An investigation of the antimicrobial and antifouling properties of marine algal metabolites
- Authors: Mann, Maryssa Gudrun Ailsa
- Date: 2008 , 2013-07-11
- Subjects: Anti-infective agents , Marine metabolites -- Therapeutic use , Marine algae , Pharmacognosy , Fouling , Marine fouling organisms
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3831 , http://hdl.handle.net/10962/d1007465 , Anti-infective agents , Marine metabolites -- Therapeutic use , Marine algae , Pharmacognosy , Fouling , Marine fouling organisms
- Description: Prevention of the accumulation of undesirable biological material i.e. biofouling upon a solid surface requires the use of antifouling systems. The solid surface may be a contact lens, an off shore oil rig or a living organism. When chemicals are employed as a mechanism of defense against biofouling, the agents involved are known as antifouling agents. Marine algae must protect themselves from fouling organisms and it is thought that one of the mechanisms used by these organisms is the production of secondary metabolites with an array of biological activities. In vitro studies have shown numerous compounds isolated from marine algae to possess antibacterial, antifungal and antimacrofouling activity. The aim of this study was to evaluate the secondary metabolite extracts of selected Southern African marine macro-algae as a potential source of compounds that inhibit biofilm formation and that could be used as antifouling agents. In this project, marine macro-algae were collected from various sites along the South African coastline. Their extracts were screened for antimicrobial activity against four ubiquitous microorganisms, Staphylococcus aureus, Klebsiella pneumoniae, Mycobacterium aurm and Candida albicans. Results of screening assays guided the fractionation of two Rhodophyta, Plocamium corallorhiza and Laurencia flexuosa. The algae were fractionated using silica gel column chromatography and compounds were isolated by semi-preparative normal phase HPLC. Compound characterization was performed using UV, IR and advanced one- and two-dimensional NMR (¹H, ¹³C NMR, COSY, HSQC, HMBC and NOESY) spectroscopy and mass spectrometry. Ten halogenated monoterpenes including four members of the small class of halogenated monoterpene aldehydes were isolated from extracts of P. corallorhiza. The compounds isolated included the known compounds 3,4,6,7-tetrachloro-3,7-dimethyl-1-octene; 4,6-dibromo-1, 1-dichloro-3,7 -dimethyl-2E,7 octadiene; 4,8-d ibromo-1,1,7 -trichloro-3, 7-dimethyl-2,5Eoctadiene;1 ,4,8-tribromo-3, 7 -dichloro-3,7-dimethyl-1 E,5E-octadiene; 8-bremo-6, 7-dichloro-3,7-dimethyl-octa-2E,4E-dienal; 4-Bromo-8-chloro-3,7-dimethyl-octa-2E,6E-dienal; 4,6- Dibromo-3,7-dimethyl-octa-2E,7-dienal; 2,4-dichloro-1-(2-chlorovinyl)-1-methyl-5-methylidene-cyclohexane and two new metabolites 4,8-chloro-3,7-dimethyl-2Z,4,6Z-octatrien-1-al and Compound 3.47. Methodology was developed for the chemical derivatization and mass spectrometric analysis of the aldehydic compounds, The aldehyde trapping reagent 0-(2,3,4,5,6-pentafluorobenzyl)hydroxylamine hydrochloride was used to derivatize the molecules, stabilizing them and allowing for their complete characterization. From Laurencia flexuosa a new cuparene sesquiterpene 4-bremo-2-(5-hydroxy-1,2,2- trimethylcyclopent-3-enyl)-5-methylphenol was isolated along with two geometric isomers of the vinyl acetylene bromofucin , An halogenated monoterpene 3S*,4R*-1-bromo-3,4,8-trichloro-9-dichloromethyl-1-E,5-E,7-Z-octatriene was also isolated but was suspected to be a contaminant and an investigation into its biological source revealed that it originated from Plocamium suhrii. A third alga, Martensia elegans was extracted based on published reports of antimicrobial compounds in related species. A new a-alkyl malate derivative was isolated and characterized. Selected compounds isolated during the course of the study were employed in preliminary assays that tested their ability to inhibit biofilm formation by Pseudomonas aeruginosa. The halogenated monoterpenes isolated from the Plocamium species were the only active compounds. 3S*,4R*-1-bromo-3,4,S-trichloro-g-dichloromethyl-1-E,5-E,7-octatriene from P. suhrii inhibited biofilm formation through antibacterial activity on planktonic cells but could not prevent biofilm formation when employed as a film on the surface of microtitre plate wells. 1,4,8-tribromo-3,7-dichloro-3,7-dimethyl-1E,5E-octadiene and 4,6-dibromo-1,1-dichloro-3,7-dimethyl-2E,7-octadiene inhibited biofilm formation when applied as a film to the microtitre plate wells but had no significant antibacterial activity. No potential antifouling agents were identified in this project but the antimicrobial activity exhibited by the crude algal extracts was highly encouraging and a number of new research areas have been identified. , KMBT_363 , Adobe Acrobat 9.54 Paper Capture Plug-in
- Full Text:
- Date Issued: 2008
- Authors: Mann, Maryssa Gudrun Ailsa
- Date: 2008 , 2013-07-11
- Subjects: Anti-infective agents , Marine metabolites -- Therapeutic use , Marine algae , Pharmacognosy , Fouling , Marine fouling organisms
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3831 , http://hdl.handle.net/10962/d1007465 , Anti-infective agents , Marine metabolites -- Therapeutic use , Marine algae , Pharmacognosy , Fouling , Marine fouling organisms
- Description: Prevention of the accumulation of undesirable biological material i.e. biofouling upon a solid surface requires the use of antifouling systems. The solid surface may be a contact lens, an off shore oil rig or a living organism. When chemicals are employed as a mechanism of defense against biofouling, the agents involved are known as antifouling agents. Marine algae must protect themselves from fouling organisms and it is thought that one of the mechanisms used by these organisms is the production of secondary metabolites with an array of biological activities. In vitro studies have shown numerous compounds isolated from marine algae to possess antibacterial, antifungal and antimacrofouling activity. The aim of this study was to evaluate the secondary metabolite extracts of selected Southern African marine macro-algae as a potential source of compounds that inhibit biofilm formation and that could be used as antifouling agents. In this project, marine macro-algae were collected from various sites along the South African coastline. Their extracts were screened for antimicrobial activity against four ubiquitous microorganisms, Staphylococcus aureus, Klebsiella pneumoniae, Mycobacterium aurm and Candida albicans. Results of screening assays guided the fractionation of two Rhodophyta, Plocamium corallorhiza and Laurencia flexuosa. The algae were fractionated using silica gel column chromatography and compounds were isolated by semi-preparative normal phase HPLC. Compound characterization was performed using UV, IR and advanced one- and two-dimensional NMR (¹H, ¹³C NMR, COSY, HSQC, HMBC and NOESY) spectroscopy and mass spectrometry. Ten halogenated monoterpenes including four members of the small class of halogenated monoterpene aldehydes were isolated from extracts of P. corallorhiza. The compounds isolated included the known compounds 3,4,6,7-tetrachloro-3,7-dimethyl-1-octene; 4,6-dibromo-1, 1-dichloro-3,7 -dimethyl-2E,7 octadiene; 4,8-d ibromo-1,1,7 -trichloro-3, 7-dimethyl-2,5Eoctadiene;1 ,4,8-tribromo-3, 7 -dichloro-3,7-dimethyl-1 E,5E-octadiene; 8-bremo-6, 7-dichloro-3,7-dimethyl-octa-2E,4E-dienal; 4-Bromo-8-chloro-3,7-dimethyl-octa-2E,6E-dienal; 4,6- Dibromo-3,7-dimethyl-octa-2E,7-dienal; 2,4-dichloro-1-(2-chlorovinyl)-1-methyl-5-methylidene-cyclohexane and two new metabolites 4,8-chloro-3,7-dimethyl-2Z,4,6Z-octatrien-1-al and Compound 3.47. Methodology was developed for the chemical derivatization and mass spectrometric analysis of the aldehydic compounds, The aldehyde trapping reagent 0-(2,3,4,5,6-pentafluorobenzyl)hydroxylamine hydrochloride was used to derivatize the molecules, stabilizing them and allowing for their complete characterization. From Laurencia flexuosa a new cuparene sesquiterpene 4-bremo-2-(5-hydroxy-1,2,2- trimethylcyclopent-3-enyl)-5-methylphenol was isolated along with two geometric isomers of the vinyl acetylene bromofucin , An halogenated monoterpene 3S*,4R*-1-bromo-3,4,8-trichloro-9-dichloromethyl-1-E,5-E,7-Z-octatriene was also isolated but was suspected to be a contaminant and an investigation into its biological source revealed that it originated from Plocamium suhrii. A third alga, Martensia elegans was extracted based on published reports of antimicrobial compounds in related species. A new a-alkyl malate derivative was isolated and characterized. Selected compounds isolated during the course of the study were employed in preliminary assays that tested their ability to inhibit biofilm formation by Pseudomonas aeruginosa. The halogenated monoterpenes isolated from the Plocamium species were the only active compounds. 3S*,4R*-1-bromo-3,4,S-trichloro-g-dichloromethyl-1-E,5-E,7-octatriene from P. suhrii inhibited biofilm formation through antibacterial activity on planktonic cells but could not prevent biofilm formation when employed as a film on the surface of microtitre plate wells. 1,4,8-tribromo-3,7-dichloro-3,7-dimethyl-1E,5E-octadiene and 4,6-dibromo-1,1-dichloro-3,7-dimethyl-2E,7-octadiene inhibited biofilm formation when applied as a film to the microtitre plate wells but had no significant antibacterial activity. No potential antifouling agents were identified in this project but the antimicrobial activity exhibited by the crude algal extracts was highly encouraging and a number of new research areas have been identified. , KMBT_363 , Adobe Acrobat 9.54 Paper Capture Plug-in
- Full Text:
- Date Issued: 2008
Application of dermal microdialysis and tape stripping methods to determine the bioavailability and/or bioequivalence of topical ketoprofen formulations
- Tettey-Amlalo, Ralph Nii Okai
- Authors: Tettey-Amlalo, Ralph Nii Okai
- Date: 2008
- Subjects: Drugs -- Therapeutic equivalency Transdermal medication High performance liquid chromatography Nonsteroidal anti-inflammatory agents -- Bioavailability Nonsteroidal anti-inflammatory agents -- Effectiveness Nonsteroidal anti-inflammatory agents -- Testing Nonsteroidal anti-inflammatory agents -- Side effects
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3796 , http://hdl.handle.net/10962/d1003274
- Description: The widespread acceptance of topical formulations intended for local and/or regional activity has prompted renewed interest in developing a model to determine the bioavailability of drugs in order to establish bioequivalence as a means of evaluating formulation performance of multisource products and also for use during formulation development. Current in vivo techniques such as blister suction and skin biopsy amongst others used to determine the bioavailability and/or bioequivalence of topical formulations are either too invasive to generate appropriate concentration-time profiles or require large numbers of study subjects thereby making the study expensive and time-consuming. Moreover, there are currently no sampling techniques that can demonstrate dermal bioavailability and/or bioequivalence of topical formulations intended for local and/or regional activity. Dermal microdialysis is a relatively new application of microdialysis that permits continuous monitoring of endogenous and/or exogenous solutes in the interstitial fluid. The technique is involves the implantation of semi-permeable membranes which are perfused with an isotonic medium at extremely slow flow rates and collection of microlitre sample volumes containing diffused drugs. Tape stripping, a relatively older technique, has been extensively used in comparative bioavailability studies of various topical formulations. However, due to shortcomings arising from reproducibility and inter-subject variation amongst others, the published FDA guidance outlining the initial protocol was subsequently withdrawn. The incorporation of transepidermal water loss with tape stripping has garnered renewed interest and has been used for the determination of drug bioavailability from a number of topical formulations. Hence the primary objective of this research is to develop and evaluate microdialysis sampling and tape stripping techniques, including the incorporation of the determination of transepidermal water loss, to assess the dermal bioavailability of ketoprofen from topical gel formulations and to develop models for bioequivalence assessment. A rapid UPLC-MS/MS method with requisite sensitivity for the analysis of samples generated from dermal microdialysis was developed and validated which accommodated the microlitre sample volumes collected. An HPLC-UV method was developed and validated for the analysis of samples generated from the in vitro microdialysis and in vivo tape stripping studies. The work presented herein contributes to a growing body of scientific knowledge seeking to develop a model for the determination of bioequivalence of pharmaceutically equivalent topical formulations intended for local and/or regional activity in human subjects.
- Full Text:
- Date Issued: 2008
- Authors: Tettey-Amlalo, Ralph Nii Okai
- Date: 2008
- Subjects: Drugs -- Therapeutic equivalency Transdermal medication High performance liquid chromatography Nonsteroidal anti-inflammatory agents -- Bioavailability Nonsteroidal anti-inflammatory agents -- Effectiveness Nonsteroidal anti-inflammatory agents -- Testing Nonsteroidal anti-inflammatory agents -- Side effects
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3796 , http://hdl.handle.net/10962/d1003274
- Description: The widespread acceptance of topical formulations intended for local and/or regional activity has prompted renewed interest in developing a model to determine the bioavailability of drugs in order to establish bioequivalence as a means of evaluating formulation performance of multisource products and also for use during formulation development. Current in vivo techniques such as blister suction and skin biopsy amongst others used to determine the bioavailability and/or bioequivalence of topical formulations are either too invasive to generate appropriate concentration-time profiles or require large numbers of study subjects thereby making the study expensive and time-consuming. Moreover, there are currently no sampling techniques that can demonstrate dermal bioavailability and/or bioequivalence of topical formulations intended for local and/or regional activity. Dermal microdialysis is a relatively new application of microdialysis that permits continuous monitoring of endogenous and/or exogenous solutes in the interstitial fluid. The technique is involves the implantation of semi-permeable membranes which are perfused with an isotonic medium at extremely slow flow rates and collection of microlitre sample volumes containing diffused drugs. Tape stripping, a relatively older technique, has been extensively used in comparative bioavailability studies of various topical formulations. However, due to shortcomings arising from reproducibility and inter-subject variation amongst others, the published FDA guidance outlining the initial protocol was subsequently withdrawn. The incorporation of transepidermal water loss with tape stripping has garnered renewed interest and has been used for the determination of drug bioavailability from a number of topical formulations. Hence the primary objective of this research is to develop and evaluate microdialysis sampling and tape stripping techniques, including the incorporation of the determination of transepidermal water loss, to assess the dermal bioavailability of ketoprofen from topical gel formulations and to develop models for bioequivalence assessment. A rapid UPLC-MS/MS method with requisite sensitivity for the analysis of samples generated from dermal microdialysis was developed and validated which accommodated the microlitre sample volumes collected. An HPLC-UV method was developed and validated for the analysis of samples generated from the in vitro microdialysis and in vivo tape stripping studies. The work presented herein contributes to a growing body of scientific knowledge seeking to develop a model for the determination of bioequivalence of pharmaceutically equivalent topical formulations intended for local and/or regional activity in human subjects.
