Polymer-lipid interactions:biomimetic self-assembly behaviour and surface properties of poly(styrene-alt-maleic acid) with diacylphosphatidylcholines

Abstract Various lubricating body fluids at tissue interfaces are composed mainly of combinations of phospholipids and amphipathic apoproteins. The challenge in producing synthetic replacements for them is not replacing the phospholipid, which is readily available in synthetic form, but replacing the apoprotein component, more specifically, its unique biophysical properties rather than its chemistry. The potential of amphiphilic reactive hypercoiling behaviour of poly(styrene-alt-maleic acid) (PSMA) was studied in combination with two diacylphosphatidylcholines (PC) of different chain lengths in aqueous solution. The surface properties of the mixtures were characterized by conventional Langmuir-Wilhelmy balance (surface pressure under compression) and the du Noüy tensiometer (surface tension of the non-compressed mixtures). Surface tension values and 31P NMR demonstrated that self-assembly of polymer-phospholipid mixtures were pH and concentration-dependent. Finally, the particle size and zeta potential measurements of this self-assembly showed that it can form negatively charged nanosized structures that might find use as drug or lipids release systems on interfaces such as the tear film or lung interfacial layers. The structural reorganization was sensitive to the alkyl chain length of the PC.

Bilirubin: An animal pigment in the Zingiberales and diverse angiosperm orders

Strelitziaceae is a tropical monocot family comprising three genera and seven species: Ravenala Adans and Phenkospermum Endl., which are monotypic, and five species of Strelitzia Aiton. All species produce woody capsular fruits that contain vibrantly colored arillate seeds. Arils of the Strelitzia species are orange, those of Phenakospermum are red, and those of Ravenala are blue. Unlike most plant pigments, which degrade after cell death, aril pigments in the family persist for decades. Chemical properties of the compounds are unusual, and do not match those of known pigment classes (carotenoids, flavonoids, betalains, and the chlorophylls). I isolated the orange pigment from the arils of Strelitzia nicolai, and performed HPLC-ESMS, UV-visible, 1H NMR and 13C NMR analyses to determine its chemical structure. These data indicated the pigment was bilirubin-IX, an orange-yellow tetrapyrrole previously known only in mammals and some other vertebrates as the breakdown product of heme. Although related tetrapyrroles are ubiquitous throughout the plant kingdom and include vital biosynthetic products such as chlorophyll and phytochromobilin, this is the first report of bilirubin in a plant, and evidence of an additional biosynthetic pathway producing orange coloration in flowers and fruits. ^ Given the unexpected presence of bilirubin, Iexamined the fruits and flowers of twelve additional angiosperm species in diverse orders for the presence of bilirubin using HPLC and LC-MS. Bilirubin was present in ten species from the orders Zingiberales, Arecales, and Myrtales, indicating its wide distribution in the plant kingdom. Bilirubin was present in low concentrations in all species except those within Strelitziaceae. It was present in particularly high concentrations in S. nicolai, S. reginae and P. guyannense, and is thus responsible for producing color in these species. ^ No studies have examined the evolutionary relationship among all species in the family. Thus, I also constructed a molecular phylogeny of the family. This information, combined with further studies on the distribution and synthesis of bilirubin in plants, will provide a basis for understanding the evolutionary history of this pigment in the plant kingdom.^

Theoretical investigations of intercellular communication in the microcirculation: Conducted responses, myoendothelial projections and endothelium derived hyperpolarizing factor

