A Step Towards Quantitative Lipoprotein Density Profiling Analysis: Applied Rayleigh Scattering

Ultracentrifugation and imaging techniques of human blood serum are precise and information-rich methods for obtaining information about an individual’s lipoprotein particle content. The information derived from lipoprotein separations via an ultracentrifuge plays a key role in the area of preventative medicine in regards to atherosclerosis. Two of the most critical lipoprotein characteristics, diameter and density, are well preserved with the proper isopycnic gradient. Currently, lipoprotein particles are stained, ultracentrifuged, and profiled through image analysis. This particular technique is helpful in determining particle density and can be correlated loosely with particle concentration. The need to completely quantify lipoprotein concentrations is imperative in assessing risk factors accurately.

Light scattering techniques, primarily Rayleigh scattering, are applied to density separated serum samples in resulting in improved qualitative data with progress in quantitative measurements through imaging alone. The Rayleigh theory dictates that a particle’s scattered intensity is based upon the incident intensity, the particle’s diameter, and the particle’s concentration when strict criteria are met within the sample and imaging apparatus. Applying this innovative imaging technique of Rayleigh scattering to ultracentrifuge tubes containing separated lipoproteins, particle concentrations at differing diameters can be calculated.

This thesis primarily goes through the time consuming task of optimizing the innovative Rayleigh scattering system so that correct quantitative estimations can be performed. Constrained by Rayleigh theory and system limitations, lipoproteins of 15 nm to 35 nm are focused upon. By doing so, previously disguised data in regards to lipoprotein subclasses is exposed. Lipoprotein diameters are estimated from Rayleigh imaged serum profiles and the estimations are confirmed through secondary size analysis achieved by dynamic light scattering instrumentation. In addition to Rayleigh optimization, a strategy for quantifying the ultracentrifuged lipoprotein particles using the recently applied scattering technique is explained in detail providing a foundation for further research. In regards to all feasibility studies presented within this thesis, much success was achieved in furthering quantitation efforts in lipoprotein density profiling.

Michael Nowlin, Texas A&M Univeristy, ©2006

Water disperses and dissolves many compounds. One such ionic species dissolvable in water is sodium chloride that by virtue of the dielectric properties of water can easily be separated into individually charged atoms, which can be hydrated (solvated). The dielectric constant (D) is inversely proportional to the distance between the ions while the attractive force is proportional to the ion charge. Thus water (D= 80) easily dissolves sodium chloride while the salt is insoluble in benzene (D=2.3). Another type of dispersal involves hydrogen bonding of water molecules to polar functional groups in the chemical structure of sugars and alcohols. Changes in the structure of water by use of surfactants or salts can be employed to “dissolve” proteins such as antibodies, collagen, some hormones, hemoglobin and myosin. Dispersal of the proteins generally depends upon the protein composition. Composition dictates which chemicals are necessary to reduce the forces of attraction that prevents solubility. A last type of dispersal includes biological chemicals. Lipids as a class, depending upon their structure, will dissolve in polar or non-polar organic solvents, but generally, this class of compounds is not soluble in water. Water insolubility allows lipids to enter special chemical bonding, which produces structures such as blood lipoproteins, cell membranes, micelles and liposomes. High-Concentration Liposome and Micelle Suspensions: Measurement by Dynamic Light Scattering describes lipoproteins and micelles with particular attention to particle size measurement of liposomes in either dilute or concentrated aqueous environments.
 