- Full Text:
- Date Issued: 2008
Evaluation of the pharmaceutical availability of erythromycin from topical formulations
- Authors: Mandimika, Nyaradzo
- Date: 2008
- Subjects: Pharmacy -- Research Chromatographic analysis Gel permeation chromatography Gels (Pharmacy) Chemistry, analytic Acne -- Treatment Sebaceous glands -- Diseases -- Treatment Drugs -- Testing Erythromycin -- Bioavailability
- Language: English
- Type: Thesis , Masters , MPharm
- Identifier: vital:3771 , http://hdl.handle.net/10962/d1003249
- Description: Erythromycin (ERY) is a macrolide antibiotic which is used in the treatment of acne vulgaris.Acne is a common skin condition that occurs when the sebaceous glands and hair shafts become infected by the bacteria Propionibacterium acnes. Acne is a chronic condition that may last for years and the severity of the effects of the disease on patients is often undermined especially in third world countries where more emphasis is placed on other more life-threatening diseases. It may cause considerable physical and emotional distress to sufferers along with the possibility of permanent scarring. Although use of topical ERY formulations is not the first line of treatment it has proven to be effective in treating inflammation of skin and skin structures cause by the responsible bacteria. To-date there are a variety of vehicles which are used in preparing topical ERY formulations namely ointment and gel bases, alcoholic solutions and pledgets. All the gel formulations on the market contain hydroxypropyl cellulose, alcohol and water along with the active ingredient(s). However, some gel formulations contain propylene glycol in addition to these excipients an example being Emgel®. Propylene glycol has been shown to affect the penetration of topically applied drugs through the skin suggesting that it would be highly likely that those formulations which contain propylene glycol may release more ERY into the skin following application. With this in mind, two ERY gel formulations were produced which contained different percentages of propylene glycol. According to the FDA guidelines, pharmacokinetic measurements in blood, plasma and/or urine of topical dermatological drug products are not feasible to document bioequivalence since the active ingredient(s) in topical formulations is/are not intended to be absorbed into the systemic circulation and in addition, concentrations in extracutaneous biological tissues would generally not be measurable. This limits determination of bioavailability and assessment of bioequivalence of such products to pharmacodynamic measurements, clinical trials and dermatopharmacokinetic (DPK) measurements such as tape stripping (TS) and microdialysis (MD).TS is a sampling technique which involves sequential removal of layers of the stratum corneum using strips of adhesive tape. This technique has found increasing use in DPK studies for investigation of drug kinetics in the skin following the application of a topical formulation. The technique has also been used as a diagnostic tool in assessing the quality of the stratum corneum in diseased skin. In the current research study, the tape stripping technique was used to investigate the pharmaceutical/biological availability of topical gel formulations containing ERY. MD is another DPK sampling technique which has been used to determine the amount of a topically applied drug that penetrates through the stratum corneum to reach deeper tissues of the skin. The in vivo sampling technique involves the insertion of microdialysis probes beneath the skin surface in the dermal tissue and allows for real-time sampling of the analyte at its target site. Recently in vitro MD has also been successfully used to assess the pharmaceutical availability of a topical corticosteroid, mometesone furoate, from topical formulations. Based on this work, microdialysis was used to determine the pharmaceutical availability of ERY from gel formulations which were developed for use in this research. The results of the pharmaceutical availability of ERY from in vivo tape stripping studies and the in vitro microdialysis studies were compared to establish correlation between the data. Pharmaceutical equivalence and bioequivalence data obtained from the respective studies on the gel formulations were investigated by statistical analysis of the data generated from both the in vitro and in vivo experiments. In summary the objectives of this research were: 1. To develop and validate a high performance liquid chromatography method suitable to analyse ERY concentrations obtained from in vitro microdialysis studies and in vivo tape stripping studies. 2. To prepare two different ERY gel formulations with different percentage content of propylene glycol. 3. To determine the pharmaceutical availability of ERY from two different gel formulations using in vitro microdialysis. 4. To develop and validate a tape stripping technique which could be used to determine percutaneous penetration and bioequivalence of the gel formulations. 5. To compare in vitro microdialysis and in vivo tape stripping data and attempt to establish a correlation between the two different approaches.
- Full Text:
- Date Issued: 2008
- Authors: Mandimika, Nyaradzo
- Date: 2008
- Subjects: Pharmacy -- Research Chromatographic analysis Gel permeation chromatography Gels (Pharmacy) Chemistry, analytic Acne -- Treatment Sebaceous glands -- Diseases -- Treatment Drugs -- Testing Erythromycin -- Bioavailability
- Language: English
- Type: Thesis , Masters , MPharm
- Identifier: vital:3771 , http://hdl.handle.net/10962/d1003249
- Description: Erythromycin (ERY) is a macrolide antibiotic which is used in the treatment of acne vulgaris.Acne is a common skin condition that occurs when the sebaceous glands and hair shafts become infected by the bacteria Propionibacterium acnes. Acne is a chronic condition that may last for years and the severity of the effects of the disease on patients is often undermined especially in third world countries where more emphasis is placed on other more life-threatening diseases. It may cause considerable physical and emotional distress to sufferers along with the possibility of permanent scarring. Although use of topical ERY formulations is not the first line of treatment it has proven to be effective in treating inflammation of skin and skin structures cause by the responsible bacteria. To-date there are a variety of vehicles which are used in preparing topical ERY formulations namely ointment and gel bases, alcoholic solutions and pledgets. All the gel formulations on the market contain hydroxypropyl cellulose, alcohol and water along with the active ingredient(s). However, some gel formulations contain propylene glycol in addition to these excipients an example being Emgel®. Propylene glycol has been shown to affect the penetration of topically applied drugs through the skin suggesting that it would be highly likely that those formulations which contain propylene glycol may release more ERY into the skin following application. With this in mind, two ERY gel formulations were produced which contained different percentages of propylene glycol. According to the FDA guidelines, pharmacokinetic measurements in blood, plasma and/or urine of topical dermatological drug products are not feasible to document bioequivalence since the active ingredient(s) in topical formulations is/are not intended to be absorbed into the systemic circulation and in addition, concentrations in extracutaneous biological tissues would generally not be measurable. This limits determination of bioavailability and assessment of bioequivalence of such products to pharmacodynamic measurements, clinical trials and dermatopharmacokinetic (DPK) measurements such as tape stripping (TS) and microdialysis (MD).TS is a sampling technique which involves sequential removal of layers of the stratum corneum using strips of adhesive tape. This technique has found increasing use in DPK studies for investigation of drug kinetics in the skin following the application of a topical formulation. The technique has also been used as a diagnostic tool in assessing the quality of the stratum corneum in diseased skin. In the current research study, the tape stripping technique was used to investigate the pharmaceutical/biological availability of topical gel formulations containing ERY. MD is another DPK sampling technique which has been used to determine the amount of a topically applied drug that penetrates through the stratum corneum to reach deeper tissues of the skin. The in vivo sampling technique involves the insertion of microdialysis probes beneath the skin surface in the dermal tissue and allows for real-time sampling of the analyte at its target site. Recently in vitro MD has also been successfully used to assess the pharmaceutical availability of a topical corticosteroid, mometesone furoate, from topical formulations. Based on this work, microdialysis was used to determine the pharmaceutical availability of ERY from gel formulations which were developed for use in this research. The results of the pharmaceutical availability of ERY from in vivo tape stripping studies and the in vitro microdialysis studies were compared to establish correlation between the data. Pharmaceutical equivalence and bioequivalence data obtained from the respective studies on the gel formulations were investigated by statistical analysis of the data generated from both the in vitro and in vivo experiments. In summary the objectives of this research were: 1. To develop and validate a high performance liquid chromatography method suitable to analyse ERY concentrations obtained from in vitro microdialysis studies and in vivo tape stripping studies. 2. To prepare two different ERY gel formulations with different percentage content of propylene glycol. 3. To determine the pharmaceutical availability of ERY from two different gel formulations using in vitro microdialysis. 4. To develop and validate a tape stripping technique which could be used to determine percutaneous penetration and bioequivalence of the gel formulations. 5. To compare in vitro microdialysis and in vivo tape stripping data and attempt to establish a correlation between the two different approaches.
- Full Text:
- Date Issued: 2008
Isolation and characterization of antiplasmodial metabolites from South African marine alga
- Authors: Afolayan, Anthonia Folake
- Date: 2008
- Subjects: Malaria -- Africa Antimalarials -- Therapeutic use Malaria -- Prevention Malaria -- Drug therapy Marine algae -- Therapeutic use Natural products -- Therapeutic use Plasmodium
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3739 , http://hdl.handle.net/10962/d1003063
- Description: Malaria is one of the three most deadly diseases in Africa. Although there are available treatments, their efficacy has been greatly reduced over the past two decades due to the development of resistance to currently available drugs. This has necessitated the search for new and effective antimalarial agents. This project approached the search for new antimalarial compounds in two ways: (i) by screening natural products isolated from marine algae against the Plasmodium parasite and (ii) by modification of selected isolated active compounds to target 1-deoxY-đ-xylulose 5-phosphate reductoisomerase (DXR), an enzyme found in the nonmevalonate isoprenoid biosynthetic pathway of Plasmodium Jalciparum. It was envisaged that such a compound would exhibit dual action on the Plasmodium parasite. Extracts obtained from 22 marine algae were prefractionated by solvent partitioning and were screened for anti plasmodial activity against the chloroquine sensitive (CQS) P. Jalciparum D 10 strain. Overall, 50% of the algae screened produced at least one crude fraction with activity against P. Jalciparum. Extracts of the algae Sargassum heterophyllum, Plocamium cornutum, Amphiroa ephedrea and Pterosiphonia cloiophylla gave the most promising results. Fractionation of S. heterophyllum afforded three tetraprenyltoluquinols (3.1, 3.2 and 3.5) and an all-trans-fucoxanthin (3.6). Three new compounds (4.5, 4.6 and 4.7) and two known halogenated monoterpenes (4.1 and 4.4) were isolated from P. cornutum. Each of the isolated compounds from both S. heterophyllum and P. cornutum showed antiplasmodial activity with IC₅₀ values ranging from 2.0 - 15.3 μM for S. heterophyllum and 13 - 230 μM for P. cornutum. Attempts to synthetically modify halogenated monoterpene 4.4 by dihydroxylation and phosphorylation in order to inhibit the DXR enzyme was unsuccessful. However, the hemiterpene analogue (5.42) of the halogenated monoterpenes was successfully phosphorylated and dihydroxylated to give compound 5.45 which showed promising activity against DXR. The result obtained indicated that the proposed phosphorylation and dihydroxylation of the halogenated monoterpene 4.4 would result in the synthesis of a potent DXR inhibitor and therefore a potential antimalarial agent with dual mode of action on the Plasmodium parasite.
- Full Text:
- Date Issued: 2008
- Authors: Afolayan, Anthonia Folake
- Date: 2008
- Subjects: Malaria -- Africa Antimalarials -- Therapeutic use Malaria -- Prevention Malaria -- Drug therapy Marine algae -- Therapeutic use Natural products -- Therapeutic use Plasmodium
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3739 , http://hdl.handle.net/10962/d1003063
- Description: Malaria is one of the three most deadly diseases in Africa. Although there are available treatments, their efficacy has been greatly reduced over the past two decades due to the development of resistance to currently available drugs. This has necessitated the search for new and effective antimalarial agents. This project approached the search for new antimalarial compounds in two ways: (i) by screening natural products isolated from marine algae against the Plasmodium parasite and (ii) by modification of selected isolated active compounds to target 1-deoxY-đ-xylulose 5-phosphate reductoisomerase (DXR), an enzyme found in the nonmevalonate isoprenoid biosynthetic pathway of Plasmodium Jalciparum. It was envisaged that such a compound would exhibit dual action on the Plasmodium parasite. Extracts obtained from 22 marine algae were prefractionated by solvent partitioning and were screened for anti plasmodial activity against the chloroquine sensitive (CQS) P. Jalciparum D 10 strain. Overall, 50% of the algae screened produced at least one crude fraction with activity against P. Jalciparum. Extracts of the algae Sargassum heterophyllum, Plocamium cornutum, Amphiroa ephedrea and Pterosiphonia cloiophylla gave the most promising results. Fractionation of S. heterophyllum afforded three tetraprenyltoluquinols (3.1, 3.2 and 3.5) and an all-trans-fucoxanthin (3.6). Three new compounds (4.5, 4.6 and 4.7) and two known halogenated monoterpenes (4.1 and 4.4) were isolated from P. cornutum. Each of the isolated compounds from both S. heterophyllum and P. cornutum showed antiplasmodial activity with IC₅₀ values ranging from 2.0 - 15.3 μM for S. heterophyllum and 13 - 230 μM for P. cornutum. Attempts to synthetically modify halogenated monoterpene 4.4 by dihydroxylation and phosphorylation in order to inhibit the DXR enzyme was unsuccessful. However, the hemiterpene analogue (5.42) of the halogenated monoterpenes was successfully phosphorylated and dihydroxylated to give compound 5.45 which showed promising activity against DXR. The result obtained indicated that the proposed phosphorylation and dihydroxylation of the halogenated monoterpene 4.4 would result in the synthesis of a potent DXR inhibitor and therefore a potential antimalarial agent with dual mode of action on the Plasmodium parasite.
- Full Text:
- Date Issued: 2008
Melatonin and anticancer therapy interactions with 5-Fluorouracil
- Authors: Cassim, Layla
- Date: 2008
- Subjects: Melatonin Melatonin -- Therapeutic use Antineoplastic agents Fluorouracil Fluorouracil -- Toxicology Cancer -- Treatment
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3746 , http://hdl.handle.net/10962/d1003224
- Description: On the basis of clinical studies, some researchers have advocated that the neurohormone and antioxidant melatonin, shown to possess intrinsic anticancer properties, be used as co-therapy in cancer patients being treated with the antineoplastic agent 5-fluorouracil, as increased patient survival times and enhanced quality of life have been observed. The focus of this research was thus to investigate the mechanisms of this seemingly beneficial drug interaction between 5-fluorouracil and melatonin. Metabolism studies were undertaken, in which it was established that there is no hepatic metabolic drug interaction between these agents by cytochrome P450, and that neither agent alters the activity of this enzyme system. Co-therapy with melatonin is thus unlikely to alter plasma levels of 5-fluorouracil by this mechanism. Novel mechanisms by which 5-fluorouracil is toxic were elucidated, such as the induction of lipid peroxidation, due to the formation of reactive oxygen species; decreases in brain serotonin, dopamine and norepinephrine levels, possibly leading to depression; hippocampal shrinkage and morphological alterations and lysis of hippocampal cells, which may underlie cognitive impairment; and a reduction in the nociceptive threshold when administered acutely. All these deleterious effects are attenuated by the co-administration of melatonin, suggesting that the agent exhibits antidepressive and analgesic properties, in addition to its known antioxidative and free radical-scavenging abilities. This suggests that melatonin cotherapy can significantly decrease 5-fluorouracil-induced toxicity, but this may also exert a protective effect on cancer cells and thus compromise the anticancer efficacy of 5-fluorouracil. It was, furthermore, found that stimulation of indoleamine 2,3-dioxygenase activity, mediated by increases in superoxide anion and interferon-γ levels, may underlie resistance to 5-fluorouracil therapy. Melatonin was shown to increase superoxide anion levels in vivo, and this is believed to be by conversion to the metabolite and known oxidant 6- hydroxymelatonin. This highlights that the possible deleterious effects of melatonin metabolites should be studied further. Serum corticosterone levels and cytokine profiles are unaltered by both 5-FU and melatonin, suggesting that these agents may be used by HIV infected individuals without promoting the progression to AIDS. It can thus be concluded that melatonin co-therapy is potentially useful in countering 5-fluorouracil toxicity.