The contractile state of microcirculatory vessels is a major determinant of the blood pressure of the whole systemic circulation. Continuous bi-directional communication exists between the endothelial cells (ECs) and smooth muscle cells (SMCs) that regulates calcium (Ca2+) dynamics in these cells. This study presents theoretical approaches to understand some of the important and currently unresolved microcirculatory phenomena. ^ Agonist induced events at local sites have been shown to spread long distances in the microcirculation. We have developed a multicellular computational model by integrating detailed single EC and SMC models with gap junction and nitric oxide (NO) coupling to understand the mechanisms behind this effect. Simulations suggest that spreading vasodilation mainly occurs through Ca 2+ independent passive conduction of hyperpolarization in RMAs. Model predicts a superior role for intercellular diffusion of inositol (1,4,5)-trisphosphate (IP3) than Ca2+ in modulating the spreading response. ^ Endothelial derived signals are initiated even during vasoconstriction of stimulated SMCs by the movement of Ca2+ and/or IP3 into the EC which provide hyperpolarizing feedback to SMCs to counter the ongoing constriction. Myoendothelial projections (MPs) present in the ECs have been recently proposed to play a role in myoendothelial feedback. We have developed two models using compartmental and 2D finite element methods to examine the role of these MPs by adding a sub compartment in the EC to simulate MP with localization of intermediate conductance calcium activated potassium channels (IKCa) and IP3 receptors (IP 3R). Both models predicted IP3 mediated high Ca2+ gradients in the MP after SMC stimulation with limited global spread. This Ca 2+ transient generated a hyperpolarizing feedback of ∼ 2–3mV. ^ Endothelium derived hyperpolarizing factor (EDHF) is the dominant form of endothelial control of SMC constriction in the microcirculation. A number of factors have been proposed for the role of EDHF but no single pathway is agreed upon. We have examined the potential of myoendothelial gap junctions (MEGJs) and potassium (K+) accumulation as EDHF using two models (compartmental and 2D finite element). An extra compartment is added in SMC to simulate micro domains (MD) which have NaKα2 isoform sodium potassium pumps. Simulations predict that MEGJ coupling is much stronger in producing EDHF than alone K+ accumulation. On the contrary, K+ accumulation can alter other important parameters (EC V m, IKCa current) and inhibit its own release as well as EDHF conduction via MEGJs. The models developed in this study are essential building blocks for future models and provide important insights to the current understanding of myoendothelial feedback and EDHF.^

One-dimensional and two-dimensional polyacrylamide gel electrophoresis: a tool for protein characterisation in aquatic samples

Dissolved organic nitrogen (DON) represents the least understood part of the nitrogen cycle. Due to recent methodological developments, proteins now represent a potentially characterisable fraction of DON at the macromolecular level. We have applied polyacrylamide gel electrophoresis to characterise proteins in samples from a range of aquatic environments in the Everglades National Park, Florida, USA. Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) showed that each sample has a complex and characteristic protein distribution. Some proteins appeared to be common to more than one site, and these might derive from dominant higher plant vegetation. Two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) provided better resolution; however, strong background hindered interpretation. Our results suggest that the two techniques can be used in parallel as a tool for protein characterisation: SDS-PAGE to provide a sample-specific fingerprint and 2D-PAGE to focus on the characterisation of individual protein molecules.

Annotated record of the detailed examination of Mn deposits from the R/V Thomas Washington ROUNDABOUT Cruise stations

The cores described are taken during the R/V Thomas Washington ROUNDABOUT Cruise from May 1988 until March 1989 by the Scripps Institute of Oceanography. A total of 159 cores and dredges were recovered and are available at Scripps Institute of Oceanography for sampling and study.

The SCUBA HAlf degree extragalactic survey - III. Identification of radio and mid-infrared counterparts to submillimetre galaxies

Determining an accurate position for a submillimetre (submm) galaxy (SMG) is the crucial step that enables us to move from the basic properties of an SMG sample - source counts and 2D clustering - to an assessment of their detailed, multiwavelength properties, their contribution to the history of cosmic star formation and their links with present-day galaxy populations. In this paper, we identify robust radio and/or infrared (IR) counterparts, and hence accurate positions, for over two-thirds of the SCUBA HAlf-Degree Extragalactic Survey (SHADES) Source Catalogue, presenting optical, 24-μm and radio images of each SMG. Observed trends in identification rate have given no strong rationale for pruning the sample. Uncertainties in submm position are found to be consistent with theoretical expectations, with no evidence for significant additional sources of error. Employing the submm/radio redshift indicator, via a parametrization appropriate for radio-identified SMGs with spectroscopic redshifts, yields a median redshift of 2.8 for the radio-identified subset of SHADES, somewhat higher than the median spectroscopic redshift. We present a diagnostic colour-colour plot, exploiting Spitzer photometry, in which we identify regions commensurate with SMGs at very high redshift. Finally, we find that significantly more SMGs have multiple robust counterparts than would be expected by chance, indicative of physical associations. These multiple systems are most common amongst the brightest SMGs and are typically separated by 2-6 arcsec, similar to 15-20/sin i kpc at z~ 2, consistent with early bursts seen in merger simulations.