Transitional properties of starch colloid with particle size reduction from micro- to nanometer

High-pressure homogenization was used to disperse starch particles in water and reduce the size from micro- to nanometer. The resultant starch colloids were characterized by particle morphology, mean size, size distribution, and zeta potential. Starch slurries were transformed from a mixture containing sediment, dispersion, and sol, to gel as a result of reduction of the particle size from 3-6µm to 10-20nm under a pressure of 207MPa. Furthermore, this process led to the transition of fluid properties without affecting the crystal structure and thermal stability of starch granules. Viscosity of the colloids increased with an increased number of homogenization passes, accompanied by a decreased particle size, narrower particle size distribution (PSD), and an increased absolute zeta potential, indicating the formation of a suspension or stable gel composed of nanoparticles. Lognormal and two other mathematical functions were established to describe the PSDs and their relationship to the homogenization passes. Hence, an environmentally friendly means of producing starch-based nanoparticles or nanogels with high yields, and predictable size and viscosity properties was presented. ©2009 Journal of Colliod & Interface Science

Liu D, Wu Q, Chen H, Chang PR.
School of Renewable Natural Resources, Louisiana State University Agricultural Center, Baton Rouge, LA 70803, USA

This Princeton research presentation describes a project to study the behavior and properties of nanoparticles made of block copolymers, find the optimal concentrations of fluorescent dyes in the nanoparticles, and determine the stability of various compounds encapsulated in nanoparticles.  Lymphocyte cells were incubated for 90 min in a solution containing PCL-PEG micelle nanoparticles of 70 nm diameter encapsulating Nile Red. Cells were then fixed and DNA was stained using Draq4.  Microtrac dynamic light scattering (DLS) equipment was used to determine whether nanoparticles are aggregating over time.  The study determined that Nile red and the AMC-vitamin E conjugate are stable in nanoparticles.  © 2006 Princeton

Katie Chin, Faculty advisor: Prof. Prud’homme, Mentor: Marian Gindy, Princeton University, NJ, USA

Striking differences in Aloe vera gel carbohydrate composition, molecular weight and particle size distributions following processing will not be addressed by dietary supplement GMPs

Abstract
Background Aloe vera gel (AVG) is a popular dietary supplement ingredient. The International Aloe Science Council (IASC) standards to confirm identity and quality of aloe products are, respectively, polysaccharide acetylation patterns and organic acid profiles. Certain polysaccharides contribute to biological activities of numerous medicinal plants, including aloe vera (AV). The quantity and quality of polysaccharides in AV supplements thus must be better understood.
Objective Determine the effect of processing methods on AVG carbohydrate composition (CC), molecular weight (MW) and particle size (PS) distributions.
Methods 3 leaves from plants grown commercially were used to prepare native gel (G) and four blended samples: not filtered (NF), filtered (F), not filtered/ freeze dried (NF/D), filtered/freeze dried (F/D). Also analyzed were two commercial AVG powders from IASC-certified vendors (C1, C2). CC was determined for all samples by HPAEC. Additional tests were performed on all samples except G: MW distributions by HPLC-SEC; PS distributions using a Microtrac® PS Analyzer.
Results CC: 1) laboratory There was variability in major sugar content between leaves: NF samples from leaf 2 contained 45.02% glucose and 38.21% mannose; samples from leaf 3 contained 28.9% glucose and 55.2% mannose. 24% of G was glucuronic acid; no other laboratory preparation contained this sugar. Blending impacted CC: compared with NF, G contained substantially more glucuronic acid, arabinose, rhamnose, and fucose and less mannose and glucose. Fructose and glucosamine, present in small quantities in NF, were not detected in G. Filtering impacted CC: compared with NF, F contained more mannose and less glucose and galactose. Filtering removed xylose, fucose and rhamnose. Freeze drying impacted CC: increasing the percentage of glucose and decreasing the percentage of mannose in filtered and not filtered samples. 2) commercial Compared with C2, C1 contained substantially more glucuronic acid, mannose, galacturonic acid and galactose and less glucose. MW: 1) laboratory Filtering did not impact MW distributions. MW distributions of not filtered samples were affected by freeze drying: freeze dried samples contained a higher percentage of >10,000 - <100,000 MW polysaccharides and a lower percentage of <10,000 MW polysaccharides. 2) commercial C1 polysaccharides were significantly higher MW (C1: 23.3% >800,000; C2: 40.1% <10,000). PS: 1) laboratory Filtration substantially reduced sample particle sizes. Freeze drying reduced the particle size of not filtered samples. For filtered samples, freeze drying virtually eliminated 1.94-11 μ particles and substantially increased the percentage of 249-592 μ particles, substantially decreased 592-995.5 μ particles and eliminated 995-2000 μ particles. 2) commercial C1 had a higher percentage of large particles.
Conclusions We report that processing significantly impacts AVG carbohydrate composition, molecular weight distribution and particle size distribution. These changes—which likely affect product quality and efficacy—suggest that dietary supplement GMPs may not adequately address these issues. ©2008 Mannatech 