- Full Text:
- Date Issued: 2008
- Authors: Cassim, Layla
- Date: 2008
- Subjects: Melatonin Melatonin -- Therapeutic use Antineoplastic agents Fluorouracil Fluorouracil -- Toxicology Cancer -- Treatment
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3746 , http://hdl.handle.net/10962/d1003224
- Description: On the basis of clinical studies, some researchers have advocated that the neurohormone and antioxidant melatonin, shown to possess intrinsic anticancer properties, be used as co-therapy in cancer patients being treated with the antineoplastic agent 5-fluorouracil, as increased patient survival times and enhanced quality of life have been observed. The focus of this research was thus to investigate the mechanisms of this seemingly beneficial drug interaction between 5-fluorouracil and melatonin. Metabolism studies were undertaken, in which it was established that there is no hepatic metabolic drug interaction between these agents by cytochrome P450, and that neither agent alters the activity of this enzyme system. Co-therapy with melatonin is thus unlikely to alter plasma levels of 5-fluorouracil by this mechanism. Novel mechanisms by which 5-fluorouracil is toxic were elucidated, such as the induction of lipid peroxidation, due to the formation of reactive oxygen species; decreases in brain serotonin, dopamine and norepinephrine levels, possibly leading to depression; hippocampal shrinkage and morphological alterations and lysis of hippocampal cells, which may underlie cognitive impairment; and a reduction in the nociceptive threshold when administered acutely. All these deleterious effects are attenuated by the co-administration of melatonin, suggesting that the agent exhibits antidepressive and analgesic properties, in addition to its known antioxidative and free radical-scavenging abilities. This suggests that melatonin cotherapy can significantly decrease 5-fluorouracil-induced toxicity, but this may also exert a protective effect on cancer cells and thus compromise the anticancer efficacy of 5-fluorouracil. It was, furthermore, found that stimulation of indoleamine 2,3-dioxygenase activity, mediated by increases in superoxide anion and interferon-γ levels, may underlie resistance to 5-fluorouracil therapy. Melatonin was shown to increase superoxide anion levels in vivo, and this is believed to be by conversion to the metabolite and known oxidant 6- hydroxymelatonin. This highlights that the possible deleterious effects of melatonin metabolites should be studied further. Serum corticosterone levels and cytokine profiles are unaltered by both 5-FU and melatonin, suggesting that these agents may be used by HIV infected individuals without promoting the progression to AIDS. It can thus be concluded that melatonin co-therapy is potentially useful in countering 5-fluorouracil toxicity.
- Full Text:
- Date Issued: 2008
Neuroprotective mechanisms of nevirapine and efavirenz in a model of neurodegeneration
- Authors: Zheve, Georgina Teurai
- Date: 2008
- Subjects: HIV infections -- Treatment AIDS (Disease) -- Treatment AIDS dementia complex -- Treatment Nervous system -- Degeneration -- Treatment Melatonin Neurotoxic agents Quinolinic acid
- Language: English
- Type: Thesis , Masters , MPharm
- Identifier: vital:3807 , http://hdl.handle.net/10962/d1003285
- Description: AIDS Dementia Complex (ADC) is a neurodegenerative disorder implicated in HIV-1 infection that is associated with elevated levels of the neurotoxin, quinolinic acid (QA) which causes a cascade of events to occur, leading to the production of reactive oxygen species (ROS), these being ultimately responsible for oxidative neurotoxicity. In clinical studies, Non-nucleoside reverse transcriptase inhibitors (NNRTIs), efavirenz (EFV) and nevirapine (NVP) have been shown to potentially delay the progressive degeneration of neurons, thus reducing the frequency and neurological deficits associated with ADC. Despite these neuroprotective implications, there is still no biochemical data to demonstrate the mechanisms through which these agents offer neuroprotection. The present study aims to elucidate and further characterize the possible antioxidant and neuroprotective mechanisms of NVP and EFV in vitro and in vivo, using QA-induced neurotoxicity as a model. Research has demonstrated that antioxidants and metal chelators have the ability to offer neuroprotection against free radical induced injury and may be beneficial in the prevention or treatment of neurodegeneration. Hence the antioxidant and metal binding properties of these agents were investigated respectively. Inorganic studies, including the 1, 1-diphenyl-2 picrylhydrazyl (DPPH) assay, show that these agents readily scavenge free radicals in vitro, thus postulating the antioxidant property of these agents. The enhancement of superoxide radical generation and iron mediated Fenton reaction by QA is related to lipid peroxidation in biological systems, the extent of which was assayed using the nitroblue tetrazolium and thiobarbituric acid method respectively. Both agents significantly curtail QA-induced lipid peroxidation and potentially scavenge superoxide anions generated by cyanide in vitro. Furthermore, in vivo results demonstrate the ability of NVP and EFV to protect hippocampal neurons against lipid peroxidation induced by QA and superoxide radicals generated as a consequence thereof. The alleviation of QA-induced oxidative stress in vitro possibly occurs through the binding of iron (II) and / or iron (III), and this argument is further strengthened by the ability of EFV and not NVP to reduce iron (II)-induced lipid peroxidation in vitro directly. In addition the ferrozine and electrochemistry assay were used to measure the extent of iron (II) Fe[superscript 2+] and iron (III) Fe[superscript 3+] chelation activity. Both assays demonstrate that these agents bind iron (II) and iron (III), and prevent redox recycling of iron and subsequent complexation of Fe[superscript 2+] with QA which enhances neuronal damage. Both NNRTIs inhibit the endogenous biosynthesis of QA by inhibiting liver tryptophan 2, 3-dioxygenase activity in vivo and subsequently increasing hippocampal serotonin levels. Furthermore, these agents reduce the turnover of hippocampal serotonin to 5-hydroxyindole acetic acid. NVP and not EFV increase 5-hydroxyindole acetic acid and norepinephrine levels in the hippocampus. The results of the pineal indole metabolism study show that NVP increases the synthesis of melatonin, but decreases N-acetylserotonin, 5-hydroxyindole acetic acid and 5-hydroxytryptophol levels. Furthermore, it shows that EFV decreases 5-hydroxyindole acetic acid and melatonin synthesis. Behavioural studies using a Morris water maze show that the post-treatment of rats with NVP and EFV significantly improves QA-induced spatial memory deficits in the hippocampus. This study therefore provides novel information regarding the neuroprotective mechanisms of NVP and EFV. These findings strengthen the argument that these NNRTIs not only have antiviral effects but possess potential neuroprotective properties, which may contribute to the effectiveness of these drugs in the treatment of ADC.
- Full Text:
- Date Issued: 2008
- Authors: Zheve, Georgina Teurai
- Date: 2008
- Subjects: HIV infections -- Treatment AIDS (Disease) -- Treatment AIDS dementia complex -- Treatment Nervous system -- Degeneration -- Treatment Melatonin Neurotoxic agents Quinolinic acid
- Language: English
- Type: Thesis , Masters , MPharm
- Identifier: vital:3807 , http://hdl.handle.net/10962/d1003285
- Description: AIDS Dementia Complex (ADC) is a neurodegenerative disorder implicated in HIV-1 infection that is associated with elevated levels of the neurotoxin, quinolinic acid (QA) which causes a cascade of events to occur, leading to the production of reactive oxygen species (ROS), these being ultimately responsible for oxidative neurotoxicity. In clinical studies, Non-nucleoside reverse transcriptase inhibitors (NNRTIs), efavirenz (EFV) and nevirapine (NVP) have been shown to potentially delay the progressive degeneration of neurons, thus reducing the frequency and neurological deficits associated with ADC. Despite these neuroprotective implications, there is still no biochemical data to demonstrate the mechanisms through which these agents offer neuroprotection. The present study aims to elucidate and further characterize the possible antioxidant and neuroprotective mechanisms of NVP and EFV in vitro and in vivo, using QA-induced neurotoxicity as a model. Research has demonstrated that antioxidants and metal chelators have the ability to offer neuroprotection against free radical induced injury and may be beneficial in the prevention or treatment of neurodegeneration. Hence the antioxidant and metal binding properties of these agents were investigated respectively. Inorganic studies, including the 1, 1-diphenyl-2 picrylhydrazyl (DPPH) assay, show that these agents readily scavenge free radicals in vitro, thus postulating the antioxidant property of these agents. The enhancement of superoxide radical generation and iron mediated Fenton reaction by QA is related to lipid peroxidation in biological systems, the extent of which was assayed using the nitroblue tetrazolium and thiobarbituric acid method respectively. Both agents significantly curtail QA-induced lipid peroxidation and potentially scavenge superoxide anions generated by cyanide in vitro. Furthermore, in vivo results demonstrate the ability of NVP and EFV to protect hippocampal neurons against lipid peroxidation induced by QA and superoxide radicals generated as a consequence thereof. The alleviation of QA-induced oxidative stress in vitro possibly occurs through the binding of iron (II) and / or iron (III), and this argument is further strengthened by the ability of EFV and not NVP to reduce iron (II)-induced lipid peroxidation in vitro directly. In addition the ferrozine and electrochemistry assay were used to measure the extent of iron (II) Fe[superscript 2+] and iron (III) Fe[superscript 3+] chelation activity. Both assays demonstrate that these agents bind iron (II) and iron (III), and prevent redox recycling of iron and subsequent complexation of Fe[superscript 2+] with QA which enhances neuronal damage. Both NNRTIs inhibit the endogenous biosynthesis of QA by inhibiting liver tryptophan 2, 3-dioxygenase activity in vivo and subsequently increasing hippocampal serotonin levels. Furthermore, these agents reduce the turnover of hippocampal serotonin to 5-hydroxyindole acetic acid. NVP and not EFV increase 5-hydroxyindole acetic acid and norepinephrine levels in the hippocampus. The results of the pineal indole metabolism study show that NVP increases the synthesis of melatonin, but decreases N-acetylserotonin, 5-hydroxyindole acetic acid and 5-hydroxytryptophol levels. Furthermore, it shows that EFV decreases 5-hydroxyindole acetic acid and melatonin synthesis. Behavioural studies using a Morris water maze show that the post-treatment of rats with NVP and EFV significantly improves QA-induced spatial memory deficits in the hippocampus. This study therefore provides novel information regarding the neuroprotective mechanisms of NVP and EFV. These findings strengthen the argument that these NNRTIs not only have antiviral effects but possess potential neuroprotective properties, which may contribute to the effectiveness of these drugs in the treatment of ADC.
- Full Text:
- Date Issued: 2008
Development and assessment of an oxytocin parenteral dosage form prepared using pluronic ® F127
- Authors: Chaibva, Faith Anesu
- Date: 2007
- Subjects: Oxytocin -- Therapeutic use , Drugs -- Dosage forms , Pregnancy -- Complications -- Management
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3747 , http://hdl.handle.net/10962/d1003225 , Oxytocin -- Therapeutic use , Drugs -- Dosage forms , Pregnancy -- Complications -- Management
- Full Text:
- Date Issued: 2007
- Authors: Chaibva, Faith Anesu
- Date: 2007
- Subjects: Oxytocin -- Therapeutic use , Drugs -- Dosage forms , Pregnancy -- Complications -- Management
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3747 , http://hdl.handle.net/10962/d1003225 , Oxytocin -- Therapeutic use , Drugs -- Dosage forms , Pregnancy -- Complications -- Management
- Full Text:
- Date Issued: 2007
Development and in vitro evaluation of a clobetasol 17-propionate topical cream formulation
- Authors: Wa Kasongo, Kasongo
- Date: 2007
- Subjects: Adrenocortical hormones , Adrenocortical hormones -- Physiological effect , Drugs -- Testing , Drug development , Dermatopharmacology
- Language: English
- Type: Thesis , Masters , MPharm
- Identifier: vital:3799 , http://hdl.handle.net/10962/d1003277 , Adrenocortical hormones , Adrenocortical hormones -- Physiological effect , Drugs -- Testing , Drug development , Dermatopharmacology
- Description: One of the primary contributing factors to the escalating costs of health care is the high cost of innovator pharmaceutical products. As a consequence, health authorities in various countries and in particular in the developing world have identified generic prescribing and generic substitution as possible strategies to contain the escalating costs of health care provision. There is therefore a need for formulation scientists in developing countries to invest more time in the research and development of generic formulations. Clobetasol 17-propionate (CP) generic cream formulations containing 0.05% w/w of the drug were manufactured and characterized using in vitro testing. Formulation development studies were preceded by the development and validation of an RP-HPLC with UV detection for the quantitation and characterization of CP in innovator and generic cream formulations during formulation development and assessment studies. Furthermore the in vitro release ates of CP release from innovator and generic cream formulations were monitored using a validated in vitro release test method developed in these studies. The formulation of CP cream products was accomplished using a variety of commercially available mixed primary emulsifiers, such as Estol® 1474, Ritapro® 200, Emulcire® 61 WL and Gelot® 64. Successful formulations were selected based on their ability to remain physically stable immediately after manufacture and for 24 hours after storage at room temperature (22°C). Estol® 1474 was found to produce an unstable cream and was therefore not investigated further. The other three emulgents produced stable creams, but only the in vitro release profile of CP from a cream manufactured to contain Gelot® 64 was found to be statistically similar to that of the innovator formulation. Therefore the cream containing Gelot® 64 was selected as the most appropriate prototype generic cream formulation and was characterized in vitro in terms of CP content, viscosity, pH and in vitro release rate. Data generated from these studies were compared to those of the innovator product, Dermovate® cream, using statistical methods. The CP content, pH and in vitro release rate data of the CP formulation were similar to those of the innovator product, however the intrinsic viscosity of Dermovate® cream was almost three (3) times greater than the intrinsic viscosity of the test formulation developed using Gelot® 64. The CP cream formulation developed in these studies was stored for 4 weeks at 40 ± 2°C and 25 ± 5% RH in an incubator and the formulation was found to be stable. A formulation has been developed and assessed and found to be suitable for use as a topical semi-solid dosage form for CP.
