Isoidide dinitrate (IIDN), a new nitrate ester, an isomer of isosorbide dinitrate (ISDN) and isomannide dinitrate (IMDN), has been studied on the isolated rabbit intestine and on the anesthetized rat blood pressure. Potency was observed as was tachyphylaxis and the effect of certain blocking agents. On the isolated rabbit intestine similar results were obtained when nitroglycerin was substituted for IIDN. In both preparations IIDN was more potent than ISDN or IMDN and ISDN more potent than IMDN. The potency dependence on configuration suggests a possible action on specific receptor sites. A type of tachyphylaxis to IIDN which was prominent in the isolated rabbit intestine was almost absent in the anesthetized rat blood pressure. This is probably due to metabolism of the drug in the intact animal and further suggests an action on receptors. In these preparations the nitrate action was not blocked by: the beta adrenergic blocking agents DCI or nethalide, the alpha adrenergic blocking agent dibenzyline, or a combination of an alpha and a beta blocking agent. Drugs that are selectively blocked by these agents were used as controls to indicate the presence of the desired blocking action. Neither the primary bretylium-like action nor the secondary reserpine-like action of guanethidine blocked the effect of IIDN on the isolated rabbit intestine. Depletion of catechol amines by pretreating the animals with reserpine did not alter the response of either preparation to the nitrate esters. The antihistamine diphenhydramine did not block the vasodepressor action of IIDN on the anesthetized rat blood pressure. It was concluded that although IIDN probably does not exert its effect through combination with adrenergic or histaminergic receptors, it possibly acts on receptors which are specific for the nitrite or nitrate group. / Medicine, Faculty of / Anesthesiology, Pharmacology and Therapeutics, Department of / Graduate

The alpha-glyceryl ethers (AGE) are widely distributed in nature, occurring in many marine organisms, land animals, plants, and humans. A series of preliminary experiments was begun to determine whether these compounds possessed possible pharmacological actions against inflammation. Anti-inflammatory tests utilized both the granuloma pouch and cotton pellet granulation methods. Possible glucocorticoid activity was checked by measuring liver glycogen deposition. The effects of AGE on growth and weight of the animals was also noted. Neither selachyl dihemisuccinate sodium (selachyl DHSS) nor selachyl alcohol decreased the exudate formation in the granuloma pouch experiments, although excellent results were obtained with hydrocortisone. The high dose of the AGE was the equivalent of ten milligrams; low doses were three hundred and six hundred micrograms. Routes of administration used were oral, intraperitoneal, and subcutaneous. In the cotton pellet granuloma test, some indication of anti-inflammatory activity was obtained, in accordance with previous results in this and other laboratories. Selachyl DHSS decreased granulation tissue formation by only 11.7% in growing rats, which is not significant; however, in mature rats, there was a decrease of 29.5%. The dosage used in this series was 30 milligrams per kilogram daily by the subcutaneous route, A definite and comparatively large increase in liver glycogen deposition was observed in rats allowed to eat freely, when given selachyl DHSS subcutaneously. These glycogen values were decreased radically by a paired feeding study in which the treated rats were allowed to eat only the same amount of food as their paired control animal. Thus food intake appears to be one of the factors responsible for the increased glycogen storage. The AGE appear to affect growth only by the Intraperitoneal route of administration. In this case, the growth of rats was retarded by 29.6%. It was also noted that these animals ate somewhat less food than the controls, although this did not appear to be an important enough difference to account for the lack of weight gain. Throughout these experiments one of the most notable observations was a lack of uniformity of the results. Factors which appeared to cause a variability in the outcome of tests included the age of the animals, length of time of preoperative housing in new quarters, route of administration, and dosage. A non-linear relationship between dose and effect appears probable. / Medicine, Faculty of / Anesthesiology, Pharmacology and Therapeutics, Department of / Graduate

