Heiko Schiffter
University of Oxford
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Featured researches published by Heiko Schiffter.
Journal of Pharmaceutical and Biomedical Analysis | 2010
Sebastian Vonhoff; Jamie Condliffe; Heiko Schiffter
The aim of this study was to develop a quick and objective method for the determination of changes in protein secondary structure by Fourier transform infrared spectroscopy (FTIR). Structural shifts from native regions (alpha-helix, intramolecular beta-sheet) to aggregated strands (intermolecular beta-sheet) were used to evaluate protein damage. FTIR spectra of 16 different proteins were recorded and quantified by peak fitting of the non-deconvolved and baseline corrected amide I bands. The resulting percentile secondary structures were correlated with the shape and intensity of the area normalized amide I bands using an interval partial least squares algorithm (iPLS). Structural elements were focused on the following regions: alpha-helix 1660-1650 cm(-1), intramolecular beta-sheet 1695-1683 cm(-1) and 1644-1620 cm(-1), intermolecular beta-sheet 1620-1595 cm(-1). Three calibration curves were created from the data sets. Calculated alpha-helix content ranged from 0% to 79.59%, intramolecular beta-sheet from 10.64% to 63.89% and intermolecular beta-sheet from 0.23% to 9.70%. The linear relationship between actual values (as determined by peak fitting) and calculated values was evaluated by correlation coefficient and root mean square error of calibration while cross-validation was performed to detect possible outliers. Results were verified by including two proteins as validation standards and comparing the calculated values to peak fitting and X-ray data. Structural changes of human serum albumin (HSA) due to elevated temperatures and the fibrillation of glucagon were quantified by calibration curve analysis. Performance and reliability of the iPLS algorithm were evaluated by comparing calculated secondary structure elements with results from peak fitting and circular dichroism. Different methods for the determination of secondary structure gave slightly different results but overall tendencies concurred. Additionally, formation of HSA aggregates could be linked to increasing beta-sheet content by comparing SEC-HPLC and turbidity analysis with results from the FTIR calibration curves. In summary, quantification of the alpha-helix to beta-sheet transition by iPLS analysis proves to be a feasible and objective way for the determination of damage to protein secondary structure.
Journal of the Royal Society Interface | 2010
Heiko Schiffter; Jamie Condliffe; Sebastian Vonhoff
The feasibility of preparing microparticles with high insulin loading suitable for needle-free ballistic drug delivery by spray-freeze-drying (SFD) was examined in this study. The aim was to manufacture dense, robust particles with a diameter of around 50 µm, a narrow size distribution and a high content of insulin. Atomization using ultrasound atomizers showed improved handling of small liquid quantities as well as narrower droplet size distributions over conventional two-fluid nozzle atomization. Insulin nanoparticles were produced by SFD from solutions with a low solid content (<10 mg ml−1) and subsequent ultra-turrax homogenization. To prepare particles for needle-free ballistic injection, the insulin nanoparticles were suspended in matrix formulations with a high excipient content (>300 mg ml−1) consisting of trehalose, mannitol, dextran (10 kDa) and dextran (150 kDa) (abbreviated to TMDD) in order to maximize particle robustness and density after SFD. With the increase in insulin content, the viscosity of the nanosuspensions increased. Liquid atomization was possible up to a maximum of 250 mg of nano-insulin suspended in a 1.0 g matrix. However, if a narrow size distribution with a good correlation between theoretical and measurable insulin content was desired, no more than 150 mg nano-insulin could be suspended per gram of matrix formulation. Particles were examined by laser light diffraction, scanning electron microscopy and tap density testing. Insulin stability was assessed using size exclusion chromatography (SEC), reverse phase chromatography and Fourier transform infrared (FTIR) spectroscopy. Densification of the particles could be achieved during primary drying if the product temperature (Tprod) exceeded the glass transition temperature of the freeze concentrate (Tg′) of −29.4°C for TMDD (3∶3∶3∶1) formulations. Particles showed a collapsed and wrinkled morphology owing to viscous flow of the freeze concentrate. With increasing insulin loading, the d (v, 0.5) of the SFD powders increased and particle size distributions got wider. Insulin showed a good stability during the particle formation process with a maximum decrease in insulin monomer of only 0.123 per cent after SFD. In accordance with the SEC data, FTIR analysis showed only a small increase in the intermolecular β-sheet of 0.4 per cent after SFD. The good physical stability of the polydisperse particles made them suitable for ballistic injection into tissue-mimicking agar hydrogels, showing a mean penetration depth of 251.3 ± 114.7 µm.