- Full Text:
- Date Issued: 2007
- Authors: Wa Kasongo, Kasongo
- Date: 2007
- Subjects: Adrenocortical hormones , Adrenocortical hormones -- Physiological effect , Drugs -- Testing , Drug development , Dermatopharmacology
- Language: English
- Type: Thesis , Masters , MPharm
- Identifier: vital:3799 , http://hdl.handle.net/10962/d1003277 , Adrenocortical hormones , Adrenocortical hormones -- Physiological effect , Drugs -- Testing , Drug development , Dermatopharmacology
- Description: One of the primary contributing factors to the escalating costs of health care is the high cost of innovator pharmaceutical products. As a consequence, health authorities in various countries and in particular in the developing world have identified generic prescribing and generic substitution as possible strategies to contain the escalating costs of health care provision. There is therefore a need for formulation scientists in developing countries to invest more time in the research and development of generic formulations. Clobetasol 17-propionate (CP) generic cream formulations containing 0.05% w/w of the drug were manufactured and characterized using in vitro testing. Formulation development studies were preceded by the development and validation of an RP-HPLC with UV detection for the quantitation and characterization of CP in innovator and generic cream formulations during formulation development and assessment studies. Furthermore the in vitro release ates of CP release from innovator and generic cream formulations were monitored using a validated in vitro release test method developed in these studies. The formulation of CP cream products was accomplished using a variety of commercially available mixed primary emulsifiers, such as Estol® 1474, Ritapro® 200, Emulcire® 61 WL and Gelot® 64. Successful formulations were selected based on their ability to remain physically stable immediately after manufacture and for 24 hours after storage at room temperature (22°C). Estol® 1474 was found to produce an unstable cream and was therefore not investigated further. The other three emulgents produced stable creams, but only the in vitro release profile of CP from a cream manufactured to contain Gelot® 64 was found to be statistically similar to that of the innovator formulation. Therefore the cream containing Gelot® 64 was selected as the most appropriate prototype generic cream formulation and was characterized in vitro in terms of CP content, viscosity, pH and in vitro release rate. Data generated from these studies were compared to those of the innovator product, Dermovate® cream, using statistical methods. The CP content, pH and in vitro release rate data of the CP formulation were similar to those of the innovator product, however the intrinsic viscosity of Dermovate® cream was almost three (3) times greater than the intrinsic viscosity of the test formulation developed using Gelot® 64. The CP cream formulation developed in these studies was stored for 4 weeks at 40 ± 2°C and 25 ± 5% RH in an incubator and the formulation was found to be stable. A formulation has been developed and assessed and found to be suitable for use as a topical semi-solid dosage form for CP.
- Full Text:
- Date Issued: 2007
Evaluation of the safety and efficacy of topical mometasone furoate formulations
- Authors: Chamboko, Bernadett Vongayi
- Date: 2007
- Subjects: Adrenocortical hormones -- Physiological effect , Drugs -- Testing , Dermatopharmacology , High performance liquid chromatography
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3748 , http://hdl.handle.net/10962/d1003226 , Adrenocortical hormones -- Physiological effect , Drugs -- Testing , Dermatopharmacology , High performance liquid chromatography
- Description: The human skin blanching assay (HSBA) is a well-researched and validated method for the bioequivalence assessment of topical corticosteroids. Traditionally, visual assessment of skin blanching has been used. Such testing methods are not conducive for interlaboratory comparisons. Regulatory bodies prefer less subjective methods of analysis. The FDA released guidelines on the assessment of bioequivalence for topical corticosteroids that recommends the use of a chromameter as a reliable method to measure skin blanching although the use of visual assessment with acceptable validation is also provided for. However, the FDA does not elucidate on the manipulation and handling of the chromameter during skin blanching measurements. The purpose of this project was several fold, which included investigations to standardize the manipulation and handling of a chromameter. In particular, measures to avoid skin whitening resulting from the effects of pressure on the skin during chromameter use were investigated. Other methods of analysis should surpass or at least be comparable to the HSBA if such methods are to be used for the assessment of topical corticosteroids. Microdialysis is a relatively new technique for assessing the rate at which drug penetrates the skin. The advantage of using this method is that there are fewer restrictions for selection of an appropriate study population unlike those required for the HSBA where one has to be both a ‘responder’ and a ‘detector’ for their results to be used in data analysis. Microdialysis was investigated by initially conducting experiments in which microdialysis probes were embedded into topical formulations containing mometasone furoate (MF) and the initial results revealed that relatively low drug was released from the formulations. These results indicated that should microdialysis be applied to measure the in vivo release of MF from such topical formulations following application to the skin, even lower concentrations of MF would likely result in the dialysate, necessitating the need for ultra-high sensitive methods of analysis. Typically, the availability of an appropriate analytical technique such as liquid chromatography coupled with mass spectrometry (LCMS) would be a pre-requisite for such in vivo studies. However, only high-pressure liquid chromatography (HPLC) and other less sensitive equipment was available in the laboratories. The study objectives were therefore focussed on in vitro assessment of the release of MF from topical formulations using microdialysis and Franz cells. In addition, the in vivo release of MF was also studied using the HSBA. Data obtained from the microdialysis experiments were compared with the data obtained from the Franz cell diffusion studies in order to provide information on the pharmaceutical availability of MF from the various topical MF dosage forms. Subsequently, pharmaceutical equivalence was investigated from the comparative pharmaceutical availability data using statistical analysis. An additional objective was to attempt to correlate in vitro with in vivo data (IVIVC) to establish a model that could be used to assess safety and efficacy of generic topical drug products. The in vivo data obtained from the HSBA were processed according to the FDA requirements and these pharmacodynamic data were subsequently compared with the microdialysis and Franz cell results. In summary the objectives of this project were: 1. To develop a system to improve the reproducibility of the use of a Minolta® chromameter and compare this with the standard/normal manipulation and handling of such instruments. 2. To develop and validate an HPLC method for the analysis of MF for use with in vitro diffusion studies using microdialysis and Franz cells. 3. To conduct a comparative HSBA on proprietary MF topical creams from two different countries in accordance with the FDA guidance. 4. To assess the pharmaceutical equivalence of topical formulations containing MF using Franz diffusion cells and in vitro microdialysis. 5. To compare the in vivo data obtained from the HSBA with those obtained in vitro using microdialysis and Franz cells.
- Full Text:
- Date Issued: 2007
- Authors: Chamboko, Bernadett Vongayi
- Date: 2007
- Subjects: Adrenocortical hormones -- Physiological effect , Drugs -- Testing , Dermatopharmacology , High performance liquid chromatography
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3748 , http://hdl.handle.net/10962/d1003226 , Adrenocortical hormones -- Physiological effect , Drugs -- Testing , Dermatopharmacology , High performance liquid chromatography
- Description: The human skin blanching assay (HSBA) is a well-researched and validated method for the bioequivalence assessment of topical corticosteroids. Traditionally, visual assessment of skin blanching has been used. Such testing methods are not conducive for interlaboratory comparisons. Regulatory bodies prefer less subjective methods of analysis. The FDA released guidelines on the assessment of bioequivalence for topical corticosteroids that recommends the use of a chromameter as a reliable method to measure skin blanching although the use of visual assessment with acceptable validation is also provided for. However, the FDA does not elucidate on the manipulation and handling of the chromameter during skin blanching measurements. The purpose of this project was several fold, which included investigations to standardize the manipulation and handling of a chromameter. In particular, measures to avoid skin whitening resulting from the effects of pressure on the skin during chromameter use were investigated. Other methods of analysis should surpass or at least be comparable to the HSBA if such methods are to be used for the assessment of topical corticosteroids. Microdialysis is a relatively new technique for assessing the rate at which drug penetrates the skin. The advantage of using this method is that there are fewer restrictions for selection of an appropriate study population unlike those required for the HSBA where one has to be both a ‘responder’ and a ‘detector’ for their results to be used in data analysis. Microdialysis was investigated by initially conducting experiments in which microdialysis probes were embedded into topical formulations containing mometasone furoate (MF) and the initial results revealed that relatively low drug was released from the formulations. These results indicated that should microdialysis be applied to measure the in vivo release of MF from such topical formulations following application to the skin, even lower concentrations of MF would likely result in the dialysate, necessitating the need for ultra-high sensitive methods of analysis. Typically, the availability of an appropriate analytical technique such as liquid chromatography coupled with mass spectrometry (LCMS) would be a pre-requisite for such in vivo studies. However, only high-pressure liquid chromatography (HPLC) and other less sensitive equipment was available in the laboratories. The study objectives were therefore focussed on in vitro assessment of the release of MF from topical formulations using microdialysis and Franz cells. In addition, the in vivo release of MF was also studied using the HSBA. Data obtained from the microdialysis experiments were compared with the data obtained from the Franz cell diffusion studies in order to provide information on the pharmaceutical availability of MF from the various topical MF dosage forms. Subsequently, pharmaceutical equivalence was investigated from the comparative pharmaceutical availability data using statistical analysis. An additional objective was to attempt to correlate in vitro with in vivo data (IVIVC) to establish a model that could be used to assess safety and efficacy of generic topical drug products. The in vivo data obtained from the HSBA were processed according to the FDA requirements and these pharmacodynamic data were subsequently compared with the microdialysis and Franz cell results. In summary the objectives of this project were: 1. To develop a system to improve the reproducibility of the use of a Minolta® chromameter and compare this with the standard/normal manipulation and handling of such instruments. 2. To develop and validate an HPLC method for the analysis of MF for use with in vitro diffusion studies using microdialysis and Franz cells. 3. To conduct a comparative HSBA on proprietary MF topical creams from two different countries in accordance with the FDA guidance. 4. To assess the pharmaceutical equivalence of topical formulations containing MF using Franz diffusion cells and in vitro microdialysis. 5. To compare the in vivo data obtained from the HSBA with those obtained in vitro using microdialysis and Franz cells.
- Full Text:
- Date Issued: 2007
Pharmaceutical analysis and quality of complementary medicines : sceletium and associated products
- Patnala, Satya Siva Rama Ranganath Srinivas
- Authors: Patnala, Satya Siva Rama Ranganath Srinivas
- Date: 2007
- Subjects: Alternative medicine , Herbs -- Therapeutic use
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3872 , http://hdl.handle.net/10962/d1018263
- Description: There has been an upsurge in the use of Complementary and Alternate Medicines (CAMs) in both developed and developing countries. Although herbal medicines have been in use for many centuries, their quality, safety and efficacy are still of major concern. Many countries are in the process of integrating CAMs into conventional health care systems based on the knowledge and use of traditional medicines. The quality control (QC) of herbal products usually presents a formidable analytical challenge in view of the complexity of the constituents in plant material and the commercial non-availability of appropriate qualified reference standards. Sceletium, a genus belonging to the family Aizoaceae, has been reported to contain psychoactive alkaloids, specifically mesembrine, mesembrenone, mesembrenol and some other related alkaloids. Sceletium is marketed as dried plant powder and as phyto-pharmaceutical dosage forms. Sceletium products and plant material marketed through health shops and on the internet are associated with unjustified claims of specific therapeutic efficacy and may be of dubious quality. Validated analytical methods to estimate Sceletium alkaloids have not previously been reported in the scientific literature and the available methods have focused only on qualitative estimation. Furthermore, since appropriate markers were not commercially available for use as reference standards, a primary objective of this study was to isolate relevant compounds, qualify them as reference standards which could be applied to develop appropriate validated qualitative and quantitative analytical methods for fingerprinting and assay of Sceletium plant material and dosage forms. The alkaloidal markers mesembrine, mesembrenone and ∆⁷ mesembrenone were isolated by solvent extraction and chromatography from dried plant material. Mesembranol and epimesembranol were synthesised by hydrogenation of the isolated mesembrine using the catalyst platinum (IV) oxide and then further purified by semi-preparative column chromatography. All compounds were subjected to analysis by ¹H, ¹³C, 2-D nuclear magnetic resonance and liquid chromatography-tandem mass spectroscopy. Mesembrine was converted to hydrochloride crystals and mesembranol was isolated as crystals from the hydrogenation reaction mass. These compounds were analysed and characterised by X-ray crystallography. A relatively simple HPLC method for the separation and quantitative analysis of five relevant alkaloidal components in Sceletium was developed and validated. The method was applied to determine the alkaloids in plant material and dosage forms containing Sceletium. An LCMS method developed during the study provided accurate identification of the five relevant Sceletium alkaloids. The method was applied for the quantitative analysis and QC of Sceletium plant material and its dosage forms. This LCMS method was found to efficiently ionize the relevant alkaloidal markers in order to facilitate their detection, identification and quantification in Sceletium plant material as well as for the assay and QC of dosage forms containing Sceletium. The chemotaxonomy of some Sceletium species and commercially available Sceletium dosage forms were successfully studied by the LCMS method. The HPLC and LCMS methods were also used to monitor the bio-conversion of some of the alkaloids while processing the plant material as per traditional method of fermentation. Additionally a high resolution CZE method was developed for the separation of several Sceletium alkaloids in relatively short analysis times. This analytical method was used successfully to fingerprint the alkaloids and quantify mesembrine in Sceletium and its products. Sceletium species grown under varying conditions at different locations, when analyzed, showed major differences in their composition of alkaloids and an enormous difference was found to exist between the various species with respect to the presence and content of alkaloids. Sceletium and its products marketed through health shops and the internet may thus have problems with respect to the quality and related therapeutic efficacy. The QC of Sceletium presents a formidable challenge as Sceletium plants and products contain a complex mixture of compounds. The work presented herein contributes to a growing body of scientific knowledge to improve the QC standards of herbal medicines and also to provide vital information regarding the selection of plant species and information on the specific alkaloidal constituents to the cultivators of Sceletium and the manufacturers of its products.
- Full Text:
- Date Issued: 2007
- Authors: Patnala, Satya Siva Rama Ranganath Srinivas
- Date: 2007
- Subjects: Alternative medicine , Herbs -- Therapeutic use
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3872 , http://hdl.handle.net/10962/d1018263
- Description: There has been an upsurge in the use of Complementary and Alternate Medicines (CAMs) in both developed and developing countries. Although herbal medicines have been in use for many centuries, their quality, safety and efficacy are still of major concern. Many countries are in the process of integrating CAMs into conventional health care systems based on the knowledge and use of traditional medicines. The quality control (QC) of herbal products usually presents a formidable analytical challenge in view of the complexity of the constituents in plant material and the commercial non-availability of appropriate qualified reference standards. Sceletium, a genus belonging to the family Aizoaceae, has been reported to contain psychoactive alkaloids, specifically mesembrine, mesembrenone, mesembrenol and some other related alkaloids. Sceletium is marketed as dried plant powder and as phyto-pharmaceutical dosage forms. Sceletium products and plant material marketed through health shops and on the internet are associated with unjustified claims of specific therapeutic efficacy and may be of dubious quality. Validated analytical methods to estimate Sceletium alkaloids have not previously been reported in the scientific literature and the available methods have focused only on qualitative estimation. Furthermore, since appropriate markers were not commercially available for use as reference standards, a primary objective of this study was to isolate relevant compounds, qualify them as reference standards which could be applied to develop appropriate validated qualitative and quantitative analytical methods for fingerprinting and assay of Sceletium plant material and dosage forms. The alkaloidal markers mesembrine, mesembrenone and ∆⁷ mesembrenone were isolated by solvent extraction and chromatography from dried plant material. Mesembranol and epimesembranol were synthesised by hydrogenation of the isolated mesembrine using the catalyst platinum (IV) oxide and then further purified by semi-preparative column chromatography. All compounds were subjected to analysis by ¹H, ¹³C, 2-D nuclear magnetic resonance and liquid chromatography-tandem mass spectroscopy. Mesembrine was converted to hydrochloride crystals and mesembranol was isolated as crystals from the hydrogenation reaction mass. These compounds were analysed and characterised by X-ray crystallography. A relatively simple HPLC method for the separation and quantitative analysis of five relevant alkaloidal components in Sceletium was developed and validated. The method was applied to determine the alkaloids in plant material and dosage forms containing Sceletium. An LCMS method developed during the study provided accurate identification of the five relevant Sceletium alkaloids. The method was applied for the quantitative analysis and QC of Sceletium plant material and its dosage forms. This LCMS method was found to efficiently ionize the relevant alkaloidal markers in order to facilitate their detection, identification and quantification in Sceletium plant material as well as for the assay and QC of dosage forms containing Sceletium. The chemotaxonomy of some Sceletium species and commercially available Sceletium dosage forms were successfully studied by the LCMS method. The HPLC and LCMS methods were also used to monitor the bio-conversion of some of the alkaloids while processing the plant material as per traditional method of fermentation. Additionally a high resolution CZE method was developed for the separation of several Sceletium alkaloids in relatively short analysis times. This analytical method was used successfully to fingerprint the alkaloids and quantify mesembrine in Sceletium and its products. Sceletium species grown under varying conditions at different locations, when analyzed, showed major differences in their composition of alkaloids and an enormous difference was found to exist between the various species with respect to the presence and content of alkaloids. Sceletium and its products marketed through health shops and the internet may thus have problems with respect to the quality and related therapeutic efficacy. The QC of Sceletium presents a formidable challenge as Sceletium plants and products contain a complex mixture of compounds. The work presented herein contributes to a growing body of scientific knowledge to improve the QC standards of herbal medicines and also to provide vital information regarding the selection of plant species and information on the specific alkaloidal constituents to the cultivators of Sceletium and the manufacturers of its products.
