Determining the critical micelle concentration of a novel lipid-lowering agent, disodium ascorbyl phytostanyl phosphate (FM-VP4), using a fluorescence depolarization procedure

The objective of this study was to determine the critical micelle concentration (CMC) of a novel water-soluble plant sterol derivative (FM-VP4) using a fluorescence depolarization method. The CMC was determined by 1,6-diphenyl-1,3,5-hexatriene (DPH) fluorescence depolarization. Test solutions of various concentrations of sodium dodecylsulphate (SDS) as a positive control or FM-VP4 in water were spiked with 2 µL of 4 mM DPH in tetrahydrofuran (THF) and left overnight to equilibrate in a dark chamber. Fluorescence of each solution was measured at room temperature using a Perseptive Biosystems Cytofluor Series 4000 multi-well plate reader. Fluorescence intensity increases as DPH is incorporated into the hydrophobic core of micelles. Thus, the CMC is the value at which an abrupt increase in intensity is observed. These points were observed at 8 mM and 0.014 mM for SDS and FM-VP4, respectively. Sodium dodecylsulphate was used as a positive control and supports the validity of our results, as the literature values of SDS are reported to be between 8-8.3 mM. The CMC of FM-VP4 is reported to be 0.014 mM.     

Development and characterization of liposomal disodium ascorbyl phytostanyl phosphates (FM-VP4)

The specific objectives of this project were (1) to develop liposomal disodium ascorbyl phytostanyl phosphate (FM-VP4) formulations, (2) to develop a liquid chromatography/mass spectrometry/mass spectrometry (LC/MS/MS) assay for quantification of FM-VP4 in liposomal formulations and plasma sample, and (3) to characterize liposomal FM-VP4 formulations by finding optimal drug-to-lipid ratios and determining the degradation of FM-VP4 in liposomes. Section 2 describes an LC/MS/MS assay developed for the identification and quantification of FM-VP4 in liposomal formulations to provide estimates of drug concentrations and encapsulation efficiency. The extra step of removing plasma proteins prior to LC/MS/MS assay yields an analysis of FM-VP4 in plasma samples. Section 3 describes experiments designed to find the optimal drug-to-lipid ratio for liposomal FM-VP4 formulations by comparing encapsulation efficiencies and varying the lipid compositions. Additionally, this section details our degradation studies to determine if liposomes have any protective effects on FM-VP4; these studies tested various lipid compositions at 37°C in rabbit plasma. The mechanism of how FM-VP4 lowers low-density lipoprotein (LDL) cholesterol and total cholesterol levels in various animal models is presently unknown. However, before the mechanism of action could be studied, FM-VP4 first had to be delivered efficiently into plasma or cultured cell. The low systemic bioavailability and cellular uptake of FM-VP4 further suggested the importance of finding an efficient delivery vehicle for this drug. This project proposed a framework for such delivery and paves the way for further investigation into how FM-VP4 works in vivo and in vitro.     

EFFECTS OF INTERPARTICLE INTERACTIONS ON STABILITY, AGGREGATION AND SEDIMENTATION IN COLLOIDAL SUSPENSIONS

Relations between interparticle effective interactions, structure formation, stability and sedimentation for a colloidal system are presented in this paper. For a binary mixture of large and small particles, the potential of the mean forces between large particles is obtained from the Ornstein-Zernike equation. We incorporated the small particles in our numerical simulations by using this potential of the mean force as the interparticle effective interaction. Our numerical results reveal the phenomenon of strong particle aggregation due to the attractive depletion force exerted by small particles. In the absence of the effect of gravity, this aggregation can result in flocculation and the formation of particle clusters thereby forming “void” structures, while under the influence of gravity, the aggregation can greatly affect the sedimentation rates. An analytical expression relating the aggregation number to the sedimentation velocity is presented. Our sedimentation experiments with a bidisperse latex suspension as well as clay particle dispersions show both the destabilizing and stabilizing effects of small particles, which are in qualitative agreement with our theoretical predictions.     

Plasma lipoprotein distribution of liposomal nystatin is influenced by protein content of high-density lipoproteins

The plasma lipoprotein distribution of free nystatin (Nys) and liposomal nystatin (L-Nys) in human plasma samples with various lipoprotein lipid and protein concentrations and compositions was investigated. To assess the lipoprotein distributions of Nys and L-Nys, human plasma was incubated with Nys and L-Nys (equivalent to 20 microg/ml) for 5 min at 37 degreesC. The plasma was subsequently partitioned into its lipoprotein and lipoprotein-deficient plasma fractions by step-gradient ultracentrifugation, and each fraction was analyzed for Nys content by high-pressure liquid chromatography. The lipid and protein contents and compositions of each fraction were determined with enzymatic kits. Following the incubation of Nys and L-Nys in human plasma the majority of Nys recovered within the lipoprotein fractions was recovered from the high-density lipoprotein (HDL) fraction. Incorporation of Nys into liposomes consisting of dimyristoylphosphatidylcholine and dimyristoylphosphatidylglycerol significantly increased the percentage of drug recovered within the HDL fraction. Furthermore, it was observed that as the amount of HDL protein decreased the amounts of Nys and L-Nys recovered within this fraction decreased. These findings suggest that the preferential distribution of Nys and L-Nys into plasma HDL may be a function of the HDL protein concentration.     

Sag method for the determination of coefficient of linear thermal expansion of carbon fibres

A method for the determination of coefficient of linear thermal expansion α has been described. The method can be used for conducting wires and fibres. It utilises magnification due to sag produced on heating the sample. A value of α = 1.0 × 10^?6/°C has been obtained for Beslon based carbon fibres produced at N.P.L. of India. Values of α for platinum and nickel wires have been determined by this method. Any contribution to sag due to a small weight hung have also been calculated and found negligible in our experiments.     

