A critical evaluation of the human skin blanching assay and comparative bioavailability studies on topical corticosteroid preparations
- Authors: Meyer, Eric
- Date: 1989
- Subjects: Dermatopharmacology Skin, Effect of drugs on Adrenocortical hormones
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3727 , http://hdl.handle.net/10962/d1001464
- Description: Several aspects of the human skin blanching assay were evaluated in an attempt to suggest improvements in the methodology of this assay. Three trials were performed in the unoccluded application mode, using two proprietary creams containing 0,1% betamethasone (as the 17-valerate). Preliminary observations of the influence of ambient temperature and relative humidity on the blanching response did not allow definite conclusions to be drawn. Studies on the number of observers required for reliable results of comparative blanching indicated that at least two trained observers should be employed. Analyses of the results of individual volunteers demonstrated the expected biological variability, and suggest that subjects selected for trials should represent a range of blanching responses. No sex-related differences in blanching responses were found, and both arms exhibited similar sensitivity to corticosteroids. Retrospective analysis of 95 040 observations of blanching responses showed that in the unoccluded application mode blanching is lowest close to the wrist, and in the occluded mode blanching is lowest close to the elbow. Studies on the method of transportation of Betnovate preparations suggest that topical formulations should not be exposed to temperature extremes during transportation. It is proposed that patients should not transport topical formulations in the holds of ships or aircraft, and that exporters and manufacturers should make use of special transportation and storage conditions. In a study of ten topical formulations from three countries it was found that there was no trend of products from one country consistently exhibiting superior blanching to products from the other two countries, or products from one country consistently exhibiting the lowest degree of blanching, although considerable differences in blanching responses were found in some cases. Interpretation of the results of these studies demonstrated the importance of employing a combination of statistical analyses, blanching profiles and AUC values when drawing conclusions regarding comparative bioavailability. A study of the blanching profiles of Betnovate cream included in all 16 trials performed during this work indicated that this preparation behaved in a similar fashion during all trials, thereby giving credence to the results of the trials
- Full Text:
- Date Issued: 1989
- Authors: Meyer, Eric
- Date: 1989
- Subjects: Dermatopharmacology Skin, Effect of drugs on Adrenocortical hormones
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3727 , http://hdl.handle.net/10962/d1001464
- Description: Several aspects of the human skin blanching assay were evaluated in an attempt to suggest improvements in the methodology of this assay. Three trials were performed in the unoccluded application mode, using two proprietary creams containing 0,1% betamethasone (as the 17-valerate). Preliminary observations of the influence of ambient temperature and relative humidity on the blanching response did not allow definite conclusions to be drawn. Studies on the number of observers required for reliable results of comparative blanching indicated that at least two trained observers should be employed. Analyses of the results of individual volunteers demonstrated the expected biological variability, and suggest that subjects selected for trials should represent a range of blanching responses. No sex-related differences in blanching responses were found, and both arms exhibited similar sensitivity to corticosteroids. Retrospective analysis of 95 040 observations of blanching responses showed that in the unoccluded application mode blanching is lowest close to the wrist, and in the occluded mode blanching is lowest close to the elbow. Studies on the method of transportation of Betnovate preparations suggest that topical formulations should not be exposed to temperature extremes during transportation. It is proposed that patients should not transport topical formulations in the holds of ships or aircraft, and that exporters and manufacturers should make use of special transportation and storage conditions. In a study of ten topical formulations from three countries it was found that there was no trend of products from one country consistently exhibiting superior blanching to products from the other two countries, or products from one country consistently exhibiting the lowest degree of blanching, although considerable differences in blanching responses were found in some cases. Interpretation of the results of these studies demonstrated the importance of employing a combination of statistical analyses, blanching profiles and AUC values when drawing conclusions regarding comparative bioavailability. A study of the blanching profiles of Betnovate cream included in all 16 trials performed during this work indicated that this preparation behaved in a similar fashion during all trials, thereby giving credence to the results of the trials
- Full Text:
- Date Issued: 1989
A critical realist account of a mentoring programme in the Faculty of Pharmacy at Rhodes University
- Authors: Oltmann, Carmen
- Date: 2009
- Subjects: Rhodes University -- Academic Development Programme Pharmacy -- Study and teaching (Higher) -- South Africa Mentoring in education -- South Africa Mentoring in Science -- South Africa Critical realism Communities of practice
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3781 , http://hdl.handle.net/10962/d1003259
- Description: This study originates from experiences I had as supervisor of the mentoring programme for first year students in the Faculty of Pharmacy, at Rhodes University. Our mentoring programme is a strategy for first year students – specifically those from previously disadvantaged backgrounds – to succeed at Rhodes University. Using an ontological meta-theory - critical realism - as my analytical lens, discourse as my unit of analysis, and Invitational Learning Theory as a theoretical tool I developed a model of mentoring based on Bhaskar’s transformational model (1993). This model illustrates the relationship between structure, culture and agency. Whilst developing this model I focussed on determining how mentors construct mentoring, and how mentoring facilitates access to a Community of Practice (CoP). Mentoring involves providing a shared space that is safe, that the mentor and mentee feel comfortable in, and that supports and challenges both the mentor and the mentee. It is a reciprocal, developmental relationship for both the mentor and the mentee that deals with issues that the mentee deems as ‘real’. Mentoring is a process, not an outcome. The mentoring strategies that the mentors employed changed as the mentors mentored. Mentors help mentees by using structures and mechanisms that worked for them, and/or by helping mentees access these structures and mechanisms. Mentoring facilitates access to a CoP by providing opportunities for engagement. This involves sharing of experiences and knowledge, and promoting discussion. The mentor helps the mentee move from being a peripheral member of the CoP to becoming a main member, i.e., becoming active, learning with and from others within the CoP. CoPs develop social capital and knowledge management. My research suggests that the knowledge, skills and attitude developed by the mentors within this study may be transferable to other aspects in Pharmacy.
- Full Text:
- Date Issued: 2009
- Authors: Oltmann, Carmen
- Date: 2009
- Subjects: Rhodes University -- Academic Development Programme Pharmacy -- Study and teaching (Higher) -- South Africa Mentoring in education -- South Africa Mentoring in Science -- South Africa Critical realism Communities of practice
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3781 , http://hdl.handle.net/10962/d1003259
- Description: This study originates from experiences I had as supervisor of the mentoring programme for first year students in the Faculty of Pharmacy, at Rhodes University. Our mentoring programme is a strategy for first year students – specifically those from previously disadvantaged backgrounds – to succeed at Rhodes University. Using an ontological meta-theory - critical realism - as my analytical lens, discourse as my unit of analysis, and Invitational Learning Theory as a theoretical tool I developed a model of mentoring based on Bhaskar’s transformational model (1993). This model illustrates the relationship between structure, culture and agency. Whilst developing this model I focussed on determining how mentors construct mentoring, and how mentoring facilitates access to a Community of Practice (CoP). Mentoring involves providing a shared space that is safe, that the mentor and mentee feel comfortable in, and that supports and challenges both the mentor and the mentee. It is a reciprocal, developmental relationship for both the mentor and the mentee that deals with issues that the mentee deems as ‘real’. Mentoring is a process, not an outcome. The mentoring strategies that the mentors employed changed as the mentors mentored. Mentors help mentees by using structures and mechanisms that worked for them, and/or by helping mentees access these structures and mechanisms. Mentoring facilitates access to a CoP by providing opportunities for engagement. This involves sharing of experiences and knowledge, and promoting discussion. The mentor helps the mentee move from being a peripheral member of the CoP to becoming a main member, i.e., becoming active, learning with and from others within the CoP. CoPs develop social capital and knowledge management. My research suggests that the knowledge, skills and attitude developed by the mentors within this study may be transferable to other aspects in Pharmacy.
