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HPMC-based liposomal mucoadhesive films with model peptide as target API
Belcáková, Hedviga ; Nobilis, Milan (advisor) ; Doležal, Pavel (referee)
Charles University, Faculty of Pharmacy in Hradec Kralove Department of Pharmaceutical Chemistry and Pharmaceutical Analysis Candidate: Hedviga Belcáková Supervisor: Prof. Milan Nobilis, CSc., Veronika Fidelj (Heidelberg University, Insitute of Pharmacy and Molecular Biotechnology) Diploma thesis title: HPMC-based liposomal mucoadhesive films with model peptide as target API This thesis describes the preformulation stage of mucoadhesive films intended for liposomal peptide delivery via buccal membrane. The evaluation consisted of thickness, maximum tensile strength, strain, moisture content, in vitro swelling and liposome integrity measurements. The chosen polymer (hypromellose, HPMC) was found to perform optimally in concentrations of 10 % with PEG 400 (5 %) acting as plasticizer and liposome concentration of 2 %. The developed preparation method showed good reproducibilty with room for improvement in the homogenization area. The choice of medium (H2O vs. PBS) showed strong influence on formulation's mechanical properties resulting in significant loss of elasticity and mucoadhesive strength. The addition of liposomes in the third stage had been carried out successfully with only occassional effect on their integrity after dissolution.
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Analytical evaluation of drugs using HPLC II.
Hanková, Eva ; Mokrý, Milan (advisor) ; Nobilis, Milan (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmaceutical Chemistry and Drug Analysis Candidate: Eva Hanková Supervisor: RNDr. Milan Mokrý, CSc. Title of Diploma Thesis: Analytical evaluation of drugs using HPLC II. Abstract This diploma thesis dealt with analytical evaluation of tiaprofenic acid using high performance liquid chromatography. To determine tiaprofenic acid, a LiChroCART® 250-4 LiChrospher® 100 RP-18 (5 μm) column and a mobile phase consisting of acetonitrile and aqueous KH2PO4 buffer 0.01 mol/l in a ratio of 40 : 60 (v/v) were used. The pH of the buffer was adjusted to 3.0 with phosphoric acid; the flow rate was set to 1 ml/min and the temperature at 22 řC. Naproxen was chosen as the internal standard, and a 263 nm wavelength was used for detection. Since the tiaprofenic acid is a chiral substance present in the form of a racemate, the selection of chromatographic conditions for the separation of its enantiomers followed. Two chiral columns in both normal and reverse mode were tested. The cellulose-based Chiralcel OD-R 250 x 4.6 mm (10 μm) column was chosen as the most suitable, using a mobile phase of n-hexane : propan-2-ol : acetic acid in a ratio of 94 : 6 : 0,1 (v/v /v) at a flow rate of 0.5 ml/min and a temperature of 22 ř C. Enantiomers were eluted at...
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The employment of HPLC in the field of chiral separations III.
Janetková, Monika ; Kučera, Radim (advisor) ; Nobilis, Milan (referee)
Department of Pharmaceutical Chemistry and Pharmaceutical Analysis, Faculty of Pharmacy in Hradec Králové, Charles University Candidate: Monika Janetková Supervisor: doc. PharmDr. Radim Kučera, Ph.D. Consultant: doc. Ing. Radim Vespalec, DrSc. Title of Thesis: The employment of HPLC in the field of chiral separations III. Boron cluster compounds are completely synthetic inorganic structures posessing unique properties. Exceptional characteristics allow them to prove competent among organic molecules predominantely used in medicine. Basic structure is made of boron and hydrogen atoms forming three-dimensional polyhedron with triangular planes. The atoms B-H-B are forming electrondeficient three-centred two-electron bond. Cloud of electrons that is shared inside polyhedron structure is the cause of delocalized negative charge of a whole molecule. Boron cluster compounds are likely to loose hydrogen proton in both, protic and aprotic solvents, ending in formation of an anion. Among their typical characteristics belong extreme hydrophobicity, metabolic and thermal stability. Considering stability and reactivity closo-, nido- and arachno-clusters with 11-12 vertices are largely used. Either of endo- or exoskeletal substitution might impair symmetry of a boron cluster molecule resulting in a formation of...