Christy Duncan, BS; Jane Ramberg, MS; Robert Sinnott, MNS, PhD

Iron oxide nanoparticles induce human microvascular endothelial cell permeability through reactive oxygen species production and microtubule remodeling

Abstract
Background Engineered iron nanoparticles are being explored for the development of biomedical applications and many other industry purposes. However, to date little is known concerning the precise mechanisms of translocation of iron nanoparticles into targeted tissues and organs from blood circulation, as well as the underlying implications of potential harmful health effects in human.
Results The confocal microscopy imaging analysis demonstrates that exposure to engineered iron nanoparticles induces an increase in cell permeability in human microvascular endothelial cells. Our studies further reveal iron nanoparticles enhance the permeability through the production of reactive oxygen species (ROS) and the stabilization of microtubules. We also showed Akt/GSK-3β signaling pathways are involved in iron nanoparticle-induced cell permeability. The inhibition of ROS demonstrate ROS play a major role in regulating Akt/GSK-3β – mediated cell permeability upon iron nanoparticle exposure. These results provide new insights into the bioreactivity of engineered iron nanoparticles which can inform potential applications in medical imaging or drug delivery.
Conclusion Our results indicate that exposure to iron nanoparticles induces an increase in endothelial cell permeability through ROS oxidative stress-modulated microtubule remodeling. The findings from this study provide new understandings on the effects of nanoparticles on vascular transport of macromolecules and drugs.©2009 Apopa et al; licensee BioMed Central Ltd.

Patrick L Apopa^1,2, Yong Qian¹, Rong Shao³, Nancy Lan Guo⁴, Diane Schwegler-Berry¹, Maricica Pacurari¹, Dale Porter^1,5, Xianglin Shi¹, Val Vallyathan¹, Vincent Castranova¹ and Daniel C Flynn^6
1The Pathology and Physiology Research Branch, Health Effects Laboratory Division, National Institute for Occupational Safety and Health, Morgantown, WV 26505, USA
²MBR Cancer Center, School of Medicine, West Virginia University, Morgantown, WV 26506-9300, USA
³Pioneer Valley Life Sciences Institute, Baystate Medical Center/University of Massachusetts at Amherst, Springfield, MA 01107, USA
⁴MBR Cancer Center/Department of Community Medicine, School of Medicine, West Virginia University, Morgantown, WV 26506-9300, USA
⁵Department of Physiology and Pharmacology, School of Medicine, West Virginia University, Morgantown, WV 26506, USA 
⁶The Commonwealth Medical College, Scranton, PA 18510, USA

Ultrafine Hydrogel Nanoparticles: Synthetic Approach and Therapeutic Application in Living Cells

Abstract
Licensed to kill: A method for obtaining ultrafine hydrophilic polyacrylamide-based nanoparticles that encapsulate meta-tetra(hydroxyphenyl) chlorin (mTHPC) has been developed. Rat C6 glioma cells exposed to infrared light were killed (red) in the presence of the mTHPC-encapsulating nanoparticles while the cells without exposure to light were still alive (green). ©2007 Wiley-VCH
Keywords
antitumor agents • drug delivery • hydrogels • nanostructures • photodynamic therapy

De Gao, Dr. ¹, Hao Xu, Dr. ², Martin A. Philbert, Prof. ², Raoul Kopelman, Prof. ^1
1Department of Chemistry, University of Michigan, Ann Arbor, MI 48109-1055, USA, Fax: (+1) 734-936-2778
²Department of Environmental Health Sciences, University of Michigan, Ann Arbor, MI 48109-2029, USA