- Full Text:
- Date Issued: 2007
An investigation into the neuroprotective effects of dehydroepiandrosterone
- Authors: Palvie, Stefanie Michelle
- Date: 2006
- Subjects: Aging -- Physiological aspects , Nervous system -- Degeneration -- Treatment , Steroid hormones , Dehydroepiandrosterone , Dehydroepiandrosterone -- Therapeutic use , Neurosciences , Neuroanatomy , Apoptosis , Pineal gland -- Physiology , Neurotoxic agents , Free radicals (Chemistry) -- Physiological effect
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3782 , http://hdl.handle.net/10962/d1003260 , Aging -- Physiological aspects , Nervous system -- Degeneration -- Treatment , Steroid hormones , Dehydroepiandrosterone , Dehydroepiandrosterone -- Therapeutic use , Neurosciences , Neuroanatomy , Apoptosis , Pineal gland -- Physiology , Neurotoxic agents , Free radicals (Chemistry) -- Physiological effect
- Description: Dehydroepiandrosterone, a C-19 steroid, is found endogenously with the highest circulating serum levels. It is converted to important steroids such as the sex hormones oestrogen and testosterone. DHEA has come under the spotlight as a purported “fountain of youth” due to its well-characterised age-related decline. The supplementation of DHEA in both the elderly and those with a pathophysiological deficiency has been shown to be of benefit, particularly with regard to wellbeing and depression. The role of DHEA in the periphery has not been elucidated beyond its role as a precursor hormone in sex steroid biosynthesis, though it has been established as a neuroactive neurosteroid, capable of exerting neuroprotective effects in the brain. Since the importance of free radicals in aging and neurodegeneration is well established, investigations were conducted on the ability of DHEA to inhibit free radical generation or scavenge existing free radicals. DHEA was able to significantly inhibit quinolinic acid-induced lipid peroxidation, a measure of membrane damage, over a range of concentrations, although the reduction did not appear to be dose-dependent. This was observed in both in vitro and in vivo studies. Thus, the ability of a compound to reduce the degree of lipid peroxidation may indicate its value as a neuroprotectant. However, DHEA did not significantly reduce cyanide induced generation of the superoxide free radical, suggesting that DHEA is not an effective free radical scavenger of the superoxide anion and that the reduction in lipid peroxidation does not occur through a scavenging mechanism. Apoptosis is a physiological process which is necessary for development and homeostasis. However, this form of programmed cell death can be initiated through various mechanisms and too much apoptotic cell death results in deleterious effects in the body. DHEA was shown not to induce apoptosis. Even the lowest concentration of DHEA investigated in this thesis shows a remarkable decrease in the degree of apoptosis caused by intrahippocampal chemical insult by the neurotoxin quinolinic acid. Cresyl violet was used to visualise tissue for histological examination which revealed that DHEA is able to preserve the normal healthy morphology of hippocampal cells which have been exposed to quinolinic acid. Cells maintained their integrity and showed little evidence of swelling associated with necrosis. Organ culture studies were performed by assessing the impact of DHEA on several pineal metabolites. The study revealed that DHEA exerted an effect on the metabolism of indoleamines in the pineal gland. Melatonin, the chief pineal hormone, did not appear to be affected while the concentrations of N-acetylserotonin, serotonin and methoxytryptamine showed significant alterations. Thus, the neuroprotective mechanism of DHEA does not appear to be mediated by an increase in the presence of melatonin. The biological importance of metal ions in neurodegeneration is also well established and thus the potential interaction between DHEA and metal ions was considered as a mechanism of action. Spectroscopic and electrochemical analyses were performed to determine whether DHEA is able to interact with metal ions as a ligand. These reveal that DHEA does not form a strong bond with the metals investigated, namely copper (II) and iron (III), but that a weak interaction is evident. These investigations were conducted in a rodent model, which has neither large amounts of endogenous DHEA, nor the enzymatic infrastructure present in humans. Thus, the theory that DHEA exerts its effects through downstream metabolic products is unlikely. However, these investigations reveal that there is merit in the statement that DHEA itself is a neuroprotective molecule, and confirm that the further investigation of DHEA is an advisable strategy in the war against neurodegeneration and aging.
- Full Text:
- Date Issued: 2006
- Authors: Palvie, Stefanie Michelle
- Date: 2006
- Subjects: Aging -- Physiological aspects , Nervous system -- Degeneration -- Treatment , Steroid hormones , Dehydroepiandrosterone , Dehydroepiandrosterone -- Therapeutic use , Neurosciences , Neuroanatomy , Apoptosis , Pineal gland -- Physiology , Neurotoxic agents , Free radicals (Chemistry) -- Physiological effect
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3782 , http://hdl.handle.net/10962/d1003260 , Aging -- Physiological aspects , Nervous system -- Degeneration -- Treatment , Steroid hormones , Dehydroepiandrosterone , Dehydroepiandrosterone -- Therapeutic use , Neurosciences , Neuroanatomy , Apoptosis , Pineal gland -- Physiology , Neurotoxic agents , Free radicals (Chemistry) -- Physiological effect
- Description: Dehydroepiandrosterone, a C-19 steroid, is found endogenously with the highest circulating serum levels. It is converted to important steroids such as the sex hormones oestrogen and testosterone. DHEA has come under the spotlight as a purported “fountain of youth” due to its well-characterised age-related decline. The supplementation of DHEA in both the elderly and those with a pathophysiological deficiency has been shown to be of benefit, particularly with regard to wellbeing and depression. The role of DHEA in the periphery has not been elucidated beyond its role as a precursor hormone in sex steroid biosynthesis, though it has been established as a neuroactive neurosteroid, capable of exerting neuroprotective effects in the brain. Since the importance of free radicals in aging and neurodegeneration is well established, investigations were conducted on the ability of DHEA to inhibit free radical generation or scavenge existing free radicals. DHEA was able to significantly inhibit quinolinic acid-induced lipid peroxidation, a measure of membrane damage, over a range of concentrations, although the reduction did not appear to be dose-dependent. This was observed in both in vitro and in vivo studies. Thus, the ability of a compound to reduce the degree of lipid peroxidation may indicate its value as a neuroprotectant. However, DHEA did not significantly reduce cyanide induced generation of the superoxide free radical, suggesting that DHEA is not an effective free radical scavenger of the superoxide anion and that the reduction in lipid peroxidation does not occur through a scavenging mechanism. Apoptosis is a physiological process which is necessary for development and homeostasis. However, this form of programmed cell death can be initiated through various mechanisms and too much apoptotic cell death results in deleterious effects in the body. DHEA was shown not to induce apoptosis. Even the lowest concentration of DHEA investigated in this thesis shows a remarkable decrease in the degree of apoptosis caused by intrahippocampal chemical insult by the neurotoxin quinolinic acid. Cresyl violet was used to visualise tissue for histological examination which revealed that DHEA is able to preserve the normal healthy morphology of hippocampal cells which have been exposed to quinolinic acid. Cells maintained their integrity and showed little evidence of swelling associated with necrosis. Organ culture studies were performed by assessing the impact of DHEA on several pineal metabolites. The study revealed that DHEA exerted an effect on the metabolism of indoleamines in the pineal gland. Melatonin, the chief pineal hormone, did not appear to be affected while the concentrations of N-acetylserotonin, serotonin and methoxytryptamine showed significant alterations. Thus, the neuroprotective mechanism of DHEA does not appear to be mediated by an increase in the presence of melatonin. The biological importance of metal ions in neurodegeneration is also well established and thus the potential interaction between DHEA and metal ions was considered as a mechanism of action. Spectroscopic and electrochemical analyses were performed to determine whether DHEA is able to interact with metal ions as a ligand. These reveal that DHEA does not form a strong bond with the metals investigated, namely copper (II) and iron (III), but that a weak interaction is evident. These investigations were conducted in a rodent model, which has neither large amounts of endogenous DHEA, nor the enzymatic infrastructure present in humans. Thus, the theory that DHEA exerts its effects through downstream metabolic products is unlikely. However, these investigations reveal that there is merit in the statement that DHEA itself is a neuroprotective molecule, and confirm that the further investigation of DHEA is an advisable strategy in the war against neurodegeneration and aging.
- Full Text:
- Date Issued: 2006
An investigation into the neuroprotective effects of estrogen and progesterone in a model of homocysteine-induced neurodegeration
- Authors: Wu, Wing Man
- Date: 2006
- Subjects: Homocysteine , Estrogen , Estrogen -- Therapeutic use , Progesterone , Hormone receptors , Methyl aspartate , Oxidative stress , Alzheimer's disease -- Treatment , Nervous system -- Degeneration -- Prevention
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3806 , http://hdl.handle.net/10962/d1003284 , Homocysteine , Estrogen , Estrogen -- Therapeutic use , Progesterone , Hormone receptors , Methyl aspartate , Oxidative stress , Alzheimer's disease -- Treatment , Nervous system -- Degeneration -- Prevention
- Description: Homocysteine (Hcy) is a sulfur containing amino acid and is a potent neurotoxin. It has been shown that elevated levels of Hcy, termed hyperhomocysteinemia, plays a role in the pathologies of Alzheimer’s disease (AD) and age-related cognitive decline. Hcy is a glutamate agonist, which causes in increase in Ca[superscript (2+)] influx via the activation of NMDA class of excitatory amino acid receptors, which results in neuronal cell death and apoptosis. Estrogen and progesterone are female hormones that are responsible for reproduction and maternal behaviour. However, in the last decade, it is evident that both female hormones have neuroprotective properties in many animal models of neurodegeneration. Collectively, both estrogen and progesterone reduce the consequences of the oxidative stress by enhancing the antioxidant defence mechanisms, reducing excitotoxicity by altering glutamate receptor activity and reducing the damage caused by lipid peroxidation. However, the mechanisms by which estrogen and progesterone provide such neuroprotection probably depend on the type and concentration of hormone present. Moreover, numerous studies have shown that hormone replacement therapy (HRT, estrogen and progestins) or estrogen-only replacement therapy (ERT) may prevent or delay the onset of AD and improve cognition for women with AD. Clinical trials have also shown that women taking HRT may modify the effects of Hcy levels on cognitive functioning. Oxidative stress increases in the aging brain and thus has a powerful effect on enhanced susceptibility to neurodegenerative disease. The detection and measurement of lipid peroxidation and superoxide anion radicals in the brain tissue supports the involvement of free radical reactions in neurotoxicity and in neurodegenerative disorders. The hippocampus is an important region of the brain responsible for the formation of memory. However, agents that induce stress in this area have harmful effects and could lead to dementia. This study aims to investigate and clarify the neuroprotective effects of estrogen and progesterone, using Hcy-induced neurodegenerative models. The initial studies demonstrate that estrogen and progesterone have the ability to scavenge potent free radicals. Histological studies undertaken reveal that both estrogen and progesterone protect against Hcy-induced neuronal cell death. In addition, immunohistochemical investigations show that Hcy-induced apoptosis in the hippocampus can be inhibited by both estrogen and progesterone. However, estrogen also acts at the NMDA receptor as an agonist, while progesterone blocks at the NMDA receptor. These mechanisms reduce the ability of Hcy to cause damage to neurons, since Hcy-induced neurotoxicity is dependent on the overstimulation of the NMDA receptor. SOD and GPx are important enzymatic antioxidants which can react with ROS and neutralize them before these inflict damage in the brain. Hcy can increase oxidative stress by inhibiting expression and function of these antioxidants. However, it has been shown that the antioxidant abilities of both estrogen and progesterone can up-regulate the activities of SOD and GPx. These results provide further evidence that estrogen and progesterone act as antioxidants and are free radical scavengers. The discovery of neuroprotective agents is becoming important as accumulating evidence indicates the protective role of both estrogen and progesterone in Hcy-induced neurodegeneration. Thus further work in clinical trials is needed to examine whether reducing Hcy levels with HRT can become the treatment of neurodegenerative disorders, such as Alzheimer’s disease.
- Full Text:
- Date Issued: 2006
- Authors: Wu, Wing Man
- Date: 2006
- Subjects: Homocysteine , Estrogen , Estrogen -- Therapeutic use , Progesterone , Hormone receptors , Methyl aspartate , Oxidative stress , Alzheimer's disease -- Treatment , Nervous system -- Degeneration -- Prevention
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3806 , http://hdl.handle.net/10962/d1003284 , Homocysteine , Estrogen , Estrogen -- Therapeutic use , Progesterone , Hormone receptors , Methyl aspartate , Oxidative stress , Alzheimer's disease -- Treatment , Nervous system -- Degeneration -- Prevention
- Description: Homocysteine (Hcy) is a sulfur containing amino acid and is a potent neurotoxin. It has been shown that elevated levels of Hcy, termed hyperhomocysteinemia, plays a role in the pathologies of Alzheimer’s disease (AD) and age-related cognitive decline. Hcy is a glutamate agonist, which causes in increase in Ca[superscript (2+)] influx via the activation of NMDA class of excitatory amino acid receptors, which results in neuronal cell death and apoptosis. Estrogen and progesterone are female hormones that are responsible for reproduction and maternal behaviour. However, in the last decade, it is evident that both female hormones have neuroprotective properties in many animal models of neurodegeneration. Collectively, both estrogen and progesterone reduce the consequences of the oxidative stress by enhancing the antioxidant defence mechanisms, reducing excitotoxicity by altering glutamate receptor activity and reducing the damage caused by lipid peroxidation. However, the mechanisms by which estrogen and progesterone provide such neuroprotection probably depend on the type and concentration of hormone present. Moreover, numerous studies have shown that hormone replacement therapy (HRT, estrogen and progestins) or estrogen-only replacement therapy (ERT) may prevent or delay the onset of AD and improve cognition for women with AD. Clinical trials have also shown that women taking HRT may modify the effects of Hcy levels on cognitive functioning. Oxidative stress increases in the aging brain and thus has a powerful effect on enhanced susceptibility to neurodegenerative disease. The detection and measurement of lipid peroxidation and superoxide anion radicals in the brain tissue supports the involvement of free radical reactions in neurotoxicity and in neurodegenerative disorders. The hippocampus is an important region of the brain responsible for the formation of memory. However, agents that induce stress in this area have harmful effects and could lead to dementia. This study aims to investigate and clarify the neuroprotective effects of estrogen and progesterone, using Hcy-induced neurodegenerative models. The initial studies demonstrate that estrogen and progesterone have the ability to scavenge potent free radicals. Histological studies undertaken reveal that both estrogen and progesterone protect against Hcy-induced neuronal cell death. In addition, immunohistochemical investigations show that Hcy-induced apoptosis in the hippocampus can be inhibited by both estrogen and progesterone. However, estrogen also acts at the NMDA receptor as an agonist, while progesterone blocks at the NMDA receptor. These mechanisms reduce the ability of Hcy to cause damage to neurons, since Hcy-induced neurotoxicity is dependent on the overstimulation of the NMDA receptor. SOD and GPx are important enzymatic antioxidants which can react with ROS and neutralize them before these inflict damage in the brain. Hcy can increase oxidative stress by inhibiting expression and function of these antioxidants. However, it has been shown that the antioxidant abilities of both estrogen and progesterone can up-regulate the activities of SOD and GPx. These results provide further evidence that estrogen and progesterone act as antioxidants and are free radical scavengers. The discovery of neuroprotective agents is becoming important as accumulating evidence indicates the protective role of both estrogen and progesterone in Hcy-induced neurodegeneration. Thus further work in clinical trials is needed to examine whether reducing Hcy levels with HRT can become the treatment of neurodegenerative disorders, such as Alzheimer’s disease.