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Modern separation techniques for the analysis of biological material in clinical research
Kasalová, Eva ; Solich, Petr (advisor) ; Nobilis, Milan (referee) ; Friedecký, David (referee)
Charles University, Faculty of Pharmacy in Hradec Kralove Department of Analytical Chemistry Candidate: Mgr. Eva Kasalová Supervisor: Prof. RNDr. Petr Solich, CSc. Supervisor specialist: Assoc. Prof. RNDr. Dagmar Solichová, Ph.D. Title of Doctoral Thesis: Modern separation techniques for the analysis of biological material in clinical research This doctoral thesis deals with development and validation of a chromatographic determination of liposoluble vitamins and with optimization of biological materials preparation. Clinical monitoring of these vitamins, especially vitamin D in different biological materials is one of the most discuss theme in the last decade. In the theoretical part of this dissertation the target analytes - vitamins A, E and D with their biological activity are characterized - as well as the biological material used in the Research Laboratory, especially a detailed description of the composition of breast milk, its irreplaceable role in neonatal nutrition and description of breast milk processing in Milk Bank. This part is followed by a comprehensive chapter dealing with the treatment of the biological matrix before analysis. The most commonly used sample pretreatment techniques such as extraction methods are described here, with a great description of new trends in these...
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The employment of HPLC in the field of chiral separations III.
Janetková, Monika ; Kučera, Radim (advisor) ; Nobilis, Milan (referee)
Department of Pharmaceutical Chemistry and Pharmaceutical Analysis, Faculty of Pharmacy in Hradec Králové, Charles University Candidate: Monika Janetková Supervisor: doc. PharmDr. Radim Kučera, Ph.D. Consultant: doc. Ing. Radim Vespalec, DrSc. Title of Thesis: The employment of HPLC in the field of chiral separations III. Boron cluster compounds are completely synthetic inorganic structures posessing unique properties. Exceptional characteristics allow them to prove competent among organic molecules predominantely used in medicine. Basic structure is made of boron and hydrogen atoms forming three-dimensional polyhedron with triangular planes. The atoms B-H-B are forming electrondeficient three-centred two-electron bond. Cloud of electrons that is shared inside polyhedron structure is the cause of delocalized negative charge of a whole molecule. Boron cluster compounds are likely to loose hydrogen proton in both, protic and aprotic solvents, ending in formation of an anion. Among their typical characteristics belong extreme hydrophobicity, metabolic and thermal stability. Considering stability and reactivity closo-, nido- and arachno-clusters with 11-12 vertices are largely used. Either of endo- or exoskeletal substitution might impair symmetry of a boron cluster molecule resulting in a formation of...
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Analytical evaluation of drugs using HPLC I.
Dobrá, Hana ; Mokrý, Milan (advisor) ; Nobilis, Milan (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmaceutical Chemistry and Drug Analysis Student: Hana Dobrá Supervisor: RNDr. Milan Mokrý, CSc. Title of master's thesis: Analytical evaluation of drugs by using HPLC For the development of chromatographic conditions for suprofen separation were used four conventional different C18 columns. Most suitable appeared column is LiChroCART 125-4 HPLC Cartridge R, LiChrospher 100 RP - 18, 5 mm. Many different mobile phases were used and the most appropriate was mobile phase consisting of acetonitrile and potassium phosphate aqueous buffer in ratio 35/65 with pH 3.03. By using this mobile phase suprofen's retention time was 7.8 minutes. We were also looking for a suitable internal standard. From a few used compounds as the best was chosen naproxen, which was eluted behind the peak of suprofen and provided a separated peak. Many conditions for the chiral separation of suprofen's enantiomers were tried. Finally, suprofen was separated by normal phase mode on a Chiralcel OD-R, 0.46 cm x 25 cm, from Daicel Chemical Industries using a mobile phase of hexane and propan-2-ol in a ratio of 98:2, with the addition of acetic acid at volume 0.6 millilitres, at a flow rate 0.5 ml / min, retention times take about 60 minutes. Analysis was reduced...
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Vliv vybraných látek ovlivňujících centrální nervový systém na kostní metabolismus
Fekete, Soňa ; Živný, Pavel (advisor) ; Horák, Pavel (referee) ; Nobilis, Milan (referee)
The increase in life expectancy of the world population is associated with challenges regarding health issues. For instance, osteoporosis is a medical condition mostly observed in elderly people, in which the quality and quantity of the bone are severely affected. Not only for women but also for men, osteoporosis is recognized as an important public health issue. Osteoporosis is a systemic skeletal disorder and is a result of loss of skeletal mass. Osteoporosis is characterized by low bone mass, microarchitectural deterioration of bone tissue and an increase in bone fragility and susceptibility to fracture. Osteoporotic fractures are a significant cause of morbidity and mortality. The longterm use of drugs such as antiepileptics and antidepressants could affect the onset of osteoporosis. The aim of the study was to evaluate the effect of orchidectomy, the effect of newer antiepileptic (levetiracetam, lacosamide, topiramat, lamotrigine) and antidepressive drugs (mirtazapine, venlafaxine and trazodone) on bone metabolism in healthy male Wistar rats. The first specific aim was to determine the effect of orchidectomy on bone metabolism in rats. We found that after 12 weeks post-orchidectomy there was a negative effect on bone metabolism in rats. These results established these animals as suitable models for...