Journal of Pharmaceutical Sciences | 2014
Sunali Bhatnagar; Heiko Schiffter; Constantin C. Coussios
Ultrasound (US) exposure of soft tissues, such as the skin, has been shown to increase permeability, enhancing the passage of drug molecules via passive processes such as diffusion. However, US regimes have not been exploited to enhance active convective transport of drug molecules from a donor layer, such as a gel, into another medium. A layered tissue-mimicking material (TMM) was used as a model for a drug donor layer and underlying soft tissue to test penetration of agents in response to a range of US parameters. Influence of agent molecular mass (3-2000 kDa), US frequency (0.256/1.1 MHz) and US pressure (0-10 MPa) on transport was characterised. Agents of four different molecular sizes were embedded within the TMM with or without cavitation nuclei (CN) and US applied to achieve inertial cavitation. Post-insonation, samples were analysed to determine the concentration and penetration distance of agent transported. US exposure substantially enhanced transport. At both US frequencies, enhancement of transport was significantly higher (p < 0.05) above the cavitation threshold, and CN reduced the pressure at which cavitation, and therefore transport, was achieved. Acoustic cavitation activity and related phenomena was the predominant transport mechanism, and addition of CN significantly enhanced transport within a range of clinically applicable acoustic pressures.
Modern Drying Technology, Set | 2014
Mirko Peglow; Thomas Metzger; Geoffrey Lee; Heiko Schiffter; Robert Hampel; Stefan Heinrich; Evangelos Tsotsas
Knowledge of the amount of moisture contained in particles before, during and after drying is an elementary requirement in drying technology. This moisture amount, for example for quality control, can easily be determined on more or less large samples of particles by weighing. However, other tasks, such as the design of industrial convective dryers impose much more serious challenges. To reliably design a convective industrial dryer, kinetic data referring to the specific product are necessary. Since information on drying kinetics is usually not available, it has to be gained experimentally. In this case, it is not sufficient to measure the mass of moisture contained in the product at a certain point of time, but the change of this mass with time has to be resolved as accurately as possible. Additionally, the change of mass with time must refer to the single particle. The reason for this second requirement is that gas conditions change in particle systems. This results – even if every particle has exactly the sameproperties and the particles are perfectlymixed – inmore or less significant differences between the drying kinetics of the entire particle system and the drying kinetics of the single particle. Experimental techniques for the determination of single particle drying kinetics will be discussed in Section 1.2, with emphasis on the magnetic suspension balance. On the other hand, it is evident thatmeasurements on single particleswill give only very low signals and, hence, be confronted with severe limitations of resolution and accuracy, evenwhenusing very sensitive instruments. This is especially true for small particles (powdery products). Therefore, we may be forced to investigate drying kinetics of an entire particle system such as a packed orfluidized bed. This is typically done by measuring gas humidity at the outlet of the dryer, instead of solids moisture content. It should be borne in mind that the results of such indirect measurements must be scaled down to the single particle by an appropriate model in order to obtain unbiased access to product-specific drying kinetics. Important instruments for