Exceptional activity of gallium(III) chloride and chlorogallate(III) ionic liquids for Baeyer–Villiger oxidation

Baeyer–Villiger oxidation of cyclic ketones, using H2O2 as the oxidising agent, was systematically studied using a range of metal chlorides in different solvents, and in neat chlorogallate(III) ionic liquids. The extremely high activity of GaCl3 in promoting oxidation with H2O2, irrespective of solvent, was reported for the first time. The activity of all other metal chlorides was strongly solvent-dependent. In particular, AlCl3 was very active in a protic solvent (ethanol), and tin chlorides, SnCl4 and SnCl2, were active in aprotic solvents (toluene and dioxane). In order to eliminate the need for volatile organic solvent, a Lewis acidic chlorogallate(III) ionic liquid was used in the place of GaCl3, which afforded typically 89–94% yields of lactones in 1–120 min, at ambient conditions. Raman and 71Ga NMR spectroscopic studies suggest that the active species, in both GaCl3 and chlorogallate(III) ionic liquid systems, are chlorohydroxygallate(III) anions, [GaCl3OH]−, which are the products of partial hydrolysis of GaCl3 and chlorogallate(III) anions; therefore, the presence of water is crucial.

Synthesis and evaluation of temperature- and glucose-sensitive nanoparticles based on phenylboronic acid and N-vinylcaprolactam for insulin delivery

Poly N-vinylcaprolactam-co-acrylamidophenylboronic acid p(NVCL-co-AAPBA) was prepared from N-vinylcaprolactam (NVCL) and 3-acrylamidophenylboronic acid (AAPBA), using 2,2-azobisisobutyronitrile (AIBN) as initiator. The synthesis and structure of the polymer were examined by Fourier Transform infrared spectroscopy (FT-IR) and 1H-NMR. Dynamic light scattering (DLS), lower critical solution temperature (LCST) and transmission electron microscopy (TEM) were utilized to characterize the nanoparticles, CD spectroscopy was used to determine if there were any changes to the conformation of the insulin, and cell and animal toxicity were also investigated. The prepared nanoparticles were found to be monodisperse submicron particles and were glucose- and temperature-sensitive. In addition, the nanoparticles have good insulin-loading characteristics, do not affect the conformation of the insulin and show low-toxicity to cells and animals. These p(NVCL-co-AAPBA) nanoparticles may have some value for insulin or other hypoglycemic protein delivery.

An investigation into the folding and assembly of engineered antibodies and novel antibody formats

Monoclonal antibodies and novel antibody formats are currently one of the principal therapeutic in the biopharmaceutical industry worldwide and are widely used in the treatment of autoimmune diseases and cancer. It is for this reason that the productivity and quality of antibody production requires improvement; specifically investigations into the engineering of antibodies and any issues that may arise from the production of these therapeutics. The work presented in this thesis describes an investigation into the folding and assembly of seven antibodies plus the novel antibody format FabFv. IgG is comprised of two identical HCs and two identical LCs. The folding process of immunoglobulin is controlled by the CH1 domain within the HC. The CH1 domain remains in a disordered state and is sequestered by BiP in the endoplasmic reticulum. Upon the addition of a folded CL domain, BiP is displaced, the CH1 domain is able to fold and the complete IgG protein can then be secreted from the cell. The results presented in this thesis however, have outlined an additional mechanism for the folding of the CH1 domain. We have shown that the CH1 domain is able to fold in the absence of LC resulting in the secretion of HC dimers in a VH dependent manner. The proposed mechanism for the secretion of HC dimers suggests that some VH domains can interact with each other in order to bring the CH1 domains in close proximity to enable folding to occur. As HC dimer secretion is a hindrance in antibody production, this result has highlighted an engineering target to improve antibody yield. Examination of the folding of IgG4 with the variable region A33 has revealed the inability to secrete LC dimers, cleavage of the HC during expression and secretion of HC dimers in the Fab, FabFv and full length forms. The attributes described have also been shown to be variable region dependent. This has introduced a new concept that the variable domain is important in determining the expression and secretion of antibodies and their individual chains. Pulse chase and 2D gel electrophoresis analysis of the novel antibody format FabFv has revealed that the folding and expression of the LC and HC causes multimeric species of FabFv to be secreted, as opposed to the monomeric form which is the desired therapeutic. Our hypothesis is that this process occurs via a LC dependent mechanism. The proposed hypothesis suggests that further engineering to the LC could diminish the formation and secretion of FabFv multimers. The results from these investigations can be applied to increase the productivity of therapeutics and increase the biological understanding of the domain interactions of IgG during folding, assembly and secretion.

