Characterization of Ion-Exchange Fibers for Controlled Drug Delivery

Increasing attention has been focused on methods that deliver pharmacologically active compounds (e.g. drugs, peptides and proteins) in a controlled fashion, so that constant, sustained, site-specific or pulsatile action can be attained. Ion-exchange resins have been widely studied in medical and pharmaceutical applications, including controlled drug delivery, leading to commercialisation of some resin based formulations. Ion-exchangers provide an efficient means to adjust and control drug delivery, as the electrostatic interactions enable precise control of the ion-exchange process and, thus, a more uniform and accurate control of drug release compared to systems that are based only on physical interactions. Unlike the resins, only few studies have been reported on ion-exchange fibers in drug delivery. However, the ion-exchange fibers have many advantageous properties compared to the conventional ion-exchange resins, such as more efficient compound loading into and release from the ion-exchanger, easier incorporation of drug-sized compounds, enhanced control of the ion-exchange process, better mechanical, chemical and thermal stability, and good formulation properties, which make the fibers attractive materials for controlled drug delivery systems. In this study, the factors affecting the nature and strength of the binding/loading of drug-sized model compounds into the ion-exchange fibers was evaluated comprehensively and, moreover, the controllability of subsequent drug release/delivery from the fibers was assessed by modifying the conditions of external solutions. Also the feasibility of ion-exchange fibers for simultaneous delivery of two drugs in combination was studied by dual loading. Donnan theory and theoretical modelling were applied to gain mechanistic understanding on these factors. The experimental results imply that incorporation of model compounds into the ion-exchange fibers was attained mainly as a result of ionic bonding, with additional contribution of non-specific interactions. Increasing the ion-exchange capacity of the fiber or decreasing the valence of loaded compounds increased the molar loading, while more efficient release of the compounds was observed consistently at conditions where the valence or concentration of the extracting counter-ion was increased. Donnan theory was capable of fully interpreting the ion-exchange equilibria and the theoretical modelling supported precisely the experimental observations. The physico-chemical characteristics (lipophilicity, hydrogen bonding ability) of the model compounds and the framework of the fibrous ion-exchanger influenced the affinity of the drugs towards the fibers and may, thus, affect both drug loading and release. It was concluded that precisely controlled drug delivery may be tailored for each compound, in particularly, by choosing a suitable ion-exchange fiber and optimizing the delivery system to take into account the external conditions, also when delivering two drugs simultaneously.

Purification of pharmaceuticals and nutraceutical compounds by sub- and supercritical extraction and chromatography

This thesis discusses the use of sub- and supercritical fluids as the medium in extraction and chromatography. Super- and subcritical extraction was used to separate essential oils from herbal plant Angelica archangelica. The effect of extraction parameters was studied and sensory analyses of the extracts were done by an expert panel. The results of the sensory analyses were compared to the analytically determined contents of the extracts. Sub- and supercritical fluid chromatography (SFC) was used to separate and purify high-value pharmaceuticals. Chiral SFC was used to separate the enantiomers of racemic mixtures of pharmaceutical compounds. Very low (cryogenic) temperatures were applied to substantially enhance the separation efficiency of chiral SFC. The thermodynamic aspects affecting the resolving ability of chiral stationary phases are briefly reviewed. The process production rate which is a key factor in industrial chromatography was optimized by empirical multivariate methods. General linear model was used to optimize the separation of omega-3 fatty acid ethyl esters from esterized fish oil by using reversed-phase SFC. Chiral separation of racemic mixtures of guaifenesin and ferulic acid dimer ethyl ester was optimized by using response surface method with three variables per time. It was found that by optimizing four variables (temperature, load, flowate and modifier content) the production rate of the chiral resolution of racemic guaifenesin by cryogenic SFC could be increased severalfold compared to published results of similar application. A novel pressure-compensated design of industrial high pressure chromatographic column was introduced, using the technology developed in building the deep-sea submersibles (Mir 1 and 2). A demonstration SFC plant was built and the immunosuppressant drug cyclosporine A was purified to meet the requirements of US Pharmacopoeia. A smaller semi-pilot size column with similar design was used for cryogenic chiral separation of aromatase inhibitor Finrozole for use in its development phase 2.