- Full Text:
- Date Issued: 2009
Adherence to antiretroviral therapy in children in Zimbabwe: a randomized control trial to validate a new self-reported adherence monitoring tool
- Authors: Mugore, Linnetie
- Date: 2014
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/54734 , vital:26607
- Description: Background: Among children taking antiretroviral therapy (ART), self-reports have been widely reported to over-estimate adherence levels. Pill count adherence levels are often lower than self-reported levels, with unannounced home pill count adherence being lower than facility based pill count adherence. There is often poor agreement between pill count adherence levels and those measured using other objective adherence measuring methods such as Medication Event Monitoring Systems (MEMS®), which is widely viewed as the gold standard for adherence measurement. Objectives: The aim of this study was to design and evaluate a new self-reported paediatric adherence monitoring tool, assess the feasibility of using pill count methods in monitoring adherence and identify challenges to reporting adherence among children on ART in rural and urban Zimbabwe. Methods A dual centre, superiority, parallel design RCT was conducted to evaluate the newly-developed visually- and verbally-cued „past 10 days‟ tool for the assessment of adherence in children on ART at two sites in Zimbabwe; Harare Central Children‟s Hospital in an urban setting, and Murambinda Mission Hospital, a rural site. Child-caregiver pairs presenting to one of these facilities for the child‟s review of ART and refill of the medication were recruited, signed informed consent obtained, and were randomised for self-reported adherence monitoring into either the experimental group („new 10-day tool‟) or the control group („PACTG-style‟ self-report tool). Data (demographic, socioeconomic, and reported adherence) were collected in individual interviews with child-caregiver pairs. Additional adherence monitoring methods used for both groups included the Morisky-8-Item Medication Adherence Scale (MMAS-8) and a facility based pill count. FGDs were held with groups of caregivers and groups of children ≥13 years of age to understand reasons for non-adherence as well as issues around reporting non-adherence. Superiority testing was conducted by comparing adherent proportions and their confidence intervals (95% CI). Further concurrent validity test was done using the Mann-Whitney U test to evaluate the relationship between the new tool and the MMAS-8 scores. Agreement between the child and caregiver reports of adherence was used as a test of reliability of the new tool using the kappa statistic. Socio-demographic, clinical and care-related factors associated with adherence were identified using reported adherence in both child and caregiver groups in a logistic regression model. Two pill count methods were assessed for feasibility using the proportions of children with complete data for calculating adherence levels, and their CI and a comparison of the two methods, a routinely-used method and one that incorporated the reported residual quantity (RRQ) of medication at last refill. Results : Analysis included 245 child-caregiver pairs, 123 in the experimental group and 122 in the control group. The median age for children was 9 years. In the experimental group, adherence by caregiver and child reports ranged from 94.3% - 98.4% and 78.4% - 96.1%, and those in the control group ranged from 89.2% - 97.5% and 71.2% - 98.1%, respectively. There was no significant difference between adherence levels in the two groups. Adherence levels measured by both the experimental and control tools were found to be associated with MMAS-8 adherence levels (p <0.05). Agreement between child- and caregiver-reported adherence was moderate though significant (kappa; 0.407, p <0.05). Only about half of the children had adequate data to compute pill counts. Proportions adherent at 95% cut-off were 39% by the „routine pill count‟ and 58% by the „Pill count RRQ‟. Being an orphan was associated with child reported-adherence whereas use of non-human reminders, having a maternal relative as a primary caregiver and knowledge of dose frequency, were all associated with caregiver-reported adherence. Major causes of non-adherence mentioned during the FGDs included interference of medication administration times with scheduling of routine socio-economic activities and lack of support from some non-biological caregivers. Reporting of non-adherence appeared to be hampered by perceptions of negative reactions by healthcare workers to these reports and by caregivers being unaware that the child missed some doses. Conclusions: The „new 10-day tool‟ was not shown to be superior to the „PACTG-style tool‟ in detecting non-adherence, however this new tool was found to be a valid and reliable adherence monitoring tool that included a moderately long recall period of 10 days, can be applied without the need for the respondent to remember names of individual medicines in the
- Full Text:
- Date Issued: 2014
- Authors: Mugore, Linnetie
- Date: 2014
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/54734 , vital:26607
- Description: Background: Among children taking antiretroviral therapy (ART), self-reports have been widely reported to over-estimate adherence levels. Pill count adherence levels are often lower than self-reported levels, with unannounced home pill count adherence being lower than facility based pill count adherence. There is often poor agreement between pill count adherence levels and those measured using other objective adherence measuring methods such as Medication Event Monitoring Systems (MEMS®), which is widely viewed as the gold standard for adherence measurement. Objectives: The aim of this study was to design and evaluate a new self-reported paediatric adherence monitoring tool, assess the feasibility of using pill count methods in monitoring adherence and identify challenges to reporting adherence among children on ART in rural and urban Zimbabwe. Methods A dual centre, superiority, parallel design RCT was conducted to evaluate the newly-developed visually- and verbally-cued „past 10 days‟ tool for the assessment of adherence in children on ART at two sites in Zimbabwe; Harare Central Children‟s Hospital in an urban setting, and Murambinda Mission Hospital, a rural site. Child-caregiver pairs presenting to one of these facilities for the child‟s review of ART and refill of the medication were recruited, signed informed consent obtained, and were randomised for self-reported adherence monitoring into either the experimental group („new 10-day tool‟) or the control group („PACTG-style‟ self-report tool). Data (demographic, socioeconomic, and reported adherence) were collected in individual interviews with child-caregiver pairs. Additional adherence monitoring methods used for both groups included the Morisky-8-Item Medication Adherence Scale (MMAS-8) and a facility based pill count. FGDs were held with groups of caregivers and groups of children ≥13 years of age to understand reasons for non-adherence as well as issues around reporting non-adherence. Superiority testing was conducted by comparing adherent proportions and their confidence intervals (95% CI). Further concurrent validity test was done using the Mann-Whitney U test to evaluate the relationship between the new tool and the MMAS-8 scores. Agreement between the child and caregiver reports of adherence was used as a test of reliability of the new tool using the kappa statistic. Socio-demographic, clinical and care-related factors associated with adherence were identified using reported adherence in both child and caregiver groups in a logistic regression model. Two pill count methods were assessed for feasibility using the proportions of children with complete data for calculating adherence levels, and their CI and a comparison of the two methods, a routinely-used method and one that incorporated the reported residual quantity (RRQ) of medication at last refill. Results : Analysis included 245 child-caregiver pairs, 123 in the experimental group and 122 in the control group. The median age for children was 9 years. In the experimental group, adherence by caregiver and child reports ranged from 94.3% - 98.4% and 78.4% - 96.1%, and those in the control group ranged from 89.2% - 97.5% and 71.2% - 98.1%, respectively. There was no significant difference between adherence levels in the two groups. Adherence levels measured by both the experimental and control tools were found to be associated with MMAS-8 adherence levels (p <0.05). Agreement between child- and caregiver-reported adherence was moderate though significant (kappa; 0.407, p <0.05). Only about half of the children had adequate data to compute pill counts. Proportions adherent at 95% cut-off were 39% by the „routine pill count‟ and 58% by the „Pill count RRQ‟. Being an orphan was associated with child reported-adherence whereas use of non-human reminders, having a maternal relative as a primary caregiver and knowledge of dose frequency, were all associated with caregiver-reported adherence. Major causes of non-adherence mentioned during the FGDs included interference of medication administration times with scheduling of routine socio-economic activities and lack of support from some non-biological caregivers. Reporting of non-adherence appeared to be hampered by perceptions of negative reactions by healthcare workers to these reports and by caregivers being unaware that the child missed some doses. Conclusions: The „new 10-day tool‟ was not shown to be superior to the „PACTG-style tool‟ in detecting non-adherence, however this new tool was found to be a valid and reliable adherence monitoring tool that included a moderately long recall period of 10 days, can be applied without the need for the respondent to remember names of individual medicines in the
- Full Text:
- Date Issued: 2014
African traditional medicine-antiretroviral interactions : effects of Sutherlandia frutescens on the pharmacokinetics of Atazanavir
- Authors: Müller, Adrienne Carmel
- Date: 2011 , 2011-03-28
- Subjects: Antiretroviral agents , Medicinal plants , Traditional medicine , AIDS (Disease) -- Treatment , HIV infections -- Drug therapy , Drug interactions , Pharmacokinetics
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3859 , http://hdl.handle.net/10962/d1013373
- Description: In response to the urgent call for investigations into antiretroviral (ARV)-African traditional medicine (ATM) interactions, this research was undertaken to ascertain whether chronic administration of the ATM, Sutherlandia frutescens (SF) may alter the bioavailability of the protease inhibitor (PI), atazanavir (ATV), which may impact on the safety or efficacy of the ARV. Prior to investigating a potential interaction between ATV and SF in vitro and in vivo, a high performance liquid chromatography method with ultraviolet detection (HPLC-UV) was developed and validated for the bioanalysis of ATV in human plasma and liver microsomes. An improved and efficient analytical method with minimal use of solvents and short run time was achieved in comparison to methods published in the literature. In addition, the method was selective, linear, accurate and precise for quantitative analysis of ATV in these studies. Molecular docking studies were conducted to compare the binding modes and affinities of ATV and two major SF constituents, Sutherlandioside B and Sutherlandin C, with the efflux transporter, P-glycoprotein (P-gp) and the CYP450 isoenzyme, CYP3A4 to determine the potential for these phytochemicals to competitively inhibit the binding of ATV to these two proteins, which are mediators of absorption and metabolism. These studies revealed that modulation of P-gp transport of ATV by Sutherlandioside B and Sutherlandin C was not likely to occur via competitive inhibition. The results further indicated that weak competitive inhibition of CYP3A4 may possibly occur in the presence of either of these two SF constituents. The Caco-2 cell line was used as an in vitro model of human intestinal absorption. Accumulation studies in these cells were conducted to ascertain whether extracts and constituents of SF have the ability to alter the absorption of ATV. The results showed that the aqueous extract of SF significantly reduced ATV accumulation, suggesting decreased ATV absorption, whilst a triterpenoid glycoside fraction isolated from SF exhibited an opposing effect. Analogous responses were elicited by the aqueous extract and a triterpenoid glycoside fraction in similar accumulation studies in P-gp overexpressing Madin–Darby Canine Kidney Strain II cells (MDCKII-MDR1), which signified that the effects of this extract and component on ATV transport in the Caco-2 cells were P-gp-mediated. The quantitative analysis of ATV in human liver microsomes after co-incubation with extracts and components of SF was conducted to determine the effects of SF on the metabolism of ATV. The aqueous and methanolic extracts of SF inhibited ATV metabolism, whilst the triterpenoid glycoside fraction had a converse effect. Analogous effects by the extracts were demonstrated in experiments conducted in CYP3A4-transfected microsomes, suggesting that the inhibition of ATV metabolism in the liver microsomes by these SF extracts was CYP3A4-mediated. A combination of Sutherlandiosides C and D also inhibited CYP3A4-mediated ATV metabolism, which was in contrast to the response elicited by the triterpenoid fraction in the liver microsomes, where other unidentified compounds, shown to be present therein, may have contributed to the activation of ATV metabolism. The in vitro studies revealed the potential for SF to alter the bioavailability of ATV, therefore a clinical study in which the effect of a multiple dose regimen of SF on the pharmacokinetics (PK) of a single dose of ATV was conducted in healthy male volunteers. The statistical analysis showed that the 90 % confidence intervals around the geometric mean ratios (ATV + SF/ATV alone) for both Cmax and AUC0-24 hours, fell well below the lower limit of the "no-effect" boundary of 0.8 – 1.25, implying that the bioavailability of ATV was significantly reduced in this cohort of subjects. It may thus be concluded that if the reduction in bioavailability observed in this clinical study is found to be clinically relevant, co-administration of SF commercial dosage forms and ATV in HIV/AIDS patients may potentially result in subtherapeutic ATV levels, which may in turn contribute to ATV resistance and/or treatment failure. This research has therefore highlighted the potential risk for toxicity or lack of efficacy of ARV regimens which may result when ATMs and PIs are used concurrently and that patients and health care practitioners alike should be aware of these perils.