Ritodrine is a beta-adrenergic agonist commonly used to inhibit premature labor contractions in women. The primary goal of ritodrine tocolysis is to prolong gestation, however, other indications may include fetal distress. The purpose of the current study was to examine the metabolic and behavioral effects of ritodrine in the fetus, using the chronically instrumented pregnant sheep as an experimental model. Ritodrine was infused continuously into 11 fetal lambs at a rate of 2.6 ug/minute for a period of 8, 12 or 24 hours. Samples were taken simultaneously at predetermined time periods from a fetal femoral artery, umbilical vein, maternal femoral artery and uterine vein for the analysis of fetal and maternal arterial and umbilical and uterine venous blood gases, acid-base status, hematocrit, ritodrine concentration, uterine and umbilical blood flow, and glucose, lactate and oxygen concentrations and fluxes. Cardiovascular and behavioral variables were monitored continuously. The average concentration of ritodrine in fetal arterial plasma was 20.0 ± 2.7 ng/ml (range 9.5 to 3 4.7 ng/ml) at the end of the infusion. This concentration is within the range of cord levels obtained in ritodrine exposed human fetuses at birth (7 to 79 ng/mL) . Fetal arterial plasma ritodrine levels at 8 hours post-infusion were still sufficiently elevated to exert considerable fetal effects. The apparent tolerance of the fetus to given plasma levels of drug varied considerably among animals. The infusion of ritodrine resulted in many typical beta-adrenergic receptor mediated responses in the fetus. Fetal arterial glucose levels rose to 79% above the control by the end of the infusion. This was associated with an increase in fetal glucose delivery (70% above the control), a decrease in the umbilical veno-arterial glucose concentration difference and a tendency for fetal glucose uptake to decline. Fetal arterial plasma lactate concentrations rose more than fivefold during the infusion of ritodrine. This was associated with a rise in fetal lactate delivery (540% above the control), a slight increase in the umbilical veno-arterial plasma lactate concentration difference and a tendency for fetal lactate uptake to rise. Fetal oxygen consumption rose progressively and significantly throughout the infusion of ritodrine and during the first 8 hours of post-infusion, reaching a maximum of 22% above the control by 8 hours post-infusion. Umbilical blood flow remained unchanged, therefore umbilical oxygen delivery was not increased to meet the additional oxygen demands of the fetus. The rise in fetal oxygen consumption was hence accomplished through an increase in fetal fractional oxygen extraction (from a control value of 32.0±1.1% to a maximum of 51.6±1.8% by 1.5 hours of infusion). The rise in fetal oxygen extraction resulted in concurrent declines in fetal arterial Po₂ (78% of the control) and O₂ content (55% of the control) and a widening of the veno-arterial oxygen content difference. By the end of the infusion, umbilical venous Po₂ and O₂ content values had also fallen significantly to 78% and 75% of the control respectively. These latter changes resulted in a concurrent 25% decline in fetal oxygen delivery which in turn contributed to the rise in fetal oxygen extraction. Fetal arterial and umbilical venous pH declined rapidly and significantly from control values of 7.370±0.004 and 7.401±0.005 to 7.274±0.025 and 7.306±0.023 respectively by the end of the infusion. The acidemia was reflected by significant declines in base excess values and appeared to be entirely metabolic in nature, resulting from elevated blood lactate levels. The acidemia likely contributed to the progressive fall in fetal blood O₂ content through a rightward shift of the oxyhemoglobin dissociation curve (Bohr effect). The rise in fetal oxygen consumption was reflected by a similar (although nonsignificant) increase in uterine oxygen consumption. Uteroplacental oxygen consumption appeared to remain unaltered. The rise in uterine oxygen consumption was not accompanied by a corresponding increase in uterine oxygen delivery, hence uterine oxygen extraction rose to 23.8±1.9% (from a control value of 19.5±1.6%) by 1.5 hours post-infusion. The rise in uterine oxygen extraction resulted in significant declines in uterine venous Po₂ and CO₂ values which likely contributed to the fall in fetal oxygen delivery. Fetal heart rate increased significantly to 21% (34 beats per minute, bpm) above the control (162±7 bpm) during the first 1.5 hours of ritodrine infusion. It remained elevated by an average of 16% (26 bpm) throughout the remainder of the infusion and the first 8 hours of post-infusion, returning to the control by the end of the post-infusion period. Fetal arterial pressure remained unchanged from the control (46.2±1.5 mm Hg). The incidence of fetal breathing activity fell significantly from an overall average control value of 43.2±2.6% to an average of 28.1±6.8% during the ritodrine infusion period. In most animals, breathing was most depressed near the end of the infusion. The incidence of low voltage electrocortical (ECoG) activity also fell significantly by an average of 7.5% while that of high voltage ECoG rose by 7.3%. Alterations in intermediate voltage activity were not observed. The incidence of fetal rapid eye movement also tended to fall by an average of 8.2% during the infusion of ritodrine. These behavioral changes may have resulted from the development of fetal hypoxemia, rather than as a direct effect of ritodrine. In conclusion, these data have demonstrated that ritodrine infusion to fetal lambs results in significant physiological and behavioral changes in the fetus. These effects may put the fetus at risk, particularly in situations where fetal oxygen delivery is already reduced, as in various states of compromised pregnancy. / Medicine, Faculty of / Obstetrics and Gynaecology, Department of / Graduate

This thesis describes the synthesis of a series of 5-alkideniminoxy-5-phenylhydantoins for the purpose of studying their anticonvulsant properties.