- Full Text:
- Date Issued: 2006
An investigation into the neuroprotective properties of acyclovir
- Authors: Müller, Adrienne Carmel
- Date: 2006
- Subjects: Acyclovir -- Therapeutic use , Acyclovir -- Physiological effect , Nervous system -- Degeneration -- Treatment , Memory disorders -- Treatment , Quinolinic acid
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3776 , http://hdl.handle.net/10962/d1003254 , Acyclovir -- Therapeutic use , Acyclovir -- Physiological effect , Nervous system -- Degeneration -- Treatment , Memory disorders -- Treatment , Quinolinic acid
- Description: Accumulating evidence suggests that quinolinic acid has a role to play in disorders involving impairment of learning and memory. In the present study, the effect of the guanosine analogue antiherpetic, acyclovir, on quinolinic acid-induced spatial memory deficits was investigated, as well as some of the mechanisms which underlie this effect. Behavioural studies using a Morris water maze show that post-treatment of rats with acyclovir significantly improves spatial memory deficits induced by intrahippocampal injections of quinolinic acid. Histological analysis of the hippocampi show that the effect of acyclovir is related to its ability to alleviate quinolinic acid-induced necrotic cell death, through interference with some of the mechanisms of neurodegeneration. However, acyclovir is unable to alter a quinolinic acid-induced increase in glutamate release in the rat hippocampus, even though it alleviates quinolinic acid induced oxidative stress by scavenging the superoxide anion in vitro and in vivo in whole rat brain and hippocampus respectively. Due to the inverse relationship which exists between superoxide anion and glutathione levels, acyclovir also curtails the quinolinic acid-induced decrease in hippocampal glutathione levels. Acyclovir suppresses quinolinic acid-induced lipid peroxidation in vitro and in vivo, in whole rat brain and hippocampus respectively, through its alleviation of oxidative stress and possibly through the binding of iron (II) and / or iron (III), preventing the participation and redox recycling of iron (II) in the Fenton reaction, which quinolinic acid is thought to enhance by weak binding of ferrous ions. This argument is further strengthened by the ability of the drug to suppress iron (II)-induced lipid peroxidation in vitro directly. Inorganic studies including ultraviolet and visible spectroscopy, electrochemistry and the ferrozine assay show that acyclovir binds to iron (II) and iron (III) and that quinolinic acid forms an easily oxidisable association with iron (II). Acyclovir inhibits the endogenous biosynthesis of quinolinic acid by inhibiting the activity of liver tryptophan-2,3-dioxygenase, intestinal indoleamine-2,3-dioxygenase and rat liver 3-hydroxyanthranillic acid oxygenase in vitro and in vivo, possibly through competitive inhibition of haeme, scavenging of superoxide anion and binding of iron (II) respectively. An inverse relationship exists between tryptophan-2,3-dioxygenase activity and brain serotonin levels. Acyclovir administration in rats induces a rise in forebrain serotonin and 5-hydroxyindole acetic acid and reduces the turnover of forebrain serotonin to 5-hydroxyindole acetic acid. Furthermore, it shows that acyclovir does not alter forebrain norepinephrine levels. The results of the pineal indole metabolism study show that acyclovir increases 5-hydroxytryptophol, N-acetylserotonin and the neurohormone melatonin, but decreases 5-hydroxyindole acetic acid. The results of this study show that acyclovir has some neuroprotective properties which may make it useful in the alleviation of the anomalous neurobiology in neurodegenerative disorders.
- Full Text:
- Date Issued: 2006
- Authors: Müller, Adrienne Carmel
- Date: 2006
- Subjects: Acyclovir -- Therapeutic use , Acyclovir -- Physiological effect , Nervous system -- Degeneration -- Treatment , Memory disorders -- Treatment , Quinolinic acid
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3776 , http://hdl.handle.net/10962/d1003254 , Acyclovir -- Therapeutic use , Acyclovir -- Physiological effect , Nervous system -- Degeneration -- Treatment , Memory disorders -- Treatment , Quinolinic acid
- Description: Accumulating evidence suggests that quinolinic acid has a role to play in disorders involving impairment of learning and memory. In the present study, the effect of the guanosine analogue antiherpetic, acyclovir, on quinolinic acid-induced spatial memory deficits was investigated, as well as some of the mechanisms which underlie this effect. Behavioural studies using a Morris water maze show that post-treatment of rats with acyclovir significantly improves spatial memory deficits induced by intrahippocampal injections of quinolinic acid. Histological analysis of the hippocampi show that the effect of acyclovir is related to its ability to alleviate quinolinic acid-induced necrotic cell death, through interference with some of the mechanisms of neurodegeneration. However, acyclovir is unable to alter a quinolinic acid-induced increase in glutamate release in the rat hippocampus, even though it alleviates quinolinic acid induced oxidative stress by scavenging the superoxide anion in vitro and in vivo in whole rat brain and hippocampus respectively. Due to the inverse relationship which exists between superoxide anion and glutathione levels, acyclovir also curtails the quinolinic acid-induced decrease in hippocampal glutathione levels. Acyclovir suppresses quinolinic acid-induced lipid peroxidation in vitro and in vivo, in whole rat brain and hippocampus respectively, through its alleviation of oxidative stress and possibly through the binding of iron (II) and / or iron (III), preventing the participation and redox recycling of iron (II) in the Fenton reaction, which quinolinic acid is thought to enhance by weak binding of ferrous ions. This argument is further strengthened by the ability of the drug to suppress iron (II)-induced lipid peroxidation in vitro directly. Inorganic studies including ultraviolet and visible spectroscopy, electrochemistry and the ferrozine assay show that acyclovir binds to iron (II) and iron (III) and that quinolinic acid forms an easily oxidisable association with iron (II). Acyclovir inhibits the endogenous biosynthesis of quinolinic acid by inhibiting the activity of liver tryptophan-2,3-dioxygenase, intestinal indoleamine-2,3-dioxygenase and rat liver 3-hydroxyanthranillic acid oxygenase in vitro and in vivo, possibly through competitive inhibition of haeme, scavenging of superoxide anion and binding of iron (II) respectively. An inverse relationship exists between tryptophan-2,3-dioxygenase activity and brain serotonin levels. Acyclovir administration in rats induces a rise in forebrain serotonin and 5-hydroxyindole acetic acid and reduces the turnover of forebrain serotonin to 5-hydroxyindole acetic acid. Furthermore, it shows that acyclovir does not alter forebrain norepinephrine levels. The results of the pineal indole metabolism study show that acyclovir increases 5-hydroxytryptophol, N-acetylserotonin and the neurohormone melatonin, but decreases 5-hydroxyindole acetic acid. The results of this study show that acyclovir has some neuroprotective properties which may make it useful in the alleviation of the anomalous neurobiology in neurodegenerative disorders.
- Full Text:
- Date Issued: 2006
An investigation into the neuroprotective properties of the non-steroidal anti-inflammatory agents tolmetin, sulindac and turmeric
- Authors: Dairam, Amichand
- Date: 2006
- Subjects: Nonsteroidal anti-inflammatory agents Antioxidants Tolmetin -- Therapeutic use Sulindac -- Therapeutic use Turmeric -- Therapeutic use Nervous system -- Degeneration -- Prevention Alzheimer's disease
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3752 , http://hdl.handle.net/10962/d1003230
- Description: Accumulating evidence suggests that anti-inflammatory agents and antioxidants have neuroprotective properties and may be beneficial in the treatment of neurodegenerative disorders. In the present study, the possible neuroprotective properties of tolmetin, sulindac and turmeric were investigated. The antioxidant effects of tolmetin and sulindac were determined by inducing free radical generation with quinolinic acid (QA), cyanide or iron (II) in rat brain homogenates or primary hippocampal neurons. Tolmetin and sulindac significantly reduce lipid peroxidation and scavenge the superoxide anion. Metal binding studies were conducted to determine whether metal chelation is a possible mechanism through which these agents reduce QA and iron (II)-induced lipid peroxidation. UV/VIS, infrared spectroscopy as well as electrochemical studies show that both agents bind to iron (II) and/or iron (III). Histological examination of the hippocampus showed that pre-treatment of animals with tolmetin or sulindac offers protection against intrahippocampal injections of QA. These agents also attenuate QA-induced apoptosis and reduce the loss of neurons in the hippocampus. The co-incubation of primary hippocampal neurons with the NSAIDS also enhanced cell viability which is significantly reduced by QA. Behavioural studies using a water maze showed that the treatment of animals after QA-induced neurotoxicity reduces QA-induced spatial memory loss. Tolmetin and sulindac also reduced glutathione depletion and protein oxidation in rat hippocampus. Both NSAIDS inhibit liver tryptophan 2,3-dioxygenase activity in vitro and in vivo and subsequently increased hippocampal serotonin levels. However, both NSAIDS also reduce dopamine levels in rat striatum. Tolmetin but not sulindac increased the synthesis of melatonin by the pineal gland. The active components of turmeric known as the curcuminoids were separated using preparative thin layer chromatography (TLC). The purity was confirmed by TLC, NMR and mass spectrometry. The environmental toxin lead, induces lipid peroxidation and reduces primary hippocampal neuronal viability. The co-incubation of the neurons with the curcuminoids significantly reduces lead-induced lipid peroxidation and enhances neuronal cell viability in the presence of lead. Lead-induced spatial memory deficit is also attenuated with curcumin, demethoxycurcumin but not bisdemethoxycurcumin. The curcuminoids also reduce lead-induced hippocampal glutathione depletion and protein oxidation. Metal binding studies show that the curcuminoids bind to lead and is another possible mechanism through which the curcuminoids reduce lead-induced neurotoxicity. The findings of this study indicate a possible role of tolmetin, sulindac and turmeric in neurodegenerative disorders such as Alzheimer’s disease. However, tolmetin and sulindac reduce dopamine levels.
- Full Text:
- Date Issued: 2006
- Authors: Dairam, Amichand
- Date: 2006
- Subjects: Nonsteroidal anti-inflammatory agents Antioxidants Tolmetin -- Therapeutic use Sulindac -- Therapeutic use Turmeric -- Therapeutic use Nervous system -- Degeneration -- Prevention Alzheimer's disease
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3752 , http://hdl.handle.net/10962/d1003230
- Description: Accumulating evidence suggests that anti-inflammatory agents and antioxidants have neuroprotective properties and may be beneficial in the treatment of neurodegenerative disorders. In the present study, the possible neuroprotective properties of tolmetin, sulindac and turmeric were investigated. The antioxidant effects of tolmetin and sulindac were determined by inducing free radical generation with quinolinic acid (QA), cyanide or iron (II) in rat brain homogenates or primary hippocampal neurons. Tolmetin and sulindac significantly reduce lipid peroxidation and scavenge the superoxide anion. Metal binding studies were conducted to determine whether metal chelation is a possible mechanism through which these agents reduce QA and iron (II)-induced lipid peroxidation. UV/VIS, infrared spectroscopy as well as electrochemical studies show that both agents bind to iron (II) and/or iron (III). Histological examination of the hippocampus showed that pre-treatment of animals with tolmetin or sulindac offers protection against intrahippocampal injections of QA. These agents also attenuate QA-induced apoptosis and reduce the loss of neurons in the hippocampus. The co-incubation of primary hippocampal neurons with the NSAIDS also enhanced cell viability which is significantly reduced by QA. Behavioural studies using a water maze showed that the treatment of animals after QA-induced neurotoxicity reduces QA-induced spatial memory loss. Tolmetin and sulindac also reduced glutathione depletion and protein oxidation in rat hippocampus. Both NSAIDS inhibit liver tryptophan 2,3-dioxygenase activity in vitro and in vivo and subsequently increased hippocampal serotonin levels. However, both NSAIDS also reduce dopamine levels in rat striatum. Tolmetin but not sulindac increased the synthesis of melatonin by the pineal gland. The active components of turmeric known as the curcuminoids were separated using preparative thin layer chromatography (TLC). The purity was confirmed by TLC, NMR and mass spectrometry. The environmental toxin lead, induces lipid peroxidation and reduces primary hippocampal neuronal viability. The co-incubation of the neurons with the curcuminoids significantly reduces lead-induced lipid peroxidation and enhances neuronal cell viability in the presence of lead. Lead-induced spatial memory deficit is also attenuated with curcumin, demethoxycurcumin but not bisdemethoxycurcumin. The curcuminoids also reduce lead-induced hippocampal glutathione depletion and protein oxidation. Metal binding studies show that the curcuminoids bind to lead and is another possible mechanism through which the curcuminoids reduce lead-induced neurotoxicity. The findings of this study indicate a possible role of tolmetin, sulindac and turmeric in neurodegenerative disorders such as Alzheimer’s disease. However, tolmetin and sulindac reduce dopamine levels.