- Full Text:
- Date Issued: 2011
- Authors: Müller, Adrienne Carmel
- Date: 2011 , 2011-03-28
- Subjects: Antiretroviral agents , Medicinal plants , Traditional medicine , AIDS (Disease) -- Treatment , HIV infections -- Drug therapy , Drug interactions , Pharmacokinetics
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3859 , http://hdl.handle.net/10962/d1013373
- Description: In response to the urgent call for investigations into antiretroviral (ARV)-African traditional medicine (ATM) interactions, this research was undertaken to ascertain whether chronic administration of the ATM, Sutherlandia frutescens (SF) may alter the bioavailability of the protease inhibitor (PI), atazanavir (ATV), which may impact on the safety or efficacy of the ARV. Prior to investigating a potential interaction between ATV and SF in vitro and in vivo, a high performance liquid chromatography method with ultraviolet detection (HPLC-UV) was developed and validated for the bioanalysis of ATV in human plasma and liver microsomes. An improved and efficient analytical method with minimal use of solvents and short run time was achieved in comparison to methods published in the literature. In addition, the method was selective, linear, accurate and precise for quantitative analysis of ATV in these studies. Molecular docking studies were conducted to compare the binding modes and affinities of ATV and two major SF constituents, Sutherlandioside B and Sutherlandin C, with the efflux transporter, P-glycoprotein (P-gp) and the CYP450 isoenzyme, CYP3A4 to determine the potential for these phytochemicals to competitively inhibit the binding of ATV to these two proteins, which are mediators of absorption and metabolism. These studies revealed that modulation of P-gp transport of ATV by Sutherlandioside B and Sutherlandin C was not likely to occur via competitive inhibition. The results further indicated that weak competitive inhibition of CYP3A4 may possibly occur in the presence of either of these two SF constituents. The Caco-2 cell line was used as an in vitro model of human intestinal absorption. Accumulation studies in these cells were conducted to ascertain whether extracts and constituents of SF have the ability to alter the absorption of ATV. The results showed that the aqueous extract of SF significantly reduced ATV accumulation, suggesting decreased ATV absorption, whilst a triterpenoid glycoside fraction isolated from SF exhibited an opposing effect. Analogous responses were elicited by the aqueous extract and a triterpenoid glycoside fraction in similar accumulation studies in P-gp overexpressing Madin–Darby Canine Kidney Strain II cells (MDCKII-MDR1), which signified that the effects of this extract and component on ATV transport in the Caco-2 cells were P-gp-mediated. The quantitative analysis of ATV in human liver microsomes after co-incubation with extracts and components of SF was conducted to determine the effects of SF on the metabolism of ATV. The aqueous and methanolic extracts of SF inhibited ATV metabolism, whilst the triterpenoid glycoside fraction had a converse effect. Analogous effects by the extracts were demonstrated in experiments conducted in CYP3A4-transfected microsomes, suggesting that the inhibition of ATV metabolism in the liver microsomes by these SF extracts was CYP3A4-mediated. A combination of Sutherlandiosides C and D also inhibited CYP3A4-mediated ATV metabolism, which was in contrast to the response elicited by the triterpenoid fraction in the liver microsomes, where other unidentified compounds, shown to be present therein, may have contributed to the activation of ATV metabolism. The in vitro studies revealed the potential for SF to alter the bioavailability of ATV, therefore a clinical study in which the effect of a multiple dose regimen of SF on the pharmacokinetics (PK) of a single dose of ATV was conducted in healthy male volunteers. The statistical analysis showed that the 90 % confidence intervals around the geometric mean ratios (ATV + SF/ATV alone) for both Cmax and AUC0-24 hours, fell well below the lower limit of the "no-effect" boundary of 0.8 – 1.25, implying that the bioavailability of ATV was significantly reduced in this cohort of subjects. It may thus be concluded that if the reduction in bioavailability observed in this clinical study is found to be clinically relevant, co-administration of SF commercial dosage forms and ATV in HIV/AIDS patients may potentially result in subtherapeutic ATV levels, which may in turn contribute to ATV resistance and/or treatment failure. This research has therefore highlighted the potential risk for toxicity or lack of efficacy of ARV regimens which may result when ATMs and PIs are used concurrently and that patients and health care practitioners alike should be aware of these perils.
- Full Text:
- Date Issued: 2011
An investigation into the antidepressant activity of hypericum perforatum
- Authors: Stephens, Linda Lee
- Date: 2005
- Subjects: Hypericum perforatum -- Physiological effect Hypericum perforatum -- Therapeutic use Antidepressants
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3793 , http://hdl.handle.net/10962/d1003271
- Description: Hypericum perforatum is a herbal medicine that has been used for centuries for the treatment of depression. Many studies have been conducted in the Northern hemisphere on the efficacy of the HP extracts produced there. These studies include clinical trials and pharmacological investigations using a standardised HP extract or a fraction of the HP extract containing certain compounds, such as hypericin, pseudohypericin, hyperforin and several of the flavonoids thought to be responsible for the antidepressant activity. The mechanism of action of HP and its constituents is still not completely clear and it is speculated that the antidepressant activity is the result of several of the compounds acting synergistically. HP is indigenous to and also cultivated in the Western Cape of South Africa. Extracts from these plants are sold in the local health shops and there are no previous studies evaluating the efficacy of these products. The aim of this thesis is to investigate the antidepressant activity of one of these products and two of its constituents, quercetin and caffeic acid, to gain further insight into their mode of antidepressant action and to compare these results with similar studies which used a standardised extract produced in the northern hemisphere. The first study investigated the effect of HP, quercetin and caffeic acid on pineal metabolism. Changes in the synthesis of melatonin produced by the pineal gland have been implicated in depression. The results showed an increase in the level of melatonin produced in the animals treated with quercetin, which suggests that this compound may mediate antidepressant activity through such a mechanism. There are no previous reports on the in vivo effects of HP or any of its constituents on pineal metabolism. The second study investigated the effect of HP, quercetin and caffeic acid on the activity of the liver enzyme, tryptophan-2,3-dioxygenase (TDO). Inhibition of this enzyme has been shown to increase plasma levels of tryptophan, a precursor of serotonin and thereby result in increased serotonin levels in the brain. Low levels of serotonin in the brain have been implicated in depression. This study revealed significant inhibition of TDO by caffeic acid and this suggests that this constituent of HP could be contributing to its antidepressant activity through such a mechanism. There are no previous reports investigating the in vivo effect of HP or any of its constituents on TDO activity. Modulation of the levels of indoleamines, serotonin (5-HT) and dopamine (DA) as well as the metabolites, 3,4 dihydroxyphenyl acetic acid (DOPAC), 5-hydroxyindole acetic acid (5-HIAA) and homovallinic acid (HVA) in the brain have been implicated in the neuropharmacology of depression. Different studies using enzyme-linked immunosorbant assay (ELISA), high performance liquid chromatography with electrochemical detection (HPLC-ECD) and liquid chromatography-mass spectrometry (LC-MS) were used to determine changes in the levels of these indoleamines brought about after treatment with HP caffeic acid and quercetin. The results of the ELISA study showed significant increases in 5-HT levels in the brains of the animals treated with caffeic acid and quercetin. The results of the HPLC-ECD studies also revealed significant increases in 5-HT levels and a decrease in the turnover of 5-HT in the animals treated with quercetin. A significant increase in DA levels in the animals treated with quercetin was shown in both the HPLC-ECD and LC-MS studies. There was also an increase in DA turnover in the animals treated with HP shown in the HPLC-ECD and LC-MS studies. These results suggest that HP and its constituents, quercetin and caffeic acid mediate their antidepressant effects through serotonergic and dopaminergic neurotransmission. Adaptive changes in the density of b-adrenergic (b-AR), 5-HT2 and N-methyl-D-aspartate (NMDA) receptors have been implicated in depression. Several studies, investigating the effect of treatment with HP and quercetin on these different receptor densities, were undertaken using radioactive binding assays. Treatment with HP resulted in significant down regulation of b-AR and NMDA receptor densities and up-regulation of 5HT2 receptors. The effects on the b-AR and 5-HT2 receptors are similar to the results reported using HP in the Northern hemisphere, but the effect on the NMDA receptors is novel providing insight into the mode of action of HP. Apoptosis of neuronal cells has been implicated in neuro-degenerative and depressive disorders. Detection of apoptosis, using fluorescent microscopy observed through the labelling of DNA strand breaks, showed a decrease in the amount of apoptosis in the animals treated with HP and quercetin. This adds further support for the use of HP as an antidepressant and these results are similar to results reported from the Northern hemisphere. The results of all these studies suggest that the quality of the locally produced tincture is similar in efficacy to that of the standardised product of the Northern hemisphere.