Healthcare social media is emerging in recent years with increasing attention on people’s health. Online review websites are not only diversi.ed with medicine, hospitals, or doctors but abundant in amount. To discover knowledge from these online reviews, several computational models are proposed. Online healthcare review websites are facing challenges in con.ict of interests among various healthcare stakeholders. To avoid legal complaints and better sustain under such circumstance, we propose a decoupling approach for designing healthcare review websites. Objective components such as medical condition and treatment are remained as the primary parts, as they are generic, impersonal and directly related to patients themselves. Subjective components, however, such as comments to doctors or hospitals are decoupled as secondary parts for sensitive and controversial informa­tion and are optional to reviewers. Our proposed approach shows better .exibility in managing of contents in different levels of details and ability of balancing the right of expression of reviewers with other stakeholders. To identity the patient-reported adverse reactions in drug reviews, we propose a consumer-oriented coding scheme using wordnet synonym and derivational related form. Signi.cant discrepancy of incidences of adverse reactions is discovered be­tween online reviews and clinical trials. We proposed an adverse reaction report ratio model for integrated interpretation of adverse reactions reported in online re­views versus those from clinical trial. Our estimation on average adverse reactions shows high correlation with drug acceptability score obtained from a large-scale meta-analysis. To investigate the impact of key adverse reactions in patients’ perspective, we propose a topic model named Fisher’s Linear Discriminant Analysis Projected Non­negative Matrix Factorization (FLDA-projected-NMF) for discovering discrimina­tive features and topics with additional class information. With satisfaction scores provided in the reviews, discriminative features and topics on satisfaction are dis­covered and polarities of adverse reactions are estimated based on the discriminative feature weights. Discriminative features and topics on medication duration and on age group are obtained as well. Our method outperforms other supervised methods in evaluation of topic sentiment score and topic interpretation measured by entropy. Patient-reported adverse reaction terms are mined from reviews with comment class label. Some new adverse reactions in depression drug and statin drug are also dis­covered. To further study patients’ behaviors, we use structural equation modeling for studying the relationship of factors in patients’ treatment experience with patients’ quality of life. In covariance model, most adverse reactions are found of small co­variance except nausea, headache and dizziness. In measurement model, coef.cients of individual adverse reactions on latent adverse reaction are correlated to the inci­dence of adverse reactions. In structural model, we model the relationship of latent adverse reaction, rating score, positive sentiment and negative sentiment. Compari­son between the measurement models of rating scores of depression drug and statin drug shows that there could be latent factors to account for the variances of latent rating, which shows correlations with the severity of adverse reactions.

Charles Univesity, Faculty of Pharmacy in Hradec Králové Department of Pharmaceutical Chemistry and Pharmaceutical Analysis Candidate: Jana Tomanová Supervisor: doc. PharmDr. Radim Kučera, Ph.D. Consultant: PharmDr. Vít Šesták, Ph.D. Title of the Diploma Thesis: LC-HRMS analysis of selected drugs in biological material I. High performance liquid chromatography is one of the most widespread separation analytical techniques. It is used in various medical, pharmaceutical and industrial laboratories. It uses different affinities of the substances in the analysed mixture to the mobile and stationary phases. Mass spectrometry is one of the instrumental techniques by which charged particles are separated in the gas phase based on the mass- to-charge ratio. Nowadays, great emphasis is placed on the fact that the individual analytical methods which are being developed are subsequently validated according to a guideline issued by national or international authorities. Validation is a process that verifies that a method is suitable for its intended use. In this thesis, the conditions for the determination of warfarin in patient's serum by means of mass spectrometer with LTQ XL linear ion trap were optimized and validated. The optimization of the method was based on the already developed and validated...

Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmaceutical Chemistry and Pharmaceutical Analysis Candidate: Kristýna Švarcová Consultant: RNDr. Martin Mžik, Ph.D. Supervisor: doc. PharmDr. Radim Kučera, Ph.D. Title: LC-HRMS analysis of selected drugs in biological material II. Anxiety disorders affect up to 25 % of the population in their life, the lifetime prevalence is 13,6 %. Insomnia affect at least one third of the population in their life, the prevalence is 15-40 %. Drugs used to treat anxiety and insomnia easily cross the blood-brain barrier and affect the CNS. It is therefore necessary to determine the correct dosing schedule and monitor treatment. The result is optimization of treatment, reduction of side effects and increased patient adherence to treatment. The rapid qualitative and quantitative analysis for urgent cases of intoxication is also needed. This diploma thesis deals with development and optimization of an extraction method for selected benzodiazepines (alprazolam, bromazepam, diazepam, chlordiazepoxide, clonazepam, midazolam, oxazepam) and zolpidem in human serum. Subsequently, the whole analytical method was validated for use in clinical practice using the UHPLC-HRMS. Protein precipitation with acetonitrile was the most suitable sample preparation...