A non-contrast self-navigated 3-dimensional MR technique for aortic root and vascular access route assessment in the context of transcatheter aortic valve replacement: proof of concept.

OBJECTIVES: Due to the high prevalence of renal failure in transcatheter aortic valve replacement (TAVR) candidates, a non-contrast MR technique is desirable for pre-procedural planning. We sought to evaluate the feasibility of a novel, non-contrast, free-breathing, self-navigated three-dimensional (SN3D) MR sequence for imaging the aorta from its root to the iliofemoral run-off in comparison to non-contrast two-dimensional-balanced steady-state free-precession (2D-bSSFP) imaging. METHODS: SN3D [field of view (FOV), 220-370 mm(3); slice thickness, 1.15 mm; repetition/echo time (TR/TE), 3.1/1.5 ms; and flip angle, 115°] and 2D-bSSFP acquisitions (FOV, 340 mm; slice thickness, 6 mm; TR/TE, 2.3/1.1 ms; flip angle, 77°) were performed in 10 healthy subjects (all male; mean age, 30.3 ± 4.3 yrs) using a 1.5-T MRI system. Aortic root measurements and qualitative image ratings (four-point Likert-scale) were compared. RESULTS: The mean effective aortic annulus diameter was similar for 2D-bSSFP and SN3D (26.7 ± 0.7 vs. 26.1 ± 0.9 mm, p = 0.23). The mean image quality of 2D-bSSFP (4; IQR 3-4) was rated slightly higher (p = 0.03) than SN3D (3; IQR 2-4). The mean total acquisition time for SN3D imaging was 12.8 ± 2.4 min. CONCLUSIONS: Our results suggest that a novel SN3D sequence allows rapid, free-breathing assessment of the aortic root and the aortoiliofemoral system without administration of contrast medium. KEY POINTS: • The prevalence of renal failure is high among TAVR candidates. • Non-contrast 3D MR angiography allows for TAVR procedure planning. • The self-navigated sequence provides a significantly reduced scanning time.

Development of Novel Time-Domain Electromagnetic Methods for Offshore Groundwater Studies: A Data Application from Bat Yam, Israel

Recent marine long-offset transient electromagnetic (LOTEM) measurements yielded the offshore delineation of a fresh groundwater body beneath the seafloor in the region of Bat Yam, Israel. The LOTEM application was effective in detecting this freshwater body underneath the Mediterranean Sea and allowed an estimation of its seaward extent. However, the measured data set was insufficient to understand the hydrogeological configuration and mechanism controlling the occurrence of this fresh groundwater discovery. Especially the lateral geometry of the freshwater boundary, important for the hydrogeological modelling, could not be resolved. Without such an understanding, a rational management of this unexploited groundwater reservoir is not possible. Two new high-resolution marine time-domain electromagnetic methods are theoretically developed to derive the hydrogeological structure of the western aquifer boundary. The first is called Circular Electric Dipole (CED). It is the land-based analogous of the Vertical Electric Dipole (VED), which is commonly applied to detect resistive structures in the subsurface. Although the CED shows exceptional detectability characteristics in the step-off signal towards the sub-seafloor freshwater body, an actual application was not carried out in the extent of this study. It was found that the method suffers from an insufficient signal strength to adequately delineate the resistive aquifer under realistic noise conditions. Moreover, modelling studies demonstrated that severe signal distortions are caused by the slightest geometrical inaccuracies. As a result, a successful application of CED in Israel proved to be rather doubtful. A second method called Differential Electric Dipole (DED) is developed as an alternative to the intended CED method. Compared to the conventional marine time-domain electromagnetic system that commonly applies a horizontal electric dipole transmitter, the DED is composed of two horizontal electric dipoles in an in-line configuration that share a common central electrode. Theoretically, DED has similar detectability/resolution characteristics compared to the conventional LOTEM system. However, the superior lateral resolution towards multi-dimensional resistivity structures make an application desirable. Furthermore, the method is less susceptible towards geometrical errors making an application in Israel feasible. In the extent of this thesis, the novel marine DED method is substantiated using several one-dimensional (1D) and multi-dimensional (2D/3D) modelling studies. The main emphasis lies on the application in Israel. Preliminary resistivity models are derived from the previous marine LOTEM measurement and tested for a DED application. The DED method is effective in locating the two-dimensional resistivity structure at the western aquifer boundary. Moreover, a prediction regarding the hydrogeological boundary conditions are feasible, provided a brackish water zone exists at the head of the interface. A seafloor-based DED transmitter/receiver system is designed and built at the Institute of Geophysics and Meteorology at the University of Cologne. The first DED measurements were carried out in Israel in April 2016. The acquired data set is the first of its kind. The measured data is processed and subsequently interpreted using 1D inversion. The intended aim of interpreting both step-on and step-off signals failed, due to the insufficient data quality of the latter. Yet, the 1D inversion models of the DED step-on signals clearly detect the freshwater body for receivers located close to the Israeli coast. Additionally, a lateral resistivity contrast is observable in the 1D inversion models that allow to constrain the seaward extent of this freshwater body. A large-scale 2D modelling study followed the 1D interpretation. In total, 425 600 forward calculations are conducted to find a sub-seafloor resistivity distribution that adequately explains the measured data. The results indicate that the western aquifer boundary is located at 3600 m - 3700 m before the coast. Moreover, a brackish water zone of 3 Omega*m to 5 Omega*m with a lateral extent of less than 300 m is likely located at the head of the freshwater aquifer. Based on these results, it is predicted that the sub-seafloor freshwater body is indeed open to the sea and may be vulnerable to seawater intrusion.