- Full Text:
- Date Issued: 2005
- Authors: Stephens, Linda Lee
- Date: 2005
- Subjects: Hypericum perforatum -- Physiological effect Hypericum perforatum -- Therapeutic use Antidepressants
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3793 , http://hdl.handle.net/10962/d1003271
- Description: Hypericum perforatum is a herbal medicine that has been used for centuries for the treatment of depression. Many studies have been conducted in the Northern hemisphere on the efficacy of the HP extracts produced there. These studies include clinical trials and pharmacological investigations using a standardised HP extract or a fraction of the HP extract containing certain compounds, such as hypericin, pseudohypericin, hyperforin and several of the flavonoids thought to be responsible for the antidepressant activity. The mechanism of action of HP and its constituents is still not completely clear and it is speculated that the antidepressant activity is the result of several of the compounds acting synergistically. HP is indigenous to and also cultivated in the Western Cape of South Africa. Extracts from these plants are sold in the local health shops and there are no previous studies evaluating the efficacy of these products. The aim of this thesis is to investigate the antidepressant activity of one of these products and two of its constituents, quercetin and caffeic acid, to gain further insight into their mode of antidepressant action and to compare these results with similar studies which used a standardised extract produced in the northern hemisphere. The first study investigated the effect of HP, quercetin and caffeic acid on pineal metabolism. Changes in the synthesis of melatonin produced by the pineal gland have been implicated in depression. The results showed an increase in the level of melatonin produced in the animals treated with quercetin, which suggests that this compound may mediate antidepressant activity through such a mechanism. There are no previous reports on the in vivo effects of HP or any of its constituents on pineal metabolism. The second study investigated the effect of HP, quercetin and caffeic acid on the activity of the liver enzyme, tryptophan-2,3-dioxygenase (TDO). Inhibition of this enzyme has been shown to increase plasma levels of tryptophan, a precursor of serotonin and thereby result in increased serotonin levels in the brain. Low levels of serotonin in the brain have been implicated in depression. This study revealed significant inhibition of TDO by caffeic acid and this suggests that this constituent of HP could be contributing to its antidepressant activity through such a mechanism. There are no previous reports investigating the in vivo effect of HP or any of its constituents on TDO activity. Modulation of the levels of indoleamines, serotonin (5-HT) and dopamine (DA) as well as the metabolites, 3,4 dihydroxyphenyl acetic acid (DOPAC), 5-hydroxyindole acetic acid (5-HIAA) and homovallinic acid (HVA) in the brain have been implicated in the neuropharmacology of depression. Different studies using enzyme-linked immunosorbant assay (ELISA), high performance liquid chromatography with electrochemical detection (HPLC-ECD) and liquid chromatography-mass spectrometry (LC-MS) were used to determine changes in the levels of these indoleamines brought about after treatment with HP caffeic acid and quercetin. The results of the ELISA study showed significant increases in 5-HT levels in the brains of the animals treated with caffeic acid and quercetin. The results of the HPLC-ECD studies also revealed significant increases in 5-HT levels and a decrease in the turnover of 5-HT in the animals treated with quercetin. A significant increase in DA levels in the animals treated with quercetin was shown in both the HPLC-ECD and LC-MS studies. There was also an increase in DA turnover in the animals treated with HP shown in the HPLC-ECD and LC-MS studies. These results suggest that HP and its constituents, quercetin and caffeic acid mediate their antidepressant effects through serotonergic and dopaminergic neurotransmission. Adaptive changes in the density of b-adrenergic (b-AR), 5-HT2 and N-methyl-D-aspartate (NMDA) receptors have been implicated in depression. Several studies, investigating the effect of treatment with HP and quercetin on these different receptor densities, were undertaken using radioactive binding assays. Treatment with HP resulted in significant down regulation of b-AR and NMDA receptor densities and up-regulation of 5HT2 receptors. The effects on the b-AR and 5-HT2 receptors are similar to the results reported using HP in the Northern hemisphere, but the effect on the NMDA receptors is novel providing insight into the mode of action of HP. Apoptosis of neuronal cells has been implicated in neuro-degenerative and depressive disorders. Detection of apoptosis, using fluorescent microscopy observed through the labelling of DNA strand breaks, showed a decrease in the amount of apoptosis in the animals treated with HP and quercetin. This adds further support for the use of HP as an antidepressant and these results are similar to results reported from the Northern hemisphere. The results of all these studies suggest that the quality of the locally produced tincture is similar in efficacy to that of the standardised product of the Northern hemisphere.
- Full Text:
- Date Issued: 2005
An investigation into the feasibility of incorporating didanosine into innovative solid lipid nanocarriers
- Authors: Wa Kasongo, Kasongo
- Date: 2010
- Subjects: Antiretroviral agents HIV infections -- Drug testing Didanosine Nanoparticles Drug delivery systems Nanostructured materials Lipids -- Therapeutic use
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3800 , http://hdl.handle.net/10962/d1003278
- Description: The research undertaken in these studies aimed to investigate the feasibility of developing and manufacturing innovative solid lipid carriers, such as solid lipid nanoparticles (SLN) and/or nanostructured lipid carriers (NLC) using a hot high pressure homogenization method, for didanosine(DDI). In addition, studies using in vitro differential protein adsorption were undertaken to establish whether the SLN and/or NLC have the potential to deliver DDI to the central nervous system (CNS). Prior to initiating pre-formulation, formulation development and optimization studies of DDI-Ioaded SLN and/or NLC, it was necessary to develop and validate an analytical method for the in vitro quantitation and analysis of DDI. An accurate, precise and sensitive RP-HPLC method with UV detection set at 248 nm was developed, optimized and validated for the quantitative in vitro analysis of DDI in formulations. Pre-formulation studies were designed to evaluate the thermal stability of DDI and to select and characterize lipid excipients that may be used for the manufacture of the nanocarriers. It was established that DDI is thermostable at temperatures not exceeding 163°C and therefore a hot high pressure homogenization technique could be used to manufacture DDI-loaded SLN and/or NLC. Lipid screening studies revealed that DDI is poorly soluble in both solid and liquid lipids. A combination of Precirol® ATO 5 and Transcutol® HP was found to have the best solubilizing-potential for DDI of all lipids investigated. The inclusion of Transcutol® HP into Precirol® ATO 5 changed the polymorphic form of the solid lipid from the stable 13-modification to a material that exhibited the co-existence between α- and β-polymorphic forms. The relatively high solubility of DDI in Transcutol® HP compared to Precirol® ATO 5 was an indication that a solid lipid matrix prepared from a binary mixture of Precirol® ATO 5 and Transcutol® HP was likely to have a higher loading capacity and encapsulation efficiency for DDI than a matrix consisting of Precirol® ATO 5 alone. Furthermore, the potential for the solid lipid matrix to exist in the α- and/or β-modifications when Transcutol® HP was added to Precirol® ATO 5 suggested that expulsion of DDI from a solid lipid matrix during prolonged storage periods was likely to be minimal. Therefore it was considered logical to investigate the feasibility of incorporating DDI into NLC and not in SLN. However, due to the limited solubility of DDI in lipids, formulation development of DDI-loaded NLC commenced using small quantities of DDI. Formulation development and optimization studies of DDI-loaded NLC were initially aimed at selecting a surfactant system that was capable of stabilizing NLC in an aqueous environment. Solutol® HS alone or a ternary mixture consisting of Solutol® HS, Tween® 80 and Lutrol® F68 was found to stabilize the nanoparticles in terms of particle size and the polydispersity index. The use of the ternary mixture as the surfactant system was preferred to using Solutol® HS alone as Lutrol® F68 and especially Tween® 80 have been successfully used to target the delivery of API to the brain. Aqueous DDI-free and DDI-Ioaded NLC containing increasing amounts of DDI were manufactured using hot high pressure homogenization at 800 bar for three cycles. The NLC formulations were characterized in terms of particle size, polydispersity index, zeta potential, and polymorphism, degree of crystallinity, encapsulation efficiency (EE), shape and surface morphology. The mean particle size for all formulations was below 250 nm with narrow polydispersity indices, indicating that narrow particle size distribution had been achieved. The d99% values for all formulations tested, were generated using laser diffractometry, and were below 400 nm, with span values ranging from 0.84 - 1.19 also suggesting that a narrow particle size distribution had been achieved. The zeta potential values measured in double distilled water with the conductivity adjusted to 50 μS/cm ranged from -18.4 to -11.4 mV. In addition, all the formulations showed a decrease in the degree of crystallinity as compared to the bulk lipid material and WAXS shows that the formulations existed in a single β-modification form. Furthermore DDI that had been incorporated into the NLC appeared to be molecularly dispersed in the lipid matrices. These parameters remained unaffected for most formulations following storage for two months at 25°C. In addition these formulations contained a mixture of spherical and non-spherical particles irrespective of the amount of DDI that was added during the manufacture of the formulations. These studies showed that it was feasible to develop and incorporate small amounts of DDI into NLC. However in order to use these delivery systems for oral administration of DDI to paediatric patients, strategies to improve the amount of DDI that could be loaded into the particles and to