Osteoporosis is a skeletal disease characterized with poor bone quality and low bone mineral density. The pathogenesis of osteoporosis is the imbalance of bone resorption and bone formation. Two strategies can be employed to cure osteoporosis. One is to inhibit bone resorption and the other is to stimulate bone formation. Currently, therapeutic drugs approved by FDA are mainly antiresorptive agents. Till now, there is only one bone anabolic agent approved. Obviously, more efforts should be poured into the development of bone anabolic agents. Sclerostin is a key negative regulator of osteoblast Wnt signaling making it a promising therapeutic target for bone anabolic therapy. Anti-sclerostin humanized monoclonal antibody romosozumab, which could effectively promote bone formation, has been accepted by the FDA for the review of biologic license application in 2017. However, there are several concerns about the humanized anti-sclerostin antibody, including immunogenicity, high cost of production and relative low stability. Nucleic acid aptamers are short single stranded oligonucleotides. They can bind to their targets with similar high affinity as antibodies. Moreover, aptamers have some superior advantages compared to antibodies, such as no immunogenicity, easily synthesized, and high stability. Aptamers against sclerostin could be a promising alternative to antibodies in terms of promotion of bone formation and reversal of osteoporosis. In this thesis, 20 rounds of SELEX were performed to select aptamers with high binding affinity and specificity to sclerostin. The inhibition potency of aptamer candidates to the antagonistic effect of sclerostin on Wnt signaling was also evaluated. Low KD and EC50 values of aptamer candidates against sclerostin implied a great potential of sclerostin aptamer being novel agents to promote bone formation. The study establishes the foundation for the next stage of preclinical studies and it will benefit the development of novel bone anabolic agents to reverse osteoporosis.

Includes bibliographical references / New chemotherapeutics are urgently needed to combat Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis (TB). The development of compounds that could potentiate the activity of known antimycobacterial drugs is a relatively unexplored approach to new TB drug discovery. This study aimed to generate metabolites of chlorpromazine (CPZ), a phenothiazine with demonstrated in vitro activity against Mtb, and to investigate their potential utility in combination with anti-TB drugs. 7-HydroxyCPZ (M2), CPZ-N-oxide (M3), CPZ sulfoxide (M1), nor-CPZ (M5), nor-CPZ sulfoxide (M6b) and CPZ-N-S-dioxide (M4b) were generated from CPZ using various biotransformation systems and identified by Liquid Chromatography - Mass Spectrometry (LC/MS). The identity of M2 was confirmed with reference to a 7-hydroxyCPZ standard. M3, M1, M5, M6b and M4b were synthesized de novo and used to identify the metabolites generated in the biotransformation samples. Individually, CPZ and its metabolites (M2, M3, M5) were weakly active (MIC99 >50μM) against M. smegmatis (Msm) and Mtb while M1, M6b & M4b did not exhibit a MIC99 even at very high concentrations. Generally, an improvement in activity was observed where CPZ or its metabolites were used in combination with known anti-TB drugs. The combinations that exhibited a fractional inhibition concentration index (FICI) of < 0.5 were defined as synergistic. A combination of M2 and spectinomycin (SPEC) exhibited the highest synergism against Msm (FICI 0.19) and Mtb (FICI 0.13). In vitro assays established that CPZ and M2 are bactericidal against Mtb whereas M3 and M5 are bacteriostatic on their own. In combination assays, the use of RIF with M3 and M5, bedaquiline (BDQ) with M2, and SPEC with M3 were bactericidal. At 140μM, CPZ and M1, M2, M3 treated samples exhibited a 2-fold up-regulation of the cydA (Rv1623c) gene which encodes an essential subunit of the cytochrome bd-type menaquinol oxidase in Mtb. The same observation was made for RIF/M2 and RIF/M5 treated samples. These results suggest that the metabolites retain the mechanism of action (MoA) as the parental CPZ. The Mtb 16S rRNA gene, rrs (MTB000019) was identified as the biological target for SPEC. This brought into perspective the underlying mechanisms at play when SPEC is used in combination with CPZ, its metabolites or other drugs, against mycobacteria. This study establishes the utility of combination assays in confirming the active metabolite(s) of known drugs and provides proof of concept data to support follow-up investigations of CPZ and its metabolites as potential compounds for novel combination therapies for anti-TB drug development.

Dissertation submitted to the Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, in fulfilment of the requirements for the degree of Master of Science in Medicine in the branch of Pharmacy / A coated doughnut-shaped tablet is evaluated as its ability to release model drugs at a zero-order rate for 8 to 12 hours. The doughnut-shape tablets were compressed using special designed punches.Automated technology is thus feasible for this system. The coating material , 10% w /w gelatin in HPMC K15M was directly compressed and adhered to the tablet core . / IT2018