteins and two magnetic nuclei. The most important result is a demonstration of a qualitatively different response to hydration of the internal dynamics in different frequency ranges. The amplitude of the fast (nanosecond time scale) motion gradually increases with increasing hydration, whereas that of the slow (microsecond time scale) motion increases only until the hydration level 0.2-0.3 g of water/g of protein and then shows almost no hydration dependence. The reason for such a difference is discussed in terms of the different physical natures of these two dynamic processes. Backbone and side chain nuclei show the same features of the response of dynamics with hydration despite the fact that the backbone motional amplitudes are much smaller than those of side chains. Although T4 lysozyme and alphaB-crystallin possess rather different structural and biochemical properties, both proteins show qualitatively very similar hydration responses. In addition to the internal motions, exchange NMR data enabled the identification of one more type of motion in the millisecond to second time scale that appears only at high hydration levels. This motion was attributed to the restricted librations of the protein as a whole.
Subject(s)
Bacteriophage T4/enzymology , Muramidase/chemistry , alpha-Crystallin B Chain/chemistry , Carbon Isotopes , Humans , Magnetic Resonance Spectroscopy , Muramidase/metabolism , Nitrogen Isotopes , Thermodynamics , Water/chemistryABSTRACT
When a female mosquito bites, it carries away a blood sample containing specific antibodies that can provide a history of the immune responses of its vertebrate host. This research examines the limits and reliability of a technique to detect antibodies in blood-fed mosquitoes in the laboratory. Mosquitoes were fed on blood containing a specific antibody, and then they were assayed using an enzyme-linked immunosorbent assay to determine the limits of detection of antibody over time, at different temperatures and initial antibody concentrations. The antibody, at an initial concentration of 1 microg/ml, could be detected in mosquitoes for 24-48 h after feeding. Blind tests simulating the assay of feral mosquitoes were used to test the reliability of the method and detected positive mosquitoes with few false negatives and no false positives. Specific antibodies also could be detected in mosquitoes that had been air-dried or preserved in ethanol. This research indicates that, in theory, the collection and immunological assay of blood-fed mosquitoes could be developed to detect and monitor infectious disease in wildlife.
Subject(s)
Animals, Wild/immunology , Antibodies/analysis , Culicidae/physiology , Feeding Behavior/physiology , Animals , Animals, Wild/blood , Antibodies/blood , Humans , MiceABSTRACT
Heating with microwave energy and tannin complexation of kafirin both increase the tensile strength of cast kafirin bioplastic films. The effects of these treatments on the molecular structure of kafirin and of kafirin in the film were investigated. SDS-PAGE of heated wet kafirin showed an increase in kafirin oligomers. Disulfide groups increased in heated kafirin and in films made from the heated kafirin. Fourier transform infrared (FTIR) spectroscopy of heated kafirin and films made from the heated kafirin indicated an increase in beta-sheet conformation. In contrast, kafirin complexation with tannic acid (TA) and sorghum condensed tannin (SCT) resulted in a slight decrease in beta-sheet conformation in the kafirin and a larger decrease in the kafirin in the films. Raman spectroscopy showed that, with TA, there was a shift in peak from 1710 to 1728 cm(-1) for kafirin-tannic acid complexes, indicating kafirin and tannic acid interaction. The protein conformational changes presumably facilitated cross-linking between kafirin molecules and/or between kafirin and the tannins. Thus, although both heating with microwave energy and tannin complexation cause cross-linking of kafirin to increase film tensile strength, their effects on kafirin structure appear to be different.
Subject(s)
Hot Temperature , Microwaves , Plant Proteins/chemistry , Tannins/chemistry , Electrophoresis, Polyacrylamide Gel , Protein Conformation , Protein Structure, Secondary , Sorghum/chemistry , Spectroscopy, Fourier Transform Infrared , Spectrum Analysis, Raman , Tensile StrengthABSTRACT
Synthetic and recombinant peptide models of the central repetitive domain of the high molecular weight subunits of wheat glutenin with different numbers of the consensus repeat motifs PGQGQQ + GYYPTSLQQ (21, 45, 110, and 203 residues long) and a recombinant 58,000-Da relative molecular mass (M(r) 58,000) repetitive peptide from a single subunit (1Dx5) are studied using Fourier transform IR spectroscopy. The spectra of the dry peptides are very similar; at low water contents (<76% relative humidity) there is an increase in beta-sheet structures in all peptides. However, on further hydration the content of beta sheets decrease and more beta turns are observed. The changes during the second step of hydration are very marked in the 21 and 45 residue peptides, but they are less apparent in the longer perfect repeat peptides. In the 110 and 203 residue peptides hydration results in increased contents of intermolecular beta-sheets and less beta-turn formation. In contrast, the beta-turn content of the M(r) 58,000 peptide increases during the second hydration step. The decreased extent of structure changes with increasing chain length indicates that cumulative intermolecular interactions, in particular hydrogen bonds, are an important factor in determining the structures in the solid state. The regularity of the perfect repeat sequences in the 21, 45, 110, and 203 residue peptides may favor the formation of larger stretches of intermolecular beta sheets. In contrast, the M(r) 58,000 peptide contains imperfect repeats (in common with native glutenin subunits), which may limit its ability to form intermolecular beta sheets.