The Effects of Competitive Displacement on Haloperidol's Plasma Distribution in Normolipidemic and Hyperlipidemic Plasma

Purpose. To assess whether dyslipidemia affects haloperidol's overall plasma distribution when it is in the presence of another highly protein bound drug that competes for plasma protein binding sites. Methods. We performed in vitro studies in which warfarin sodium was pre-incubated in normolipidemic and hyperlipidemic plasma samples in varying concentrations. Following the pre-incubation with warfarin, [³H]-haloperidol mixed with unlabeled haloperidol was added to the plasma samples. The plasma was separated into its lipoprotein and lipoprotein deficient fractions by density gradient ultracentrifugation and haloperidol distribution was determined. Results. Our results indicate that when normolipidemic plasma was pre-incubated with various concentrations of warfarin no significant redistribution of haloperidol was noted among the various plasma lipoprotein fractions. However, in the case of the hyperlipidemic plasma, pre-incubating with warfarin did result in a significant redistribution of haloperidol from the lipoprotein-deficient fraction to the very-low-density and low-density fractions of lipoproteins. Conclusion. Understanding how plasma lipoproteins influence competitive displacement interactions would be valuable in helping to explain and perhaps predict pharmacokinetic parameters that may affect clinical outcome. The clinical significance of competitive displacement of drugs in patients with dyslipidemia requires further study.     

Assessing the Antifungal Activity of a New Oral Lipid-Based Amphotericin B Formulation Following Administration to Rats Infected with Aspergillus Fumigatus

ABSTRACT

The purpose of this study was to assess the antifungal activity of a new oral amphotericin B (AmpB) lipid-based formulation following administration to rats infected with Aspergillus fumigatus. Aspergillus fumigatus inoculum (2.1–2.5 × 10⁷ colony forming units [CFU]) were injected via the jugular vein; 48h later male albino Sprague-Dawley rats (350–400 g) were administered either a single oral dose of AmpB incorporated into Peceol (50 mg AmpB/kg), physiologic saline (nontreated controls) or Peceol alone (vehicle control) once daily for 4 days. To assess antifungal activity Brain, Lung, Heart, Liver, Spleen and Kidney sections were homogenized with normal saline (1 mL/g of tissue) and a 0.1-mL aliquot was spread plated onto a Sabourand dextrose agar plate. The plates were incubated for 48 hr at 37°C, at which time the number of fungal CFU were determined and corrected for tissue weight. In addition, plasma galactomannan antigen concentrations were determined. Data was reported as mean ± standard error of the mean. The AmpB-Peceol oral formulation significantly decreased total fungal CFU concentrations recovered in all the organs added together, brain CFU concentrations, spleen CFU concentrations and plasma galactomannan antigen concentrations compared to baseline. No significant differences in lung, heart, liver and kidney CFU concentrations between treatment and control groups were observed. Peceol vehicle control did not exhibit any antifungal activity. These findings suggest that a new oral lipid-based formulation of AmpB incorporated into Peceol can significantly decrease brain and spleen CFU concentrations and plasma galactomannan antigen concentrations compared to non-treated controls.     

Cyclosporine A: a review of current oral and intravenous delivery systems

As early as 1978, the immunosuppressive effect of cyclosporine A (CsA), a metabolite of the fungus Tolypocladium inflatum (Borel, 1989), was reported to be effective in inhibiting organ rejection in patients receiving kidney transplants from mismatched cadaver donors (Calne et al., 1978) and in the treatment of graft-versus-host disease in patients with acute leukemia following bone marrow transplants (Powles et al., 1978). Today, CsA is still indicated to prevent rejection following solid organ transplantations, prevent and treat graft-vs-host disease following bone marrow transplants, and has also been used in the treatment of autoimmune disease such as psoriasis, rheumatoid arthritis, and nephrotic syndrome (Canadian Pharmacists Association, 2006). The effectiveness of CsA is derived from its ability to specifically and reversibly inhibit immunocompetent lymphocytes in the G₀ and G₁ phase of the cell cycle. The T-helper cells are the main target, but suppression of the T-suppressor cells also occurs. The production and release of lymphokines, including interleukin-2 are also inhibited (Novartis, 2005a). CsA can be administered intravenously as well as orally in the form of a solution or a soft gelatin capsule. The following review will focus on the evolution of the emulsion-based oral formulations from the first generation as Sandimmune® to the second generation Neoral®, both products of Novartis Pharmaceutical, as well as on the Sandimmune® commercial intravenous formulation. The potential of alternative delivery systems, including micelles, micro- and nanoparticles, and liposomes, will also be discussed.     

Advanced 3D and 4D microstructure study of single granule formation for pharmaceutical powders using synchrotron x-ray imaging

Monitoring the microstructure of the granule in the wet granulation process could play a decisive role in obtaining high-quality granules. Due to the complex, fast and opaque nature of wet granulation, it cannot be captured by conventional methods. In this study, synchrotron x-ray imaging was employed for the first time to investigate the internal real-time pore evolution during the granule formation process, based on the single droplet impact method. It was found that granules from coarser and more homogenous powders experienced a higher rate of pore evolution during nucleation with a more uniform pore distribution. Dynamic wetting studies showed the granule formation mechanisms, the crater mechanism was found for most binary mixtures with 50 wt. % excipients. According to the physical tests, the granules with lower porosity and finer pores exhibited higher hardness and a slower dissolution rate.