achieve high encapsulation efficiencies had to be developed. The limited solubility of DDI in lipid media was identified as a major factor that affected the loading capacity and encapsulation efficiency of DDI in the NLC. Therefore, a novel strategy aimed at increasing the saturation solubility of DDI in the lipid by attempting to increase the dissolution velocity of the drug in the lipid using a particle size reduction approach, was designed and investigated. DDI was dispersed in Transcutol® HP and the particle size of DDI in the liquid lipid medium was reduced gradually using hot high pressure homogenization and the product obtained from these studies was used to manufacture DDI-loaded NLC using a cold high pressure homogenization procedure. Although the encapsulation efficiency and drug loading following use of this approach was relatively high, the particles were large and showed a tendency to grow in size leading to the formation of microparticles after storage for two months at 25°C. In addition, the degree of crystallinity of the nanoparticles increased rapidly over the same storage period which led to expulsion of DDI nanoparticles for the NLC, despite the DDI loading in NLC being unaffected. It was clearly evident that this new approach of manufacturing solid lipid nanocarriers could be used as a platform not only for enhancing the loading capacity of DDI in solid lipid nanocarriers but also for other hydrophilic drugs. Differential protein adsorption patterns of DDI-loaded NLC were generated in vitro using two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) in order to establish the potential for these systems to deliver DDI to the CNS. NLC formulations containing small amounts of DDI were used as these formulations showed a better stability profile than the formulation with a higher encapsulation efficiency and drug loading capacity. Furthermore, the encapsulation efficiency and drug loading of DDI were considered sufficient for use in 2-D PAGE studies. Data obtained from 2-D PAGE analysis reveal that DDI-loaded NLC preferentially adsorb proteins in vitro that are responsible for specific brain targeting in vivo. More importantly, these studies reveal that in addition to Tween® 80 that has already been shown to have the potential to target CDDS to the brain, Solutol® HS 15 has the potential to achieve a similar objective. Consequently, DDI-loaded NLC have the potential to deliver DDI to the brain and these results may be used as a platform for conducting in vivo studies to establish whether DDI can cross the blood brain barrier and enter the CNS when administered in NLC which may in turn lead to a major breakthrough in the management of HIV/AIDS and Aids Dementia Complex (ADC).
- Full Text:
- Date Issued: 2010
- Authors: Wa Kasongo, Kasongo
- Date: 2010
- Subjects: Antiretroviral agents HIV infections -- Drug testing Didanosine Nanoparticles Drug delivery systems Nanostructured materials Lipids -- Therapeutic use
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3800 , http://hdl.handle.net/10962/d1003278
- Description: The research undertaken in these studies aimed to investigate the feasibility of developing and manufacturing innovative solid lipid carriers, such as solid lipid nanoparticles (SLN) and/or nanostructured lipid carriers (NLC) using a hot high pressure homogenization method, for didanosine(DDI). In addition, studies using in vitro differential protein adsorption were undertaken to establish whether the SLN and/or NLC have the potential to deliver DDI to the central nervous system (CNS). Prior to initiating pre-formulation, formulation development and optimization studies of DDI-Ioaded SLN and/or NLC, it was necessary to develop and validate an analytical method for the in vitro quantitation and analysis of DDI. An accurate, precise and sensitive RP-HPLC method with UV detection set at 248 nm was developed, optimized and validated for the quantitative in vitro analysis of DDI in formulations. Pre-formulation studies were designed to evaluate the thermal stability of DDI and to select and characterize lipid excipients that may be used for the manufacture of the nanocarriers. It was established that DDI is thermostable at temperatures not exceeding 163°C and therefore a hot high pressure homogenization technique could be used to manufacture DDI-loaded SLN and/or NLC. Lipid screening studies revealed that DDI is poorly soluble in both solid and liquid lipids. A combination of Precirol® ATO 5 and Transcutol® HP was found to have the best solubilizing-potential for DDI of all lipids investigated. The inclusion of Transcutol® HP into Precirol® ATO 5 changed the polymorphic form of the solid lipid from the stable 13-modification to a material that exhibited the co-existence between α- and β-polymorphic forms. The relatively high solubility of DDI in Transcutol® HP compared to Precirol® ATO 5 was an indication that a solid lipid matrix prepared from a binary mixture of Precirol® ATO 5 and Transcutol® HP was likely to have a higher loading capacity and encapsulation efficiency for DDI than a matrix consisting of Precirol® ATO 5 alone. Furthermore, the potential for the solid lipid matrix to exist in the α- and/or β-modifications when Transcutol® HP was added to Precirol® ATO 5 suggested that expulsion of DDI from a solid lipid matrix during prolonged storage periods was likely to be minimal. Therefore it was considered logical to investigate the feasibility of incorporating DDI into NLC and not in SLN. However, due to the limited solubility of DDI in lipids, formulation development of DDI-loaded NLC commenced using small quantities of DDI. Formulation development and optimization studies of DDI-loaded NLC were initially aimed at selecting a surfactant system that was capable of stabilizing NLC in an aqueous environment. Solutol® HS alone or a ternary mixture consisting of Solutol® HS, Tween® 80 and Lutrol® F68 was found to stabilize the nanoparticles in terms of particle size and the polydispersity index. The use of the ternary mixture as the surfactant system was preferred to using Solutol® HS alone as Lutrol® F68 and especially Tween® 80 have been successfully used to target the delivery of API to the brain. Aqueous DDI-free and DDI-Ioaded NLC containing increasing amounts of DDI were manufactured using hot high pressure homogenization at 800 bar for three cycles. The NLC formulations were characterized in terms of particle size, polydispersity index, zeta potential, and polymorphism, degree of crystallinity, encapsulation efficiency (EE), shape and surface morphology. The mean particle size for all formulations was below 250 nm with narrow polydispersity indices, indicating that narrow particle size distribution had been achieved. The d99% values for all formulations tested, were generated using laser diffractometry, and were below 400 nm, with span values ranging from 0.84 - 1.19 also suggesting that a narrow particle size distribution had been achieved. The zeta potential values measured in double distilled water with the conductivity adjusted to 50 μS/cm ranged from -18.4 to -11.4 mV. In addition, all the formulations showed a decrease in the degree of crystallinity as compared to the bulk lipid material and WAXS shows that the formulations existed in a single β-modification form. Furthermore DDI that had been incorporated into the NLC appeared to be molecularly dispersed in the lipid matrices. These parameters remained unaffected for most formulations following storage for two months at 25°C. In addition these formulations contained a mixture of spherical and non-spherical particles irrespective of the amount of DDI that was added during the manufacture of the formulations. These studies showed that it was feasible to develop and incorporate small amounts of DDI into NLC. However in order to use these delivery systems for oral administration of DDI to paediatric patients, strategies to improve the amount of DDI that could be loaded into the particles and to achieve high encapsulation efficiencies had to be developed. The limited solubility of DDI in lipid media was identified as a major factor that affected the loading capacity and encapsulation efficiency of DDI in the NLC. Therefore, a novel strategy aimed at increasing the saturation solubility of DDI in the lipid by attempting to increase the dissolution velocity of the drug in the lipid using a particle size reduction approach, was designed and investigated. DDI was dispersed in Transcutol® HP and the particle size of DDI in the liquid lipid medium was reduced gradually using hot high pressure homogenization and the product obtained from these studies was used to manufacture DDI-loaded NLC using a cold high pressure homogenization procedure. Although the encapsulation efficiency and drug loading following use of this approach was relatively high, the particles were large and showed a tendency to grow in size leading to the formation of microparticles after storage for two months at 25°C. In addition, the degree of crystallinity of the nanoparticles increased rapidly over the same storage period which led to expulsion of DDI nanoparticles for the NLC, despite the DDI loading in NLC being unaffected. It was clearly evident that this new approach of manufacturing solid lipid nanocarriers could be used as a platform not only for enhancing the loading capacity of DDI in solid lipid nanocarriers but also for other hydrophilic drugs. Differential protein adsorption patterns of DDI-loaded NLC were generated in vitro using two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) in order to establish the potential for these systems to deliver DDI to the CNS. NLC formulations containing small amounts of DDI were used as these formulations showed a better stability profile than the formulation with a higher encapsulation efficiency and drug loading capacity. Furthermore, the encapsulation efficiency and drug loading of DDI were considered sufficient for use in 2-D PAGE studies. Data obtained from 2-D PAGE analysis reveal that DDI-loaded NLC preferentially adsorb proteins in vitro that are responsible for specific brain targeting in vivo. More importantly, these studies reveal that in addition to Tween® 80 that has already been shown to have the potential to target CDDS to the brain, Solutol® HS 15 has the potential to achieve a similar objective. Consequently, DDI-loaded NLC have the potential to deliver DDI to the brain and these results may be used as a platform for conducting in vivo studies to establish whether DDI can cross the blood brain barrier and enter the CNS when administered in NLC which may in turn lead to a major breakthrough in the management of HIV/AIDS and Aids Dementia Complex (ADC).