Subject(s)
Glutens/analogs & derivatives , Glutens/chemistry , Peptide Fragments/chemistry , Peptide Fragments/metabolism , Protein Subunits/chemistry , Triticum/chemistry , Amino Acid Sequence , Molecular Weight , Protein Structure, Tertiary , Spectroscopy, Fourier Transform Infrared , Water/chemistryABSTRACT
The gene encoding high-molecular-weight (HMW) subunit 1Bx20 was isolated from durum wheat cv. Lira. It encodes a mature protein of 774 amino acid residues with an M(r) of 83,913. Comparison with the sequence of subunit 1Bx7 showed over 96% identity, the main difference being the substitution of two cysteine residues in the N-terminal domain of subunit 1Bx7 with tyrosine residues in 1Bx20. Comparison of the structures and stabilities of the two subunits purified from wheat using Fourier-transform infra-red and circular dichroism spectroscopy showed no significant differences. However, incorporation of subunit 1Bx7 into a base flour gave increased dough strength and stability measured by Mixograph analysis, while incorporation of subunit 1Bx20 resulted in small positive or negative effects on the parameters measured. It is concluded that the different effects of the two subunits could relate to the differences in their cysteine contents, thereby affecting the cross-linking and hence properties of the glutenin polymers.
Subject(s)
Glutens/analogs & derivatives , Triticum/genetics , Amino Acid Sequence , Circular Dichroism , Crosses, Genetic , Cysteine/chemistry , Electrophoresis, Polyacrylamide Gel , Glutens/chemistry , Molecular Sequence Data , Molecular Weight , Protein Structure, Tertiary , Sequence Homology, Amino Acid , Spectroscopy, Fourier Transform Infrared , Tyrosine/metabolismABSTRACT
The observed resistance to pepsinolysis of known food allergens has been suggested as a predictor of their allergenic risk. Consequently, resistance to pepsinolysis has become incorporated into decision tree assessment for potential allergenic risk posed by novel foods. However, existing methods take little account of the interaction between food structure and physiological conditions existing during digestion in vivo. Here we show that a range of protein allergens can adsorb to model stomach emulsions, providing a further means of resisting digestion. We also show that raising the pH and the addition of bile salts to a model stomach emulsion, thereby mimicking the duodenal environment, has the effect of desorbing the adsorbed protein.
Subject(s)
Allergens/chemistry , Emulsions/chemistry , Gastric Juice/chemistry , Adsorption , Allergens/metabolism , Animals , Bile Acids and Salts/pharmacology , Cattle , Dose-Response Relationship, Drug , Emulsions/metabolism , Proteins/chemistryABSTRACT
The cupin family comprises a family of proteins possessing a common beta-barrel structure that is thought to have originated in a prokaryotic ancestor. This structural motif is found as a single domain in fungal spherulins, fern sporulins and the germins/oxalate oxidase proteins of plants, while the globular storage proteins of plants, called legumins (11 S) and euvicilins (7 S), are two-domain cupins. The 11 S globulins are hexameric heteroligomeric proteins of M (r) approximately 360000, with each subunit comprising an acidic 30000-40000- M (r) polypeptide that is disulphide-linked to a 20000- M (r) basic polypeptide. A number of cupins have been identified as major plant food allergens, including the 7 S globulins of soybean (beta-conglycinin), peanut (conarachin; Ara h 1), walnut (Jug r 2) and lentil, and the 11 S globulins of peanut (arachin; Ara h 3), soybean (glycinin) and possibly also coconut and walnut. Other members of the cupin superfamily have not been identified as allergens, with the exception of one germin (germination-specific protein) from pepper. Cupins are generally very stable proteins. A summary of our current knowledge of allergenic seed storage globulins will be presented, together with an overview of cupin structure and stability properties, as illustrated by the allergenic soya globulins, glycinin and beta-conglycinin.