Upregulation and Functionality of Neuronal Nicotinic Acetylcholine Receptors

Cigarette smoking is, in developed countries, the leading cause of premature death. In tobacco smoke, the main addictive compound is nicotine, which in the brain binds to neuronal nicotinic acetylcholine receptors (neuronal nAChRs). These have been implicated in addiction, but also in several neurological disorders including Alzheimer's and Parkinson's diseases, Tourette's syndrome, attention-deficit hyperactivity disorder (ADHD), schizophrenia, pain, depression, and autosomal-dominant noctural frontal lobe epilepsy; all of which makes nAChRs an intriguing target of study. Chronic treatment with nicotine leads to an increase in the number of nAChRs (upregulation) in the brain and changes their functionality. Changes in the properties of nAChRs are likely to occur in smokers as well, since they are exposed to nicotine for long periods of time. Several nAChR subtypes likely play a role in the formation of nicotine addiction by participating in the release of dopamine in the striatum. The aim of this study was to clarify at cellular level the changes in nAChR characteristics resulting from chronic nicotine treatment. SH-SY5Y cells, endogenously several nAChR-expressing, and SH-EP1-h-alfa7 cells, transfected with the alfa 7 nAChR subunit gene were treated chronically with nicotine. The localisation of alfa 7 and beta2 subunits was studied with confocal and electron microscopy. Functionality of nAChRs was studied with calcium fluorometry. Effects of long-term treatment with opioid compounds on nAChRs were studied by means of ligand binding. Confocal microscopy showed that in SH-SY5Y cells, alfa7 and beta2 subunits formed clusters, unlike the case in SH-EP1-h alfa7 cells, where alfa7 nAChRs were distributed more diffusely. The majority of nAChR subunits localised on endoplasmic reticulum (ER). The isomers of methadone acted as agonists at alfa7 nAChRs. Acute morphine challenge also stimulated nAChRs. Chronic treatment with methadone or morphine led to an increased number of nAChRs. In animal studies, mice received nicotine for 7 weeks. Electron microscopical analysis of the localisation of nAChRs showed in the striatum that alfa7 and beta2 nAChR subunits localised synaptically, extrasynaptically, and intracellularly, with the majority localising extrasynaptically. Chronic nicotine treatment caused an increase in the number of nAChR subunits at all studied locations. These results suggest that the alfa7 nAChR and beta2 subunit-containing nAChRs respond to chronic nicotine treatment differently. This may indicate that the functional balance of various nAChR subtypes in control of the release of dopamine is altered as a result of chronic nicotine treatment. Compounds binding both to opioid and nACh receptors may be of clinical importance.

Community Pharmacies and the Needs of Mobile EU Citizens - A study on Finns living in Spain

Migration within the European Union (EU) has increased since the Union was established. Community pharmacies provide open access to health care services and can be the first, most frequently used or even the only contact with a nation s health care system among mobile community residents. In some of the mass-migration areas in Southern Europe, most of the customers may represent mobile citizens of foreign background. This has not always been taken into consideration in the development of community pharmacy services. Mobile patients have been on the EU's health policy agenda, but they have seldom been mentioned in the context of community pharmacies. In most of the EU member states, governments control the specific legislation concerning community pharmacies and there is no harmonised pharmaceutical policy or consistent minimal standards for community pharmacy services in the EU. The aim of this study was to understand medication use, the role of community pharmacies and the symptom mitigation process of mobile community residents. Finns living in Spain were used as an example to examine how community pharmacies in a EU member state meet the needs of mobile community residents. The data were collected by a survey in 2002 (response rate 53%, n= 533) and by five focus group discussions in 2006 (n=30). A large number (70%) of the respondents had moved to Spain for health reasons and suffered from chronic morbidity. Community pharmacies had an important role in the healthcare of mobile community residents and the respondents were mostly satisfied with these services. However, several medication safety risks related to community pharmacy practices were identified: 1) Availability of prescription medicines without prescription (e.g., antibiotics, sleeping pills, Viagra®, asthma medications, cardiovascular medicines, psoriasis medicines and analgesics); 2) Irrational use of medicines (e.g., 41% of antibiotic users had bought their antibiotics without a prescription, and the most common reasons for antibiotic self-medication were symptomatic common colds and sore throats); 3) Language barriers between patients and pharmacy professionals; 4) Lack of medication counselling; 5) Unqualified pharmacy personnel providing pharmacotherapy. A fifth of the respondents reported experiencing problems during pharmacy visits in Spain, and the lack of a common language was the source of most of these problems. The findings of this study indicate that regulations and their enforcement can play a crucial role in actually assuring the rational and safe use of medicines. These results can be used in the development of pharmaceutical and healthcare policies in the EU. It is important to define consistent minimum standards for community pharmacy services in the EU. Then, the increasing number of mobile community residents could access safe and high quality health care services, including community pharmacy services, in every member state within the EU.