- Full Text:
- Date Issued: 2006
Application of CE, HPLC and LC-MS-MS for the analysis and quality control of Ginkgo biloba dosage forms
- Authors: Dubber, Mary-Jean
- Date: 2006
- Subjects: Pharmaceutical chemistry -- Quality control Ginkgo Micelles Capillary electrophoresis High performance liquid chromatography Drugs -- Dosage forms Flavonoids Terpenes Herbals
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3757 , http://hdl.handle.net/10962/d1003235
- Description: Natural products are complex mixtures of compounds with therapeutic effects which are often reported to be due to the synergistic action of multiple and sometimes unknown components. Consequently, standardization of these products is complex and a lack of effective quality control (QC) criteria in most countries has led to marketing of commercial products with questionable quality, safety and efficacy (QSE). The aim of this study was therefore to develop qualitative and quantitative analytical methods for use in the QC of Ginkgo biloba solid oral dosage forms. Initially, a micellar electrokinetic chromatography (MEKC) method was developed for the identification of the flavonol glycosides, rutin and quercitrin as well as 3 flavonol aglycones, quercetin, kaempferol and isorhamnetin in crude extracts of 4 Ginkgo biloba solid oral dosage forms using ultraviolet (UV) detection. A reversed-flow cyclodextrin-modified MEKC method was subsequently developed for the simultaneous determination of the aforementioned flavonols as well as ginkgolide A, B, C, J and bilobalide (all positive markers) in Ginkgo commercial products. A non-aqueous capillary electrophoresis (CE) method was also developed for fingerprinting the presence of ginkgolic acids (negative markers) in Ginkgo biloba leaf extracts, which are purported to be associated with toxic properties. This method was also applied to 2 Ginkgo biloba commercial products. Since the flavonols have strong UV absorbing chromophores, a reversed phase high-performance liquid chromatographic (RP-HPLC) method was developed and validated using photo-diode-array (PDA) detection which was then successfully applied to fingerprint commercially available Ginkgo biloba solid oral dosage forms as well as quantify the relevant flavonol markers present in these extracts. Sample preparation was simple, rapid and cost efficient with minimal clean-up and the employment of a minibore column which requires low mobile phase flow rates contributed to the economy of the method. Unlike the conventional QC approach, samples were not hydrolyzed and direct determination of 2 intact flavonol glycosides, together with the usual aglycone markers was facilitated which provided maximal content information for fingerprint comparisons. On the other hand, terpene trilactones possess poor chromophores and an alternative detection method to UV was required in order to obtain suitable sensitivity. RP-HPLC with evaporative light scattering detection (ELSD) was selected for quantification of these non-volatile constituents in Ginkgo dosage forms and this method was deemed suitable for the routine QC analysis of these positive markers in commercial products. Since approximately 33 flavonoids have been identified in Ginkgo biloba leaf extracts, baseline separation using UV/PDA detection normally requires complex gradient programs and long analysis times. In addition, unequivocal identification of the flavonoids with similar UV spectra and elution times cannot be guaranteed. A liquid chromatographic tandem mass spectrometric (LC-MS-MS) method was therefore developed and validated in order to ensure accurate quantification of the selected flavonol marker compounds in Ginkgo commercial products. LC-MS-MS analysis of Ginkgo extracts revealed, in addition to rutin, the possible presence of other quercetin analogues, quercetin-3-Orhamnoside-7-O-glucoside or quercetin-3-O-glucoside-7-O-rhamnoside, previously unreported in Ginkgo biloba leaf extracts or dosage forms. In terms of evaluating the most suitable analytical method for QC, CE shows exceptional potential in the future analysis of Ginkgo biloba dosage forms while HPLC-PDA and HPLC-ELSD are currently the most affordable and practical instruments for the routine analysis of the flavonols and terpenoids, respectively. LC-MS-MS proved to be pivotal for the accurate identification and quantification of the flavonols due to interference by other flavonoid compounds with similar retention times and UV spectra to the peaks of interest. All quantitative and qualitative results revealed large discrepancies in the marker content between the products regardless of which batch was analysed and product labels disclosed little relevant information. Although currently not required by most regulatory agencies, some of the usual quality criteria applied to orthodox medicines was evaluated. In particular, dissolution analysis, disintegration, tablet hardness and weight uniformity were assessed and revealed similar inconsistencies. This thesis emphasises that implementation of effective QC criteria is long overdue and is essential to ensure consistent product QSE of commercially available Ginkgo biloba solid oral dosage forms.
- Full Text:
- Date Issued: 2006
- Authors: Dubber, Mary-Jean
- Date: 2006
- Subjects: Pharmaceutical chemistry -- Quality control Ginkgo Micelles Capillary electrophoresis High performance liquid chromatography Drugs -- Dosage forms Flavonoids Terpenes Herbals
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3757 , http://hdl.handle.net/10962/d1003235
- Description: Natural products are complex mixtures of compounds with therapeutic effects which are often reported to be due to the synergistic action of multiple and sometimes unknown components. Consequently, standardization of these products is complex and a lack of effective quality control (QC) criteria in most countries has led to marketing of commercial products with questionable quality, safety and efficacy (QSE). The aim of this study was therefore to develop qualitative and quantitative analytical methods for use in the QC of Ginkgo biloba solid oral dosage forms. Initially, a micellar electrokinetic chromatography (MEKC) method was developed for the identification of the flavonol glycosides, rutin and quercitrin as well as 3 flavonol aglycones, quercetin, kaempferol and isorhamnetin in crude extracts of 4 Ginkgo biloba solid oral dosage forms using ultraviolet (UV) detection. A reversed-flow cyclodextrin-modified MEKC method was subsequently developed for the simultaneous determination of the aforementioned flavonols as well as ginkgolide A, B, C, J and bilobalide (all positive markers) in Ginkgo commercial products. A non-aqueous capillary electrophoresis (CE) method was also developed for fingerprinting the presence of ginkgolic acids (negative markers) in Ginkgo biloba leaf extracts, which are purported to be associated with toxic properties. This method was also applied to 2 Ginkgo biloba commercial products. Since the flavonols have strong UV absorbing chromophores, a reversed phase high-performance liquid chromatographic (RP-HPLC) method was developed and validated using photo-diode-array (PDA) detection which was then successfully applied to fingerprint commercially available Ginkgo biloba solid oral dosage forms as well as quantify the relevant flavonol markers present in these extracts. Sample preparation was simple, rapid and cost efficient with minimal clean-up and the employment of a minibore column which requires low mobile phase flow rates contributed to the economy of the method. Unlike the conventional QC approach, samples were not hydrolyzed and direct determination of 2 intact flavonol glycosides, together with the usual aglycone markers was facilitated which provided maximal content information for fingerprint comparisons. On the other hand, terpene trilactones possess poor chromophores and an alternative detection method to UV was required in order to obtain suitable sensitivity. RP-HPLC with evaporative light scattering detection (ELSD) was selected for quantification of these non-volatile constituents in Ginkgo dosage forms and this method was deemed suitable for the routine QC analysis of these positive markers in commercial products. Since approximately 33 flavonoids have been identified in Ginkgo biloba leaf extracts, baseline separation using UV/PDA detection normally requires complex gradient programs and long analysis times. In addition, unequivocal identification of the flavonoids with similar UV spectra and elution times cannot be guaranteed. A liquid chromatographic tandem mass spectrometric (LC-MS-MS) method was therefore developed and validated in order to ensure accurate quantification of the selected flavonol marker compounds in Ginkgo commercial products. LC-MS-MS analysis of Ginkgo extracts revealed, in addition to rutin, the possible presence of other quercetin analogues, quercetin-3-Orhamnoside-7-O-glucoside or quercetin-3-O-glucoside-7-O-rhamnoside, previously unreported in Ginkgo biloba leaf extracts or dosage forms. In terms of evaluating the most suitable analytical method for QC, CE shows exceptional potential in the future analysis of Ginkgo biloba dosage forms while HPLC-PDA and HPLC-ELSD are currently the most affordable and practical instruments for the routine analysis of the flavonols and terpenoids, respectively. LC-MS-MS proved to be pivotal for the accurate identification and quantification of the flavonols due to interference by other flavonoid compounds with similar retention times and UV spectra to the peaks of interest. All quantitative and qualitative results revealed large discrepancies in the marker content between the products regardless of which batch was analysed and product labels disclosed little relevant information. Although currently not required by most regulatory agencies, some of the usual quality criteria applied to orthodox medicines was evaluated. In particular, dissolution analysis, disintegration, tablet hardness and weight uniformity were assessed and revealed similar inconsistencies. This thesis emphasises that implementation of effective QC criteria is long overdue and is essential to ensure consistent product QSE of commercially available Ginkgo biloba solid oral dosage forms.
- Full Text:
- Date Issued: 2006
Competing interests and change within the pharmacy education system in South Africa
- Authors: Allan, Lucie
- Date: 2006
- Subjects: Pharmacy -- South Africa Pharmacy -- Study and teaching -- South Africa Pharmacy -- Practice -- South Africa Community pharmacy services -- South Africa Community health services -- South Africa
- Language: English
- Type: Thesis , Masters , MPharm
- Identifier: vital:3741 , http://hdl.handle.net/10962/d1003219
- Description: This thesis provides a historical account of the emergence of the pharmacy education system in South Africa, and an analysis of the influence of competing interest groups over the pharmacy education curriculum. It provides a critical evaluation of structural-consensus and micro-interpretive approaches to medical and pharmacy education, and sets out a macrointerpretive account of pharmacy education in South Africa. Following Margaret Archer (1979) it analyzes three forms of negotiation between competing interest groups in their efforts to change the pharmacy curriculum; these are political manipulation, external transaction and internal initiation. The thesis argues that whilst the private sector interest group (comprising of retail, wholesale and manufacturing pharmacy) dominated the pharmacy education system until 1994, since then a newly emerged government interest group has begun to compete for educational control. The priorities pursued by this interest group have consistently reflected the objectives set out in the ANC National Health Plan of 1994. The thesis maintains that given its frustration over the non-implementation of the ANC’s health policy objectives, the government interest group is likely to resort to direct political manipulation by passing legislation to alter the content of the current pharmacy curriculum. Such changes would seek to ensure that the syllabus more accurately reflects the ANC Plan’s community health and primary health care objectives. The thesis asserts that such an outcome (of direct political manipulation of the curriculum) is not inevitable, and can be avoided through a process of internally initiated change. It presents the findings of an interpretive case study into how the Rhodes University Community Experience Programme (CEP) influenced final year pharmacy students’ perceptions of the role of the pharmacist. The students’ comments were collected by means of focus group interviews, participant observation and documentary analysis. Whilst the CEP did not successfully transform their concept of the pharmacist’s role, it did succeed in influencing students’ understanding of the notions of community pharmacy and primary health care in line with the government interest group’s health objectives. This thesis concludes that internally initiated change within the pharmacy education system, would be preferable to that imposed through external political manipulation, as such change would be more likely to preserve the independent professional interests of pharmacy academics.
- Full Text:
- Date Issued: 2006
- Authors: Allan, Lucie
- Date: 2006
- Subjects: Pharmacy -- South Africa Pharmacy -- Study and teaching -- South Africa Pharmacy -- Practice -- South Africa Community pharmacy services -- South Africa Community health services -- South Africa
- Language: English
- Type: Thesis , Masters , MPharm
- Identifier: vital:3741 , http://hdl.handle.net/10962/d1003219
- Description: This thesis provides a historical account of the emergence of the pharmacy education system in South Africa, and an analysis of the influence of competing interest groups over the pharmacy education curriculum. It provides a critical evaluation of structural-consensus and micro-interpretive approaches to medical and pharmacy education, and sets out a macrointerpretive account of pharmacy education in South Africa. Following Margaret Archer (1979) it analyzes three forms of negotiation between competing interest groups in their efforts to change the pharmacy curriculum; these are political manipulation, external transaction and internal initiation. The thesis argues that whilst the private sector interest group (comprising of retail, wholesale and manufacturing pharmacy) dominated the pharmacy education system until 1994, since then a newly emerged government interest group has begun to compete for educational control. The priorities pursued by this interest group have consistently reflected the objectives set out in the ANC National Health Plan of 1994. The thesis maintains that given its frustration over the non-implementation of the ANC’s health policy objectives, the government interest group is likely to resort to direct political manipulation by passing legislation to alter the content of the current pharmacy curriculum. Such changes would seek to ensure that the syllabus more accurately reflects the ANC Plan’s community health and primary health care objectives. The thesis asserts that such an outcome (of direct political manipulation of the curriculum) is not inevitable, and can be avoided through a process of internally initiated change. It presents the findings of an interpretive case study into how the Rhodes University Community Experience Programme (CEP) influenced final year pharmacy students’ perceptions of the role of the pharmacist. The students’ comments were collected by means of focus group interviews, participant observation and documentary analysis. Whilst the CEP did not successfully transform their concept of the pharmacist’s role, it did succeed in influencing students’ understanding of the notions of community pharmacy and primary health care in line with the government interest group’s health objectives. This thesis concludes that internally initiated change within the pharmacy education system, would be preferable to that imposed through external political manipulation, as such change would be more likely to preserve the independent professional interests of pharmacy academics.
- Full Text:
- Date Issued: 2006
Development and assessment of azithromycin paediatric suppository formulations
- Authors: Mollel, Happiness
- Date: 2006
- Subjects: Azithromycin , Pediatrics , Clinical pharmacology , Pharmacokinetics , Suppositories , Drugs -- Dosage forms
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3774 , http://hdl.handle.net/10962/d1003252 , Azithromycin , Pediatrics , Clinical pharmacology , Pharmacokinetics , Suppositories , Drugs -- Dosage forms
- Description: The use of the oral route of administration for the treatment of young children with antibiotics can at times be problematic since, factors such as nausea, vomiting, taste and/or smell, in addition to the challenges associated with the administration of suspensions, may contribute to poor patient compliance. In such cases, the use of the rectal route of administration may be appropriate. Therefore, suppositories containing 250 mg azithromycin (AZI) were manufactured and assessed for potential as an antibiotic suppository dosage form. Suppositories, containing AZI dihydrate were manufactured by the fusion method, using different grades of PEG, Witepsol® and Suppocire® bases. The rate and extent of AZI release was evaluated using USP apparatus I, and samples were analyzed using a validated HPLC method. Differences in the rate and extent of AZI release were observed with the greatest amount of AZI being released from PEG formulations. The rate and extent of AZI release from formulations manufactured using fatty bases were influenced by physicochemical properties, such as melting rate and hydroxyl value, of the bases. In addition drug partitioning appeared to favor the lipid phase and had a negative impact on AZI release characteristics. Two different formulation approaches were used in an attempt to increase the rate and extent of AZI release from fatty base formulations. The use of surfactants significantly increased AZI release from formulations manufactured with fatty bases with high hydroxyl values. The use of urea or Povidone K25 in combination with AZI as a physical mixture or solid dispersion did not increase the rate and extent of AZI release from the fatty suppositories, to any significant extent. The mechanism of drug release was evaluated using several mathematical models, including the Higuchi, Korsmeyer- eppas, Zero and, First order models. In addition, in vitro dissolution profiles were characterized by the difference and similarity factors, f1 and f2 and by use of the Gohel similarity factor, Sd. AZI release kinetics were best described by the Higuchi and Korsmeyer-Peppas models and the values of the release exponent, n, revealed that drug release was a consequence of the combined effects of AZI diffusion, rate of melting of the base and partitioning of the drug which can be considered to be anomalous release.
- Full Text:
- Date Issued: 2006
- Authors: Mollel, Happiness
- Date: 2006
- Subjects: Azithromycin , Pediatrics , Clinical pharmacology , Pharmacokinetics , Suppositories , Drugs -- Dosage forms
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3774 , http://hdl.handle.net/10962/d1003252 , Azithromycin , Pediatrics , Clinical pharmacology , Pharmacokinetics , Suppositories , Drugs -- Dosage forms
- Description: The use of the oral route of administration for the treatment of young children with antibiotics can at times be problematic since, factors such as nausea, vomiting, taste and/or smell, in addition to the challenges associated with the administration of suspensions, may contribute to poor patient compliance. In such cases, the use of the rectal route of administration may be appropriate. Therefore, suppositories containing 250 mg azithromycin (AZI) were manufactured and assessed for potential as an antibiotic suppository dosage form. Suppositories, containing AZI dihydrate were manufactured by the fusion method, using different grades of PEG, Witepsol® and Suppocire® bases. The rate and extent of AZI release was evaluated using USP apparatus I, and samples were analyzed using a validated HPLC method. Differences in the rate and extent of AZI release were observed with the greatest amount of AZI being released from PEG formulations. The rate and extent of AZI release from formulations manufactured using fatty bases were influenced by physicochemical properties, such as melting rate and hydroxyl value, of the bases. In addition drug partitioning appeared to favor the lipid phase and had a negative impact on AZI release characteristics. Two different formulation approaches were used in an attempt to increase the rate and extent of AZI release from fatty base formulations. The use of surfactants significantly increased AZI release from formulations manufactured with fatty bases with high hydroxyl values. The use of urea or Povidone K25 in combination with AZI as a physical mixture or solid dispersion did not increase the rate and extent of AZI release from the fatty suppositories, to any significant extent. The mechanism of drug release was evaluated using several mathematical models, including the Higuchi, Korsmeyer- eppas, Zero and, First order models. In addition, in vitro dissolution profiles were characterized by the difference and similarity factors, f1 and f2 and by use of the Gohel similarity factor, Sd. AZI release kinetics were best described by the Higuchi and Korsmeyer-Peppas models and the values of the release exponent, n, revealed that drug release was a consequence of the combined effects of AZI diffusion, rate of melting of the base and partitioning of the drug which can be considered to be anomalous release.