Subject(s)
Allergens/chemistry , Plant Proteins/chemistry , Amino Acid Motifs , Antigens, Plant , Globulins/chemistry , Hot Temperature , Image Processing, Computer-Assisted , Models, Molecular , Seed Storage Proteins , Soybean Proteins/chemistry , Spectroscopy, Fourier Transform Infrared , TemperatureABSTRACT
Molecular motions of polycrystalline cellobiose have been investigated by measuring proton spin-lattice relaxation times, T1 and T1rho, and the second moment, M2, in both protonated and D2O exchanged forms over the temperature range 120-380 K. T1 relaxation is dominated by the motions of hydroxyl groups between 150 and 380 K, characterised by an activation energy of about 8.74 kJ/mol, whereas T1rho relaxation is driven by the motions of the same groups between 120 and 300 K. T1rho results suggest that hydroxyl groups have a distribution of dynamics. Motion of methylene groups was detected in the second-moment experiments at about 350 K, characterised by activation energy of about 40 kJ/mol. Consideration of the calculated and observed rigid-lattice second moments suggests that the reported X-ray data are incorrect for the inter-proton distance on C6'. 13C CPMAS spectra of both protonated and deuterated cellobiose have also been measured. Spectra of the deuterated material showed the existence of a second crystalline form in addition to the normal form.
Subject(s)
Cellobiose/chemistry , Cellulose/chemistry , Crystallization , Magnetic Resonance SpectroscopyABSTRACT
It has been reported that caroubin, a protein mixture obtained from carob seeds, has rheological properties similar to those of gluten. Comparative studies of the effects of hydration and temperature on caroubin and gluten were carried out with the aid of NMR, FTIR, scanning electron microscopy, and differential scanning calorimetry techniques. The results show that caroubin has a more ordered structure than gluten and that hydration has little effect on its secondary structure when compared to gluten. Caroubin is more easily accessible to water than gluten, suggesting that caroubin is more hydrophilic in nature. On hydration, caroubin, like gluten, forms fibrillar structures and sheets.
Subject(s)
Glutens/chemistry , Polysaccharides/chemistry , Seeds/chemistry , Calorimetry, Differential Scanning , Galactans , Glutens/ultrastructure , Magnetic Resonance Spectroscopy , Mannans , Microscopy, Electron, Scanning , Plant Gums , Polysaccharides/ultrastructure , Seeds/ultrastructure , Spectroscopy, Fourier Transform Infrared , Temperature , WaterABSTRACT
Crystalline alpha-D-galacturonic acid monohydrate has been studied by 13C CPMAS NMR and X-ray crystallography. The molecular dynamics were investigated by evaluating 13C spin-lattice relaxation in the rotating frame (T1rho) and chemical-shift-anisotropy properties of each carbon. Only limited molecular motions can be detected in the low frequency (< 10(4) Hz) range by 13C relaxation time measurements (T1rho) and changes of chemical shift anisotropy properties as a function of temperature. X-ray analysis (at both ambient temperature and 150 K) shows that the acid has the usual chair-shaped, pyranose ring conformation, and that the acid and water molecules are linked, through all their O-H groups, in an extensively hydrogen-bonded lattice.
Subject(s)
Hexuronic Acids/chemistry , Carbohydrate Conformation , Crystallography, X-Ray/methods , Magnetic Resonance Spectroscopy/methods , Methylation , Models, Molecular , WaterABSTRACT
Free radical scavenging reactions of green tea polyphenols (GTP) were investigated with electron spin resonance (ESR) spectroscopy in the phospholipid bilayer of liposomes, using 1, 1-diphenyl-2-picrylhydrazyl (DPPH) radical as a model. The results showed that (1) GTP reacts with DPPH radicals in the bilayer of liposomes of both 1-hexadecanoyl-2-[(cis,cis,cis,cis,cis,cis)-4,7,10, 13,16,19-docosahexaenoyl]-sn-glycero-3-phosphocholine (DHAPC) and 1, 2-di[cis-9-hexadecenoyl]-sn-glycero-3-phosphocholine) (DPPC); and (2) GTP protects DHAPC liposomes effectively from the oxidation initiated by DPPH radicals. These results provide direct evidence that GTP reacts with free radicals in the model membrane and support the hypothesis that GTP protects unsaturated phospholipids from oxidation by reacting directly with the radicals.
Subject(s)
Bepridil/analogs & derivatives , Flavonoids , Lipid Bilayers , Phenols/chemistry , Picrates , Polymers/chemistry , Tea , Bepridil/chemistry , Biphenyl Compounds , Electron Spin Resonance Spectroscopy , Free Radicals , PolyphenolsABSTRACT
The high molecular weight (HMW) subunit group of wheat seed storage proteins impart elasticity to wheat doughs and glutens. They consist of three domains: non-repetitive N- and C-terminal domains, which contain cysteine residues for covalent cross-linking, and a central domain consisting of repeated sequences. The circular dichroism and infrared (IR) spectra of an intact HMW subunit were compared with those of a peptide corresponding to the central repetitive domain expressed in Escherichia coli. This allowed the structure of the central domain to be studied in the absence of the N- and C-terminal domains and the contributions of these domains to the structure of the whole protein to be determined. In solution the peptide showed the presence of beta-turns and polyproline II-like structure. Variable temperature studies indicated an equilibrium between these two structures, the polyproline II conformation predominating at low temperatures and the beta-turn conformation at higher temperatures. IR in the hydrated solid state also indicated the presence of beta-turns and intermolecular beta-sheet structures. In contrast, spectroscopy of the whole subunit showed the presence of alpha-helix in the N- and C-terminal domains. The content of beta-sheet was also higher in the whole subunit, indicating that the N- and C-terminal domains may promote the formation of intermolecular beta-sheet structures between the repetitive sequences, perhaps by aligning the molecules to promote interaction.