Characterisation and processing of amorphous binary mixtures with low glass transition temperature

The number of drug substances in formulation development in the pharmaceutical industry is increasing. Some of these are amorphous drugs and have glass transition below ambient temperature, and thus they are usually difficult to formulate and handle. One reason for this is the reduced viscosity, related to the stickiness of the drug, that makes them complicated to handle in unit operations. Thus, the aim in this thesis was to develop a new processing method for a sticky amorphous model material. Furthermore, model materials were characterised before and after formulation, using several characterisation methods, to understand more precisely the prerequisites for physical stability of amorphous state against crystallisation. The model materials used were monoclinic paracetamol and citric acid anhydrate. Amorphous materials were prepared by melt quenching or by ethanol evaporation methods. The melt blends were found to have slightly higher viscosity than the ethanol evaporated materials. However, melt produced materials crystallised more easily upon consecutive shearing than ethanol evaporated materials. The only material that did not crystallise during shearing was a 50/50 (w/w, %) blend regardless of the preparation method and it was physically stable at least two years in dry conditions. Shearing at varying temperatures was established to measure the physical stability of amorphous materials in processing and storage conditions. The actual physical stability of the blends was better than the pure amorphous materials at ambient temperature. Molecular mobility was not related to the physical stability of the amorphous blends, observed as crystallisation. Molecular mobility of the 50/50 blend derived from a spectral linewidth as a function of temperature using solid state NMR correlated better with the molecular mobility derived from a rheometer than that of differential scanning calorimetry data. Based on the results obtained, the effect of molecular interactions, thermodynamic driving force and miscibility of the blends are discussed as the key factors to stabilise the blends. The stickiness was found to be affected glass transition and viscosity. Ultrasound extrusion and cutting were successfully tested to increase the processability of sticky material. Furthermore, it was found to be possible to process the physically stable 50/50 blend in a supercooled liquid state instead of a glassy state. The method was not found to accelerate the crystallisation. This may open up new possibilities to process amorphous materials that are otherwise impossible to manufacture into solid dosage forms.

Liquid Chromatography-Mass Spectrometry in Studies of Drug Metabolism and Permeability

Poor pharmacokinetics is one of the reasons for the withdrawal of drug candidates from clinical trials. There is an urgent need for investigating in vitro ADME (absorption, distribution, metabolism and excretion) properties and recognising unsuitable drug candidates as early as possible in the drug development process. Current throughput of in vitro ADME profiling is insufficient because effective new synthesis techniques, such as drug design in silico and combinatorial synthesis, have vastly increased the number of drug candidates. Assay technologies for larger sets of compounds than are currently feasible are critically needed. The first part of this work focused on the evaluation of cocktail strategy in studies of drug permeability and metabolic stability. N-in-one liquid chromatography-tandem mass spectrometry (LC/MS/MS) methods were developed and validated for the multiple component analysis of samples in cocktail experiments. Together, cocktail dosing and LC/MS/MS were found to form an effective tool for increasing throughput. First, cocktail dosing, i.e. the use of a mixture of many test compounds, was applied in permeability experiments with Caco-2 cell culture, which is a widely used in vitro model for small intestinal absorption. A cocktail of 7-10 reference compounds was successfully evaluated for standardization and routine testing of the performance of Caco-2 cell cultures. Secondly, cocktail strategy was used in metabolic stability studies of drugs with UGT isoenzymes, which are one of the most important phase II drug metabolizing enzymes. The study confirmed that the determination of intrinsic clearance (Clint) as a cocktail of seven substrates is possible. The LC/MS/MS methods that were developed were fast and reliable for the quantitative analysis of a heterogenous set of drugs from Caco-2 permeability experiments and the set of glucuronides from in vitro stability experiments. The performance of a new ionization technique, atmospheric pressure photoionization (APPI), was evaluated through comparison with electrospray ionization (ESI), where both techniques were used for the analysis of Caco-2 samples. Like ESI, also APPI proved to be a reliable technique for the analysis of Caco-2 samples and even more flexible than ESI because of the wider dynamic linear range. The second part of the experimental study focused on metabolite profiling. Different mass spectrometric instruments and commercially available software tools were investigated for profiling metabolites in urine and hepatocyte samples. All the instruments tested (triple quadrupole, quadrupole time-of-flight, ion trap) exhibited some good and some bad features in searching for and identifying of expected and non-expected metabolites. Although, current profiling software is helpful, it is still insufficient. Thus a time-consuming largely manual approach is still required for metabolite profiling from complex biological matrices.