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Evaluation of biologically active substances using HPLC IV.
Seidlová, Martina ; Pilařová, Pavla (advisor) ; Nobilis, Milan (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department Pharmaceutical Chemistry and Drug Kontrol Name of Degrese Paper: Evaluating of biologically active substances by HPLC IV. Candidate: Martina Seidlová Tutor: PharmDr. Pavla Pilařová, Ph.D. In this diploma work the study of dilation behaviour of medications was provided under extreme conditions for a column being able to measure pH ranged 1-12 on column Kinetex EVO C18 (150 x 3 mm). For higher pH the following medications were studied: quetiapine, and its two metabolites - norquetiapine and 7-hydroxyquetiapine, bromhexine, codeine, zolpidem and its admixture, fluoxetine and aciclovir. The best results were reached with quetiapine with its metabolites, therefore they were chosen for the following tests and possibility of separation from plasma. As a mobile phase an acidic buffer was used at pH10 using ratio 70:30 to organic phase which was made of acetonitrile and methanol (55:5). For lower pH the following medications were studied: salicylic acid, acetylsalicylic acid, aciclovir, bromhexine and fluoxetine. The best results were reached with salicylic acid and acetylsalicylic acid, therefore they were chosen for the following testing and the possibility of separation in the medicament. As a mobile phase a phosphate buffer...
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The Development of SPE Extraction Procedure for Isolation of Pravastatin and its Metabolite from Biological Matrix
Rabatinová, Martina ; Nováková, Lucie (advisor) ; Nobilis, Milan (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Candidate: Mgr. Martina Rabatinová Supervisor: Doc. PharmDr. Lucie Nováková, Ph.D. Title of Rigorous Thesis: The development of SPE extraction procedure for isolation of pravastatin and its metabolite from biological matrix Statins are a class of drugs used to lower cholesterol levels. Statins have an inhibitory activity against 3-hydroxy-3-methylglutaryl coenzyme A reductase. The main purpose of this work was to develop and validate a quantitative method for sensitive analysis of pravastatin and pravastatin lactone. The analytes were extracted from rat urine by a solid phase extraction method. In this method urine samples were prepared on C18 Discovery DSC-18. Washing and elution agents consisted of a mixture of acetonitrile and 0.01 M ammonium acetate buffer, pH 4.5. Number of extraction steps were optimized to eliminate most of matrix effects. The method involved use of ultra high performance liquid chromatography tandem mass spectrometry. Chromatography was carried out on a BEH C18 1.7 µm, 2.1×100 mm column. A mobile phase consisted of a mixture of acetonitrile and 0.5 mM ammonium acetate solution, pH 4.0. Linear gradient conditions were used. Total run time of this anal- ysis was 6 minutes...
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Development and Validation of HPLC Methods for Transport and Metabolic Studies
Hroch, Miloš ; Nobilis, Milan (advisor) ; Šatínský, Dalibor (referee) ; Šišpera, Luděk (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of Pharmaceutical Chemistry and Drug Control Candidate Ing. Miloš Hroch Consultant Doc. PharmDr. Milan Nobilis, CSc. Title of Thesis Development and validation of HPLC methods for transport and metabolic studies The presented PhD thesis deals with the development and validation of chromatographic methods in bioanalysis of drugs and their metabolites. The work provides a theoretical introduction to the field of bioanalysis, using methods of liquid chromatography. Attention was paid to recent trends in the area, particularly with respect to the new types of stationary phases. In the experimental part of the thesis, reader is introduced to a development and validation of several bio-analytical methods using high performance liquid chromatography. The first method concerns with a determination of 5-fluorouracil and its metabolites by HPLC with gradient elution and UV/VIS detection on a column Luna PFP (2). Discussed are mainly specifics for the assay of this group of polar and poorly detectable compounds. The developed and validated method is being used for TDM of patients with rectal cancer. Another discussed method deals with a determination of methotrexate polyglutamates in erythrocytes using isocratic elution on a hybrid...
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