8TH INTERNATIONAL SYMPOSIUM ON THERAPEUTIC ULTRASOUND | 2009
Sacha D. Nandlall; Manish Arora; Heiko Schiffter; Constantin C. Coussios
Thermal dose has been proposed for various hyperthermic cancer treatment modalities as a measure of heat‐induced tissue damage. However, the applicability of current thermal dose metrics to tissue is not well understood, particularly at the temperatures and rates of heating relevant to ablative cancer therapy using High‐Intensity Focussed Ultrasound (HIFU). In this work, we assess whether the most widely employed thermal dose metric, Cumulative Equivalent Minutes (CEM), can adequately quantify heat‐induced denaturation in a tissue‐mimicking material (phantom) consisting of Bovine Serum Albumin (BSA) proteins embedded in a polyacrylamide matrix. The phantom is exposed to various temperature profiles and imaged under controlled lighting conditions against a black background as it denatures and becomes progressively more opaque. Under the assumption that the mean backscattered luminous intensity provides a good measure of the extent of BSA denaturation, we establish a relationship between the amount of thermal damage caused to the phantom, exposure time, and temperature. We demonstrate that, for monotonically increasing and bounded temperature profiles, the maximal degree to which the phantom can denature is dependent on the peak temperature it reaches, irrespective of exposure duration. We also show that when the CEM is computed using the commonly employed piecewise‐constant approximation of the parameter R, the CEM values corresponding to the same degree of damage delivered using different temperature profiles do not agree well with each other in general.
Expert Review of Vaccines | 2013
Nikolas T. Weissmueller; Heiko Schiffter; Andrew J. Pollard
The central importance for global public health policy of delivering life-saving vaccines for all children makes the development of efficacious and safe needle-free alternatives to hypodermic needles, preferably in a thermostable form, a matter of pressing urgency. This paper comprehensively reviews past in vivo studies on intradermal powder immunization with vaccine formulations that do not require refrigeration. Particular emphasis is given to the immune response in relation to antigen adjuvantation. While needle-free intradermal delivery of vaccines induces a predominantly Th2-type immune response, adjuvants powerfully enhance and modulate the magnitude and nature of the elicited immune response at various effector sites.
11TH INTERNATIONAL SYMPOSIUM ON THERAPEUTIC ULTRASOUND | 2012
Sarah J. Wagstaffe; Heiko Schiffter; Manish Arora; Constantin C. Coussios
Reliable instigation of cavitation in-vivo during ultrasound therapy is notoriously difficult. Lowering the peak rarefractional pressure required to initiate cavitation (the cavitation threshold) has been previously addressed using ultrasound contrast agents in the form of encapsulated stabilized micron scale bubbles. These agents lack stability and are generally too large to extravasate into tumours and other target tissues. Solid nanoparticles are proposed as novel cavitation nucleation agents, which overcome these limitations. Such agents are manufactured to achieve high surface roughness and hydrophobicity, facilitating air entrapment upon drying, thus harboring an abundance of cavitation nucleation sites. These nanoparticulate nucleating agents have been found to lower the cavitation threshold significantly in aqueous biological media, enabling reproducible cavitation activity during repeated exposure to therapeutic ultrasound. This paper investigates the engineering of core-shell nanoparticles and e...