Perceptive variation of carboxylate ligand and probing the influence of substitution pattern on the structure of mono- and di-butylstannoxane complexes

By reacting 2- and 3-aminobenzoic acids (HL1 and HL2, respectively), as well as 2-, 3- and 4-((E)-2-[4-(dimethylamino)phenyl]diazenyl)benzoic acids (HL3, HL4 and HL5, in this order) with a n-butyltin(IV) source [ n BuSn(O)OH or n Bu2SnO], the drum-type butylstannoxane complexes of general composition [ n Bu6Sn6O6(L n )6] [L n =L1 (1), L2 (2) and L3 (3)] and the ladder-type compounds [ n Bu8Sn4O2(L n )4] [L n =L3 (5), L4 (6) and L5 (7)] were obtained and fully characterized. By reacting 1 with 2-((E)-[4-(dimethylamino)benzylidene]amino)benzoic acid (HL6), a co-crystal (4) was achieved which comprises the metal complex aggregate found in 1 and the neutral HL6 molecule. The solution properties of the compounds were assessed from 1H and 13C NMR studies and, for the metal complexes, also from 119Sn NMR. The molecular structures of 1, 2, 4-7 were confirmed by single-crystal X-ray diffraction. Compounds 1-3 and the complex moiety of 4 display hexameric Sn6O6 clusters with drum-like structures, but 5-7 reveal Sn4O2 cores with ladder-type structural motifs. Besides the observed relationship between the ligand N-functional group and obtained (drum- or ladder-type) assemblies, the relative position of the carboxylate group in the ligand itself influences its coplanarity.

High-precision force sensing using a single trapped ion

We introduce quantum sensing schemes for measuring very weak forces with a single trapped ion. They use the spin-motional coupling induced by the laser-ion interaction to transfer the relevant force information to the spin-degree of freedom. Therefore, the force estimation is carried out simply by observing the Ramsey-type oscillations of the ion spin states. Three quantum probes are considered, which are represented by systems obeying the Jaynes-Cummings, quantum Rabi (in 1D) and Jahn-Teller (in 2D) models. By using dynamical decoupling schemes in the Jaynes-Cummings and Jahn-Teller models, our force sensing protocols can be made robust to the spin dephasing caused by the thermal and magnetic field fluctuations. In the quantum-Rabi probe, the residual spin-phonon coupling vanishes, which makes this sensing protocol naturally robust to thermally-induced spin dephasing. We show that the proposed techniques can be used to sense the axial and transverse components of the force with a sensitivity beyond the yN/\wurzel{Hz}range, i.e. in the xN/\wurzel{Hz}(xennonewton, 10^−27). The Jahn-Teller protocol, in particular, can be used to implement a two-channel vector spectrum analyzer for measuring ultra-low voltages.