- Full Text:
- Date Issued: 2006
Development and assessment of propranolol sustained release dosage forms separately and in combination with hydrochlorothiazide
- Authors: Chetty, Prakash
- Date: 2006
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3749 , http://hdl.handle.net/10962/d1003227
- Description: Hypertension is a chronic illness that is often undiagnosed and untreated leading to high mortality rates in South Africa. The use of diuretics such as hydrochlorothiazide and beta blockers such as propranolol has been advocated as first line therapy for the treatment of hypertension. The study and use of controlled release dosage forms for the treatment of various disease states has gained wide interest over the past two decades. The use of controlled release systems offers improved therapeutic efficiency over conventional immediate release dosage forms, the use of which at times have often led to poor patient adherence and decreased therapeutic efficiencies. The current research objective was to develop a sustained release multi-source product for propranolol such that once daily dosing would be achieved. In addition, the sustained release product was developed using Inderal® LA 80mg capsules as a reference product. In addition the development of a suitable immediate release hydrochlorothiazide tablet was undertaken to produce a combination dosage form. The use of two different technologies, namely direct compression and wet granulation were employed to develop the sustained release dosage form. The release of propranolol from these dosage forms was assessed using USP apparatus 1 with quantitation of the relevant dissolution samples using a validated high performance liquid chromatographic method. The release profiles from the prototype and subsequent products were subjected to model independent and model dependent analyses in order to compare them to the innovator product and to elucidate the mechanisms of drug release respectively. Dissolution test results reveal that dosage forms prepared from wet granulation showed better rate retardation and more appropriate release profiles than those prepared by direct compression techniques. The subsequent model independent and model dependent analysis show that a dosage form that is comparable to the innovator product has been developed, with drug release occurring by a diffusion type mechanism.
- Full Text:
- Date Issued: 2006
- Authors: Chetty, Prakash
- Date: 2006
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3749 , http://hdl.handle.net/10962/d1003227
- Description: Hypertension is a chronic illness that is often undiagnosed and untreated leading to high mortality rates in South Africa. The use of diuretics such as hydrochlorothiazide and beta blockers such as propranolol has been advocated as first line therapy for the treatment of hypertension. The study and use of controlled release dosage forms for the treatment of various disease states has gained wide interest over the past two decades. The use of controlled release systems offers improved therapeutic efficiency over conventional immediate release dosage forms, the use of which at times have often led to poor patient adherence and decreased therapeutic efficiencies. The current research objective was to develop a sustained release multi-source product for propranolol such that once daily dosing would be achieved. In addition, the sustained release product was developed using Inderal® LA 80mg capsules as a reference product. In addition the development of a suitable immediate release hydrochlorothiazide tablet was undertaken to produce a combination dosage form. The use of two different technologies, namely direct compression and wet granulation were employed to develop the sustained release dosage form. The release of propranolol from these dosage forms was assessed using USP apparatus 1 with quantitation of the relevant dissolution samples using a validated high performance liquid chromatographic method. The release profiles from the prototype and subsequent products were subjected to model independent and model dependent analyses in order to compare them to the innovator product and to elucidate the mechanisms of drug release respectively. Dissolution test results reveal that dosage forms prepared from wet granulation showed better rate retardation and more appropriate release profiles than those prepared by direct compression techniques. The subsequent model independent and model dependent analysis show that a dosage form that is comparable to the innovator product has been developed, with drug release occurring by a diffusion type mechanism.
- Full Text:
- Date Issued: 2006
Patient education : the effect on patient behaviour
- Authors: Shiri, Clarris
- Date: 2006
- Subjects: Patient education -- South Africa -- Eastern Cape Patient compliance -- South Africa -- Eastern Cape Hypertension -- Treatment -- South Africa Health care services -- South Africa Community health services -- South Africa
- Language: English
- Type: Thesis , Masters , MPharm
- Identifier: vital:3790 , http://hdl.handle.net/10962/d1003268
- Description: Evidence suggests that the prevalence of certain non-communicable diseases, such as hypertension, is increasing rapidly, and that patients with these diseases are making significant demands on the health services of the nations in sub-Saharan Africa. However, these countries also face other health-related challenges such as communicable diseases and underdevelopmentrelated diseases. Developing countries like South Africa have limited resources, in terms of man power and financial capital, to address the challenges that they are facing. Non-communicable diseases cannot be ignored and since health care providers cannot meet the challenges, it is worthwhile to empower patients to be involved in the management of their conditions. Patient education is a tool that can be used to enable patients to manage their chronic conditions and thereby reduce the morbidity and mortality rates of these conditions. The aim of this study was to investigate the effect of a patient education intervention on participants’ levels of knowledge about hypertension and its therapy, beliefs about medicines and adherence to anti-hypertensive therapy. The intervention consisted of talks and discussions with all the participants as one group and as individuals. There was also written information given to the participants. Their levels of knowledge about hypertension and its therapy were measured using one-on-one interviews and self-administered questionnaires. Beliefs about medicines were measured using the Beliefs about Medicines Questionnaire (BMQ) whilst adherence levels were measured using pill counts, elf-reports and prescription refill records. The participants’ blood pressure readings and body mass indices were also recorded throughout the study. The parameters before and after the educational intervention were compared using statistical analyses. The participants’ levels of knowledge about hypertension and its therapy significantly increased whilst their beliefs about medicines were positively modified after the educational intervention. There were also increases, though not statistically significant, in the participants’ levels of adherence to anti-hypertensive therapy. Unexpectedly, the blood pressure readings and body mass indices increased significantly. The participants gave positive feedback regarding the educational intervention and indicated a desire for similar programmes to be run continuously. They also suggested that such programmes be implemented for other common chronic conditions such as asthma and diabetes. This study proved that patient education programmes can be implemented to modify patients’ levels of knowledge about their conditions and the therapy, beliefs about medicines and adherence to therapy. However, such programmes need to be conducted over a long period of time since changes involving behaviour take a long time.
- Full Text:
- Date Issued: 2006
- Authors: Shiri, Clarris
- Date: 2006
- Subjects: Patient education -- South Africa -- Eastern Cape Patient compliance -- South Africa -- Eastern Cape Hypertension -- Treatment -- South Africa Health care services -- South Africa Community health services -- South Africa
- Language: English
- Type: Thesis , Masters , MPharm
- Identifier: vital:3790 , http://hdl.handle.net/10962/d1003268
- Description: Evidence suggests that the prevalence of certain non-communicable diseases, such as hypertension, is increasing rapidly, and that patients with these diseases are making significant demands on the health services of the nations in sub-Saharan Africa. However, these countries also face other health-related challenges such as communicable diseases and underdevelopmentrelated diseases. Developing countries like South Africa have limited resources, in terms of man power and financial capital, to address the challenges that they are facing. Non-communicable diseases cannot be ignored and since health care providers cannot meet the challenges, it is worthwhile to empower patients to be involved in the management of their conditions. Patient education is a tool that can be used to enable patients to manage their chronic conditions and thereby reduce the morbidity and mortality rates of these conditions. The aim of this study was to investigate the effect of a patient education intervention on participants’ levels of knowledge about hypertension and its therapy, beliefs about medicines and adherence to anti-hypertensive therapy. The intervention consisted of talks and discussions with all the participants as one group and as individuals. There was also written information given to the participants. Their levels of knowledge about hypertension and its therapy were measured using one-on-one interviews and self-administered questionnaires. Beliefs about medicines were measured using the Beliefs about Medicines Questionnaire (BMQ) whilst adherence levels were measured using pill counts, elf-reports and prescription refill records. The participants’ blood pressure readings and body mass indices were also recorded throughout the study. The parameters before and after the educational intervention were compared using statistical analyses. The participants’ levels of knowledge about hypertension and its therapy significantly increased whilst their beliefs about medicines were positively modified after the educational intervention. There were also increases, though not statistically significant, in the participants’ levels of adherence to anti-hypertensive therapy. Unexpectedly, the blood pressure readings and body mass indices increased significantly. The participants gave positive feedback regarding the educational intervention and indicated a desire for similar programmes to be run continuously. They also suggested that such programmes be implemented for other common chronic conditions such as asthma and diabetes. This study proved that patient education programmes can be implemented to modify patients’ levels of knowledge about their conditions and the therapy, beliefs about medicines and adherence to therapy. However, such programmes need to be conducted over a long period of time since changes involving behaviour take a long time.
- Full Text:
- Date Issued: 2006
Pharmaceutical analysis and drug interaction studies : African potato (Hypoxis hemerocallidea)
- Purushothaman Nair, Vipin Devi Prasad
- Authors: Purushothaman Nair, Vipin Devi Prasad
- Date: 2006
- Subjects: Potatoes -- Africa , Potatoes -- Therapeutic use , AIDS (Disease) -- Treatment , HIV infections -- Drug therapy , Medicinal plants
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3865 , http://hdl.handle.net/10962/d1015802
- Description: In order for a medicinal product to produce a consistent and reliable therapeutic response, it is essential that the final composition of the product is invariable and that the active ingredient/s is/are present in appropriate, non-toxic amounts. However, due to the complexity involved in the standardization of natural products, quality control (QC) criteria and procedures for the registration and market approval of such products are conspicuously absent in most countries around the world. African Potato (AP) is of great medical interest and this particular plant has gained tremendous popularity following the endorsement by the South African Minister of Health as a remedy for HIV/ AIDS patients. Very little information has appeared in the literature to describe methods for the quantitative analysis of hypoxoside, an important component in AP. It has also been claimed that sterols and sterolins present in AP are responsible for its medicinal property but is yet to be proven scientifically. To-date, no QC methods have been reported for the simultaneous quantitative analysis of the combination, β- sitosterol (BSS)/ stigmasterol (STG)/ stigmastanol (STN), purported to be present in preparations containing AP. The effect of concomitant administration of AP and other herbal medicines on the safety and efficacy of conventional medicines has not yet been fully determined. Amongst the objectives of this study was to develop and validate quantitative analytical methods that are suitable for the assay and quality control of plant material, extracts and commercial formulations containing AP. Hypoxoside was isolated from AP and characterized for use as a reference standard for the quality control of AP products and a stability-indicating HPLC/ UV assay method for the quantitative determination of hypoxoside was developed. In addition, a quantitative capillary zone electrophoretic (CZE) method was developed to determine hypoxoside, specifically for its advantages over HPLC. A HPLC method was also developed and validated for the quantitative analysis of BSS, STG and STN in commercially available oral dosage forms containing AP material or extracts thereof. The antioxidant activity of an aqueous extract of lyophilized corms of AP along with hypoxoside and rooperol were investigated. In comparison with the AP extracts and also with hypoxoside, rooperol showed significant antioxidant activity. The capacity of AP, (extracts, formulations, hypoxoside and rooperol as well as sterols to inhibit in vitro metabolism of drug substrates by human cytochrome P450 (CYP) enzymes such as CYP 3A4, 3A5 and CYP19 were investigated. Samples were also assessed for their effect on drug transport proteins such as P-glycoprotein (P-gp). Various extracts of AP, AP formulations, stigmasterol and the norlignans, in particular the aglycone rooperol, exhibited inhibitory effects on CYP 3A4, 3A5 and CYP19 mediated metabolism.These results suggest that concurrent therapy with AP and other medicines, in particular antiretroviral drugs, can have important implications for safety and efficacy. Large discrepancies in marker content between AP products were found. Dissolution testing of AP products was investigated as a QC tool and the results also revealed inconsistencies between different AP products.
- Full Text:
- Date Issued: 2006
- Authors: Purushothaman Nair, Vipin Devi Prasad
- Date: 2006
- Subjects: Potatoes -- Africa , Potatoes -- Therapeutic use , AIDS (Disease) -- Treatment , HIV infections -- Drug therapy , Medicinal plants
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3865 , http://hdl.handle.net/10962/d1015802
- Description: In order for a medicinal product to produce a consistent and reliable therapeutic response, it is essential that the final composition of the product is invariable and that the active ingredient/s is/are present in appropriate, non-toxic amounts. However, due to the complexity involved in the standardization of natural products, quality control (QC) criteria and procedures for the registration and market approval of such products are conspicuously absent in most countries around the world. African Potato (AP) is of great medical interest and this particular plant has gained tremendous popularity following the endorsement by the South African Minister of Health as a remedy for HIV/ AIDS patients. Very little information has appeared in the literature to describe methods for the quantitative analysis of hypoxoside, an important component in AP. It has also been claimed that sterols and sterolins present in AP are responsible for its medicinal property but is yet to be proven scientifically. To-date, no QC methods have been reported for the simultaneous quantitative analysis of the combination, β- sitosterol (BSS)/ stigmasterol (STG)/ stigmastanol (STN), purported to be present in preparations containing AP. The effect of concomitant administration of AP and other herbal medicines on the safety and efficacy of conventional medicines has not yet been fully determined. Amongst the objectives of this study was to develop and validate quantitative analytical methods that are suitable for the assay and quality control of plant material, extracts and commercial formulations containing AP. Hypoxoside was isolated from AP and characterized for use as a reference standard for the quality control of AP products and a stability-indicating HPLC/ UV assay method for the quantitative determination of hypoxoside was developed. In addition, a quantitative capillary zone electrophoretic (CZE) method was developed to determine hypoxoside, specifically for its advantages over HPLC. A HPLC method was also developed and validated for the quantitative analysis of BSS, STG and STN in commercially available oral dosage forms containing AP material or extracts thereof. The antioxidant activity of an aqueous extract of lyophilized corms of AP along with hypoxoside and rooperol were investigated. In comparison with the AP extracts and also with hypoxoside, rooperol showed significant antioxidant activity. The capacity of AP, (extracts, formulations, hypoxoside and rooperol as well as sterols to inhibit in vitro metabolism of drug substrates by human cytochrome P450 (CYP) enzymes such as CYP 3A4, 3A5 and CYP19 were investigated. Samples were also assessed for their effect on drug transport proteins such as P-glycoprotein (P-gp). Various extracts of AP, AP formulations, stigmasterol and the norlignans, in particular the aglycone rooperol, exhibited inhibitory effects on CYP 3A4, 3A5 and CYP19 mediated metabolism.These results suggest that concurrent therapy with AP and other medicines, in particular antiretroviral drugs, can have important implications for safety and efficacy. Large discrepancies in marker content between AP products were found. Dissolution testing of AP products was investigated as a QC tool and the results also revealed inconsistencies between different AP products.
- Full Text:
- Date Issued: 2006