International Journal of Hyperthermia | 2010
Sacha D. Nandlall; Heiko Schiffter; Sebastian Vonhoff; Miriam Bazan-Peregrino; Manish Arora; Constantin C. Coussios
Purpose: In controlled laboratory studies of hyperthermia and thermal ablation, translucent hydrogels containing bovine serum albumin (BSA) are often employed as tissue-mimicking materials due to the change in their opacity that takes place as they accumulate heat damage. In this work we demonstrate the biological relevance of this optical metric of thermal damage, as well as establish the physical mechanisms that link it with quantifiable damage to the proteins embedded in the gel. Materials and methods: We applied Fourier transform infrared (FTIR) spectroscopy, turbidity analysis using ultraviolet-visible (UV/VIS) spectroscopy, and size exclusion chromatography (SEC) to samples of heat-treated, aqueous bovine serum albumin (BSA). We also measured the rates of survival in heated suspensions of breast cancer cells using a colorimetric assay. Results: Using FTIR spectroscopy and SEC, we show that the intermolecular β-sheet content of the protein ensemble rises in heat treatments above 60°C, which causes aggregate formation. Furthermore, by applying UV/VIS spectroscopy we demonstrate that the opacity of the hydrogel increases past 60°C due to the formation of insoluble protein aggregates that scatter incident light. Finally, we illustrate that the viability of human breast cancer cells follows a similar trend to measurements of BSA polyacrylamide hydrogel opacity at various temperatures from 37°C to 90°C. Conclusions: Our work establishes a causal link between the degree of BSA denaturation in hydrogel and the opacity of the medium. Furthermore, our results demonstrate that BSA hydrogels provide a simple physical model for quantifying biologically relevant heat damage in real time during controlled laboratory studies of hyperthermia and thermal ablation.
Journal of the Acoustical Society of America | 2010
Jamie Condliffe; Heiko Schiffter; Robin O. Cleveland; Constantin C. Coussios
Needle-free injection is a novel technique for transdermal drug and vaccine delivery, the efficacy of which depends on the number density and mean penetration depth of particles beneath the skin. To date, these parameters have been assessed optically, which is time-consuming and unsuitable for use in vivo. The present work describes the development of a scanning acoustic microscopy technique to map and size particle distributions following injection. Drug particles were modeled using a polydisperse distribution of polystyrene spheres, mean diameter 30.0 mum, and standard deviation 16.7 mum, injected into agar-based tissue-mimicking material, and later, as polydisperse stainless steel spheres, mean diameter 46.0 mum, and standard deviation 13.0 mum, injected both into agar and into porcine skin. A focused broadband immersion transducer (10-75 MHz), driven in pulse-echo mode, was scanned over the surface of the injected samples. Recorded echo signals were post-processed to deduce particle penetration depth (30-300 mum). Furthermore, post-injection size distribution of the spheres was calculated using a novel, automated spectral analysis technique. Experimental results were validated optically and found to predict penetration depth and particle size accurately. The availability of simultaneous particle penetration depth and particle size information makes it possible for the first time to optimize particle design for specific drug delivery applications.
PLOS ONE | 2017
Nikolas T. Weissmueller; Leanne Marsay; Heiko Schiffter; Robert Carlisle; Christine S. Rollier; Robert K. Prud’homme; Andrew J. Pollard
Powder-injectors use gas propulsion to deposit lyophilised drug or vaccine particles in the epidermal and sub epidermal layers of the skin. We prepared dry-powder (Tg = 45.2 ± 0.5°C) microparticles (58.1 μm) of a MenY-CRM197 glyconjugate vaccine (0.5% wt.) for intradermal needle-free powder injection (NFPI). SFD used ultrasound atomisation of the liquid vaccine-containing excipient feed, followed by lyophilisation above the glass transition temperature (Tg’ = − 29.9 ± 0.3°C). This resulted in robust particles (density~ 0.53 ±0.09 g/cm3) with a narrow volume size distribution (mean diameter 58.1 μm, and span = 1.2), and an impact parameter (ρvr ~ 11.5 kg/m·s) sufficient to breach the Stratum corneum (sc). The trehalose, manitol, dextran (10 kDa), dextran (150 kDa) formulation, or TMDD (3:3:3:1), protected the MenY-CRM197 glyconjugate during SFD with minimal loss, no detectable chemical degradation or physical aggregation. In a capsular group Y Neisseria meningitidis serum bactericidal assay (SBA) with human serum complement, the needle free vaccine, which contained no alum adjuvant, induced functional protective antibody responses in vivo of similar magnitude to the conventional vaccine injected by hypodermic needle and syringe and containing alum adjuvant. These results demonstrate that needle-free vaccination is both technically and immunologically valid, and could be considered for vaccines in development.