- Full Text:
- Date Issued: 2010
An investigation into the neuroprotective properties of acetylsalicylic acid and acetaminophen
- Authors: Maharaj, Himant
- Date: 2005
- Subjects: Aspirin Acetaminophen Analgesics Alzheimer's disease -- Treatment Parkinson's disease
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3769 , http://hdl.handle.net/10962/d1003247
- Description: The potent analgesic property of acetylsalicylic acid and acetaminophen makes these the most commonly used analgesics in the world. Easy accessibility and cost effectiveness of these agents are attractive to patients seeking pain relief. However, the abuse of nonnarcotic analgesics such as acetaminophen and acetylsalicylic acid by alcoholics and patients seeking to relieve dysphoric moods is well documented. These agents therefore impact on the brain neurotransmitter levels and therefore all processes involved in the synthesis and metabolism of neurotransmitters may be affected. The use of non-narcotic analgesics has been reported to reduce the incidence of neurodegenerative disorders such as Alzheimer’s disease (AD) and Parkinson’s disease (PD). The mode of action by which acetylsalicylic acid and acetaminophen elicit neuroprotection is however unclear as many mechanisms of action have been inconclusively postulated. The first part of this study aims to elucidate the various mechanisms by which acetylsalicylic acid and acetaminophen affect the enzymes responsible for the catabolism of tryptophan, which is a precursor for the mood elevating neurotransmitter serotonin, as well as to investigate whether these agents alter the interplay between serotonin and pineal indole metabolism. The second part of this study focuses on the neuroprotective properties of acetylsalicylic acid and acetaminophen utilizing the neurotoxic metabolite of the kynurenine pathway, quinolinic acid and the potent Parkinsonian neurotoxin, 1-methyl-4-phenylpyridinium (MPP+). The ability of acetylsalicylic acid and acetaminophen to alter TRP metabolism was determined by investigating the effects of these agents on the primary enzymes of the kynurenine pathway i.e. tryptophan 2, 3-dioxygenase and indoleamine 2,3-dioxygenase as well as to investigate whether these agents would have any effects on 3-hydroxyanthranilic acid oxygenase. 3-Hydroxyanthranilic acid oxygenase is the enzyme responsible for the synthesis of quinolinic acid. Acetylsalicylic acid and acetaminophen alter tryptophan metabolism by inhibiting tryptophan 2, 3-dioxygenase and indoleamine 2,3-dioxygenase thus increasing the availability of tryptophan for the production of serotonin. Acetylsalicylic acid and acetaminophen also inhibit 3-hydroxyanthranilic acid oxygenase thus implying that these agents could reduce quinolinic acid production. Acetaminophen administration in rats induces a rise in serotonin and norepinephrine in the forebrain. Acetylsalicylic acid curtails the acetaminophen-induced rise in brain norepinephrine levels as well as enhances serotonin metabolism, indicating that analgesic preparations containing both agents would be advantageous, as this would prevent acetaminophen-induced mood elevation. The results from the pineal indole metabolism study show that acetylsalicylic acid enhances pineal metabolism of serotonin whereas acetaminophen induces an increase in melatonin levels in the pineal gland. Neuronal damage due to oxidative stress has been implicated in several neurodegenerative disorders such as AD and PD. The second part of the study aims to elucidate and characterize the mechanism by which acetylsalicylic acid and acetaminophen afford neuroprotection. The hippocampus is an important region of the brain responsible for memory. Agents such as quinolinic acid that are known to induce stress in this area have detrimental effects and could lead to various types of dementia. The striatum is also a vulnerable region to oxidative stress and hence (MPP+), which is toxic for this particular region of the brain, was also used as a neurotoxin. The results show that ASA and acetaminophen alone and in combination, are potent superoxide anion scavengers. In addition, the results imply that these agents offer protection against oxidative stress and lipid peroxidation induced by several neurotoxins in rat brain particularly, the hippocampus and striatum. Histological studies, using Nissl staining and Acid fuchsin, show that acetylsalicylic acid and acetaminophen are able to protect hippocampal neurons against quinolinic acidinduced necrotic cell death. Immunohistochemical investigations show that QA induces apoptotic cell death in the hippocampus, which is inhibited by ASA and acetaminophen. In addition, ASA and acetaminophen inhibited MPP+ induced apoptotic cell death in the rat striatum. The study also sought to elucidate possible mechanisms by which ASA and acetaminophen exert neuroprotective effects in the presence of MPP+ as these agents are shown to prevent the MPP+-induced reduction in dopamine levels. The results show that acetylsalicylic acid and acetaminophen inhibit the action of this neurotoxin on the mitochondrial electron transport chain, a common source of free radicals in the cell. In addition, these agents were shown to block the neurotoxic effects of MPP+ on the enzymatic defence system of the brain i.e. superoxide dismutase, glutathione peroxidase and catalase. The reduction in glutathione levels induced by MPP+ is significantly inhibited by acetylsalicylic acid and acetaminophen. The results imply that these agents are capable of not only scavenging free radicals but also enhance the cell defence mechanism against toxicity in the presence of MPP+. These agents also block the MPP+-induced inhibition of dopamine uptake into the cell. This would therefore reduce auto-oxidation of dopamine thus implying another mechanism by which these agents exert a neuroprotective role in MPP+-induced neurotoxicity. The discovery of neuroprotective properties of acetylsalicylic acid and acetaminophen is important considering the high usage of these agents and the increased incidence in neurological disorders. The findings of this thesis point to the need for clinical studies to be conducted as the results show acetylsalicylic acid and acetaminophen to have a definite role to play as antioxidants. This study therefore provides novel information regarding the neuroprotective effects of these agents and favours the use of these agents in the treatment of neurodegenerative disorders, such as AD and PD, in which oxidative stress is implicated.