Subject(s)
Peptides/chemistry , Plant Proteins/chemistry , Triticum/chemistry , Amino Acid Sequence , Circular Dichroism , Peptides/genetics , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Repetitive Sequences, Amino Acid , Spectrophotometry, Ultraviolet , Spectroscopy, Fourier Transform InfraredABSTRACT
The behaviour of water in the presence of proteins and DNA as elucidated by nuclear magnetic resonance is reviewed. The picture that emerges is that in dilute solution only those water molecules in the interior of the biopolymers or in clefts have their motions substantially affected. In concentrated systems the situation is more complicated because many more motions have to be considered, but there is no evidence of special effects due to the biopolymers being present. The case of nonfreezing water in protein solutions is considered, and it is suggested that this is not evidence for 'bound' water but simply due to the effects of the inhibition of protein precipitation.
Subject(s)
DNA/metabolism , Magnetic Resonance Spectroscopy , Proteins/metabolism , Water/metabolism , Biopolymers/chemistry , Biopolymers/metabolism , DNA/chemistry , Freezing , Protein Binding , Proteins/chemistry , Protons , Solutions , Water/chemistryABSTRACT
The solid state 13C CPMAS NMR spectra of plant cell walls are often complex owing to superposition of resonances from different polysaccharides and the heterogeneity of the cell wall assembly. In this paper, we describe the application of a set of proton relaxation-induced spectral editing (PRISE) experiments which combine 1H relaxation properties (T1, T1rho, T2) with 13C high resolution spectroscopy (CPMAS) to relate the dynamics of the plant cell walls and model systems to their domain structural details. With PRISE it has been found that in plant cell wall materials, cellulose is always associated with the long components of spin-lattice relaxation in both the laboratory and rotating frames whereas non-cellulose polysaccharides (pectin and hemicellulose) are associated with the short ones. For the proton T2 relaxation, cellulose is only associated with the short component (below 20 micros), pectin contributes to both the short component and the long one.
Subject(s)
Cell Wall/chemistry , Magnetic Resonance Spectroscopy , Plants/chemistry , Carbon Isotopes , Cellulose , Magnetic Resonance Spectroscopy/methods , Pectins , Polysaccharides , Solanum tuberosumABSTRACT
New water-soluble and lipid-soluble spin probes suitable for the ESR investigation of the physical states and interactions of components of dough have been developed. This paper reports some preliminary findings on the suitability of these probes for this type of investigation. Rotational correlation times have been measured for the spin probes in water, dough, oil, and a starch/water mixture. An increase in rotational correlation time of the spin probe corresponds to an increase in microviscosity of the medium. Changes observed in correlation times of the water-soluble spin probes in doughs and in water/starch mixtures clearly correspond to a gelatinization process when the mixtures are heated above 60 degrees C. These irreversible changes, clearly important in the baking process, were monitored by following the change of mobility of a spin probe in doughs over a wide temperature range. The similarity of the results from the two sets of experiments suggests that the phenomenon of the increase of correlation times with temperature in doughs is attributable to the starch component. The lipid-soluble spin probe was found to be located preferentially in the lipid phase of the dough.
Subject(s)
Flour , Lipids , Water , Electron Spin Resonance Spectroscopy , Food HandlingABSTRACT
13C NMR with magic angle spinning (MAS) has been employed to investigate the cell walls of potatoes and Chinese water chestnuts over a range of hydration levels. Both single-pulse excitation (SPEMAS) and cross-polarization (CPMAS) experiments were carried out. Hydration led to a substantial increase in signal intensities of galactan and galacturonan in the SPEMAS spectra and a decrease in line width, implying mobilization in the backbone and side chains of pectin. In CPMAS spectra of both samples, noncellulose components showed signal loss as hydration increased. However, the signals of some galacturonan in the 3(1) helix configuration remained in the spectra even when the water content was as high as 110%. Cellulose was unaffected. It is concluded that the pectic polysaccharides experience a distribution of molecular conformations and mobility, whereas cellulose remained as typical rigid solid.