- Full Text:
- Date Issued: 2005
- Authors: Maharaj, Himant
- Date: 2005
- Subjects: Aspirin Acetaminophen Analgesics Alzheimer's disease -- Treatment Parkinson's disease
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3769 , http://hdl.handle.net/10962/d1003247
- Description: The potent analgesic property of acetylsalicylic acid and acetaminophen makes these the most commonly used analgesics in the world. Easy accessibility and cost effectiveness of these agents are attractive to patients seeking pain relief. However, the abuse of nonnarcotic analgesics such as acetaminophen and acetylsalicylic acid by alcoholics and patients seeking to relieve dysphoric moods is well documented. These agents therefore impact on the brain neurotransmitter levels and therefore all processes involved in the synthesis and metabolism of neurotransmitters may be affected. The use of non-narcotic analgesics has been reported to reduce the incidence of neurodegenerative disorders such as Alzheimer’s disease (AD) and Parkinson’s disease (PD). The mode of action by which acetylsalicylic acid and acetaminophen elicit neuroprotection is however unclear as many mechanisms of action have been inconclusively postulated. The first part of this study aims to elucidate the various mechanisms by which acetylsalicylic acid and acetaminophen affect the enzymes responsible for the catabolism of tryptophan, which is a precursor for the mood elevating neurotransmitter serotonin, as well as to investigate whether these agents alter the interplay between serotonin and pineal indole metabolism. The second part of this study focuses on the neuroprotective properties of acetylsalicylic acid and acetaminophen utilizing the neurotoxic metabolite of the kynurenine pathway, quinolinic acid and the potent Parkinsonian neurotoxin, 1-methyl-4-phenylpyridinium (MPP+). The ability of acetylsalicylic acid and acetaminophen to alter TRP metabolism was determined by investigating the effects of these agents on the primary enzymes of the kynurenine pathway i.e. tryptophan 2, 3-dioxygenase and indoleamine 2,3-dioxygenase as well as to investigate whether these agents would have any effects on 3-hydroxyanthranilic acid oxygenase. 3-Hydroxyanthranilic acid oxygenase is the enzyme responsible for the synthesis of quinolinic acid. Acetylsalicylic acid and acetaminophen alter tryptophan metabolism by inhibiting tryptophan 2, 3-dioxygenase and indoleamine 2,3-dioxygenase thus increasing the availability of tryptophan for the production of serotonin. Acetylsalicylic acid and acetaminophen also inhibit 3-hydroxyanthranilic acid oxygenase thus implying that these agents could reduce quinolinic acid production. Acetaminophen administration in rats induces a rise in serotonin and norepinephrine in the forebrain. Acetylsalicylic acid curtails the acetaminophen-induced rise in brain norepinephrine levels as well as enhances serotonin metabolism, indicating that analgesic preparations containing both agents would be advantageous, as this would prevent acetaminophen-induced mood elevation. The results from the pineal indole metabolism study show that acetylsalicylic acid enhances pineal metabolism of serotonin whereas acetaminophen induces an increase in melatonin levels in the pineal gland. Neuronal damage due to oxidative stress has been implicated in several neurodegenerative disorders such as AD and PD. The second part of the study aims to elucidate and characterize the mechanism by which acetylsalicylic acid and acetaminophen afford neuroprotection. The hippocampus is an important region of the brain responsible for memory. Agents such as quinolinic acid that are known to induce stress in this area have detrimental effects and could lead to various types of dementia. The striatum is also a vulnerable region to oxidative stress and hence (MPP+), which is toxic for this particular region of the brain, was also used as a neurotoxin. The results show that ASA and acetaminophen alone and in combination, are potent superoxide anion scavengers. In addition, the results imply that these agents offer protection against oxidative stress and lipid peroxidation induced by several neurotoxins in rat brain particularly, the hippocampus and striatum. Histological studies, using Nissl staining and Acid fuchsin, show that acetylsalicylic acid and acetaminophen are able to protect hippocampal neurons against quinolinic acidinduced necrotic cell death. Immunohistochemical investigations show that QA induces apoptotic cell death in the hippocampus, which is inhibited by ASA and acetaminophen. In addition, ASA and acetaminophen inhibited MPP+ induced apoptotic cell death in the rat striatum. The study also sought to elucidate possible mechanisms by which ASA and acetaminophen exert neuroprotective effects in the presence of MPP+ as these agents are shown to prevent the MPP+-induced reduction in dopamine levels. The results show that acetylsalicylic acid and acetaminophen inhibit the action of this neurotoxin on the mitochondrial electron transport chain, a common source of free radicals in the cell. In addition, these agents were shown to block the neurotoxic effects of MPP+ on the enzymatic defence system of the brain i.e. superoxide dismutase, glutathione peroxidase and catalase. The reduction in glutathione levels induced by MPP+ is significantly inhibited by acetylsalicylic acid and acetaminophen. The results imply that these agents are capable of not only scavenging free radicals but also enhance the cell defence mechanism against toxicity in the presence of MPP+. These agents also block the MPP+-induced inhibition of dopamine uptake into the cell. This would therefore reduce auto-oxidation of dopamine thus implying another mechanism by which these agents exert a neuroprotective role in MPP+-induced neurotoxicity. The discovery of neuroprotective properties of acetylsalicylic acid and acetaminophen is important considering the high usage of these agents and the increased incidence in neurological disorders. The findings of this thesis point to the need for clinical studies to be conducted as the results show acetylsalicylic acid and acetaminophen to have a definite role to play as antioxidants. This study therefore provides novel information regarding the neuroprotective effects of these agents and favours the use of these agents in the treatment of neurodegenerative disorders, such as AD and PD, in which oxidative stress is implicated.
- Full Text:
- Date Issued: 2005
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
An investigation into the physico-chemical and neuroprotective properties of melatonin and 6-hydroxymelatonin
- Authors: Maharaj, Deepa Sukhdev
- Date: 2003
- Subjects: Melatonin Nervous system -- Degeneration -- Treatment
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3768 , http://hdl.handle.net/10962/d1003246
- Description: Until the beginning of this decade the antioxidant, melatonin, had been considered as little more than a tranquilizing hormone, responsible for regulating certain circadian and circannual rhythms. However, it is the discovery of melatonin as a free radical scavenger that has generated the most interest in recent years. The reduction of melatonin with age has been associated with neurodegenerative diseases such as Alzheimer’s disease (AD)and therefore, melatonin has been implicated to have an important clinical role in neuroprotection. Thus, for several years melatonin has attracted increasing attention from the general press with many advertisements touting this indoleamine to act as an aphrodisiac, rejuvenator, protector against diseases and a general wonder drug. However, melatonin formulations appear with no labelling for the correct storage conditions, dosage and side effects, as well as no control for purity and self-medicating with an unregulated product. In addition, there is much controversy surrounding the antioxidative properties of the indolemaine, 6-hydroxymelatonin (6-OHM). Therefore, the first part of this study aims to elucidate the physico-chemical and various stability characteristics of the pineal antioxidant, melatonin, while the second part is devoted to investigating the neuroprotective properties of the primary hepatic metabolite of melatonin, 6-OHM. The physical properties of melatonin were determined using various chemical techniques. This information served to both characterize and confirm the identity of melatonin raw material used in this study. In addition, this information serves to be essential as the physical properties of melatonin have not been reported in detail in literature, to date. Thereafter, using a validated high performance liquid chromatography (HPLC) method, the various physico-chemical and stability characteristics of melatonin were determined. Melatonin was shown to be extremely lipophilic, while the hygroscopic study indicates that melatonin raw material is extremely hygroscopic at temperatures above 40°C, whereas melatonin tablets are hygroscopic when left out of the original container. This study highlights the need for consumers to be aware of the proper storage of melatonin tablets to improve the stability and ensure long term integrity of the compound. Since, melatonin is most often administered orally, thus exposing it to a large variations in pH, within the gastrointestinal tract, it was decided to investigate the stability of melatonin over a range of pH’s and temperatures. The findings imply that melatonin is relatively stable at body temperature when ingested orally and that orally administered slow release preparations of melatonin should be relatively stable and therefore exhibit favourable bioavailability. However melatonin was shown to be unstable in solution. This provides important information and a challenge to the formulators of this drug substance in a liquid dosage form. An assessment of the photostability of melatonin dosage forms using International Committee on Harmonization (ICH) conditions revealed melatonin to be light sensitive and thus indicates a need for careful consideration of the packaging of these drug products. In addition a detailed assessment of the photochemistry and photoproducts formed during the UV photodegradation of melatonin is reported. Melatonin is shown to rapidly degrade in the presence of UV light, with the presence of oxygen accelerating the photodegradation. N1-acetyl-N2-formyl-5-methoxykynurenamine(AFMK) and 6-OHM were identified as the major photoproducts formed and these agents have been shown previously to retain antioxidant activity. One of the concerns of using melatonin in sunscreens is its photostability. However, it is reported in this study that the degraded solution of melatonin still possesses equipotent free radical scavenging ability as melatonin, despite the absence of melatonin in solution. In addition, melatonin is shown to reduce UV-induced oxidative stress in rat skin homogenate. Thus, these results make melatonin a likely candidate for inclusion in sunscreen preparations. Neuronal damage due to oxidative stress has been implicated in several neurodegenerative disorders. 6-OHM is not only formed as the major hepatic metabolite of melatonin, but also when melatonin reacts with toxic radicals as well as UV light. Thus the second part of the study aims to elucidate and further characterize the mechanism behind 6-OHM’s neuroprotection. The results show 6-OHM to be a more potent singlet oxygen and superoxide anion scavenger than melatonin. In addition, the results show 6-OHM to offer protection against, oxidative stress and lipid peroxidation induced by several neurotoxins in the rat brain and hippocampus. The hippocampus is an important region of the brain responsible for the formation of memory and any agent that induces stress in this area has detrimental effects and could lead to various types of dementia. Such agents include quinolinic acid (QA) and iron (II). Histological studies undertaken reveal that 6-OHM is able to protect hippocampal neurons against QA and iron (II) induced necrotic cell death. Immunohistochemical investigations showed that QA moderately induces apoptotic cell death in the hippocampus which is inhibited by both melatonin and 6-OHM. The study sought to elucidate possible mechanisms by which 6-OHM exerts its neuroprotective capabilities and the results show 6-OHM to inhibit the action of cyanide on the mitochondrial electron transport chain (ETC), one of the most common sources of free radicals. In addition, 6-OHM treatment alone, increased ETC activity above basal control levels and the results show 6-OHM to increase complex I activity in the mitochondrial ETC. Electrochemical, ultraviolet/visible spectroscopy (UV/Vis) and HPLC assessment show that an interaction exists between 6-OHM and iron (III) and 6-OHM is able to reduce iron (III) to a more biologically usable form viz. iron (II) which can be incorporated into important biomolecules such as heme. One dire consequence of this interaction is the ready provision of iron (II) to drive the Fenton reaction. However the biological and histological assessments show 6-OHM to prevent iron (II)-induced lipid peroxidation and necrotic cell death and thus, provide evidence of its antioxidant properties. The results also show 6-OHM to promote Hsp70 induction in the hippocampus. Heat shock proteins, especially Hsp 70 plays a role in cytoprotection by capturing denatured proteins and facilitating the refolding of these proteins once the stress has been relieved. 6-OHM treatment alone and together with QA was shown to increase the level of expression of Hsp70, both inducible and cognate forms of the protein. This suggests that 6-OHM helps to protect against cellular protein damage induced by any form of stress the cell may encounter. Melatonin treatment alone and in combination with QA was shown to prevent increases in the level of Hsp70 in the hippocampus, indicating that melatonin was able to reduce oxidative stress induced by QA such that Hsp70 expression was not required. The discovery of neuroprotective agents, such as melatonin and 6-OHM, is becoming important considering the rapid rise in the elderly population and the proportionate increase in neurological disorders. The findings of this study indicate the need for important information regarding the correct storage conditions and stability characteristics of melatonin dosage forms. In addition, the results indicate that 6-OHM has a definite role to play as an antioxidant. Thus further research may favour the use of these agents in the treatment of several neurodegenerative disorders.
- Full Text:
- Date Issued: 2003
- Authors: Maharaj, Deepa Sukhdev
- Date: 2003
- Subjects: Melatonin Nervous system -- Degeneration -- Treatment
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3768 , http://hdl.handle.net/10962/d1003246
- Description: Until the beginning of this decade the antioxidant, melatonin, had been considered as little more than a tranquilizing hormone, responsible for regulating certain circadian and circannual rhythms. However, it is the discovery of melatonin as a free radical scavenger that has generated the most interest in recent years. The reduction of melatonin with age has been associated with neurodegenerative diseases such as Alzheimer’s disease (AD)and therefore, melatonin has been implicated to have an important clinical role in neuroprotection. Thus, for several years melatonin has attracted increasing attention from the general press with many advertisements touting this indoleamine to act as an aphrodisiac, rejuvenator, protector against diseases and a general wonder drug. However, melatonin formulations appear with no labelling for the correct storage conditions, dosage and side effects, as well as no control for purity and self-medicating with an unregulated product. In addition, there is much controversy surrounding the antioxidative properties of the indolemaine, 6-hydroxymelatonin (6-OHM). Therefore, the first part of this study aims to elucidate the physico-chemical and various stability characteristics of the pineal antioxidant, melatonin, while the second part is devoted to investigating the neuroprotective properties of the primary hepatic metabolite of melatonin, 6-OHM. The physical properties of melatonin were determined using various chemical techniques. This information served to both characterize and confirm the identity of melatonin raw material used in this study. In addition, this information serves to be essential as the physical properties of melatonin have not been reported in detail in literature, to date. Thereafter, using a validated high performance liquid chromatography (HPLC) method, the various physico-chemical and stability characteristics of melatonin were determined. Melatonin was shown to be extremely lipophilic, while the hygroscopic study indicates that melatonin raw material is extremely hygroscopic at temperatures above 40°C, whereas melatonin tablets are hygroscopic when left out of the original container. This study highlights the need for consumers to be aware of the proper storage of melatonin tablets to improve the stability and ensure long term integrity of the compound. Since, melatonin is most often administered orally, thus exposing it to a large variations in pH, within the gastrointestinal tract, it was decided to investigate the stability of melatonin over a range of pH’s and temperatures. The findings imply that melatonin is relatively stable at body temperature when ingested orally and that orally administered slow release preparations of melatonin should be relatively stable and therefore exhibit favourable bioavailability. However melatonin was shown to be unstable in solution. This provides important information and a challenge to the formulators of this drug substance in a liquid dosage form. An assessment of the photostability of melatonin dosage forms using International Committee on Harmonization (ICH) conditions revealed melatonin to be light sensitive and thus indicates a need for careful consideration of the packaging of these drug products. In addition a detailed assessment of the photochemistry and photoproducts formed during the UV photodegradation of melatonin is reported. Melatonin is shown to rapidly degrade in the presence of UV light, with the presence of oxygen accelerating the photodegradation. N1-acetyl-N2-formyl-5-methoxykynurenamine(AFMK) and 6-OHM were identified as the major photoproducts formed and these agents have been shown previously to retain antioxidant activity. One of the concerns of using melatonin in sunscreens is its photostability. However, it is reported in this study that the degraded solution of melatonin still possesses equipotent free radical scavenging ability as melatonin, despite the absence of melatonin in solution. In addition, melatonin is shown to reduce UV-induced oxidative stress in rat skin homogenate. Thus, these results make melatonin a likely candidate for inclusion in sunscreen preparations. Neuronal damage due to oxidative stress has been implicated in several neurodegenerative disorders. 6-OHM is not only formed as the major hepatic metabolite of melatonin, but also when melatonin reacts with toxic radicals as well as UV light. Thus the second part of the study aims to elucidate and further characterize the mechanism behind 6-OHM’s neuroprotection. The results show 6-OHM to be a more potent singlet oxygen and superoxide anion scavenger than melatonin. In addition, the results show 6-OHM to offer protection against, oxidative stress and lipid peroxidation induced by several neurotoxins in the rat brain and hippocampus. The hippocampus is an important region of the brain responsible for the formation of memory and any agent that induces stress in this area has detrimental effects and could lead to various types of dementia. Such agents include quinolinic acid (QA) and iron (II). Histological studies undertaken reveal that 6-OHM is able to protect hippocampal neurons against QA and iron (II) induced necrotic cell death. Immunohistochemical investigations showed that QA moderately induces apoptotic cell death in the hippocampus which is inhibited by both melatonin and 6-OHM. The study sought to elucidate possible mechanisms by which 6-OHM exerts its neuroprotective capabilities and the results show 6-OHM to inhibit the action of cyanide on the mitochondrial electron transport chain (ETC), one of the most common sources of free radicals. In addition, 6-OHM treatment alone, increased ETC activity above basal control levels and the results show 6-OHM to increase complex I activity in the mitochondrial ETC. Electrochemical, ultraviolet/visible spectroscopy (UV/Vis) and HPLC assessment show that an interaction exists between 6-OHM and iron (III) and 6-OHM is able to reduce iron (III) to a more biologically usable form viz. iron (II) which can be incorporated into important biomolecules such as heme. One dire consequence of this interaction is the ready provision of iron (II) to drive the Fenton reaction. However the biological and histological assessments show 6-OHM to prevent iron (II)-induced lipid peroxidation and necrotic cell death and thus, provide evidence of its antioxidant properties. The results also show 6-OHM to promote Hsp70 induction in the hippocampus. Heat shock proteins, especially Hsp 70 plays a role in cytoprotection by capturing denatured proteins and facilitating the refolding of these proteins once the stress has been relieved. 6-OHM treatment alone and together with QA was shown to increase the level of expression of Hsp70, both inducible and cognate forms of the protein. This suggests that 6-OHM helps to protect against cellular protein damage induced by any form of stress the cell may encounter. Melatonin treatment alone and in combination with QA was shown to prevent increases in the level of Hsp70 in the hippocampus, indicating that melatonin was able to reduce oxidative stress induced by QA such that Hsp70 expression was not required. The discovery of neuroprotective agents, such as melatonin and 6-OHM, is becoming important considering the rapid rise in the elderly population and the proportionate increase in neurological disorders. The findings of this study indicate the need for important information regarding the correct storage conditions and stability characteristics of melatonin dosage forms. In addition, the results indicate that 6-OHM has a definite role to play as an antioxidant. Thus further research may favour the use of these agents in the treatment of several neurodegenerative disorders.
- Full Text:
- Date Issued: 2003
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
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
Application of high-performance liquid chromatography to the analysis, stability and pharmacokinetics of erythromycin
- Authors: Stubbs, Christopher
- Date: 1988
- Subjects: Erythromycin High performance liquid chromatography
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3809 , http://hdl.handle.net/10962/d1004372
- Description: Erythromycin is a macrolide antibiotic used mainly in the treatment of infections caused by gram-positive organisms. Erythromycin base is rap idly degraded in acidic media necessitating the use of structurally modified erythromycin derivatives or acid resistant dosage forms in order to decrease gastric inactivation of the drug. The majority of pharmacokinetic studies to-date have utilized relatively non-specific microbiological assay procedures which are unable to differentiate between concentrations of active erythromycin base and the inactive pro-drug derivatives. A high-performance liquid chromatographic (HPLC) technique is described for the simultaneous determination of erythromycin base and propionate (inactive pro-drug form) in human serum and urine following the oral administration of erythromycin estolate, an acid stable derivative of erythromycin. The method involves a solid-phase extraction step prior to chromatography on a C18 reversed-phase column with coulometric electrochemical detection. Sample handling and storage techniques are presented which minimize hydrolysis of the inactive ester moiety between sample collection and analysis, thereby more accurately reflecting the in vivo situation than in previously published studies. Results from single dose pharmacokinetic studies indicate that only 10-15% of the total erythromycin concentration in vivo is present as the active base component following oral administration of erythromycin estolate. This percentage increases to approximately 25% during multiple dose administration. Novel urinary excretion data are presented which reveal that approximately 40% and 55% of the total erythromycin excreted in urine is excreted as erythromycin base following single and multiple dosages respectively. Computer fitting of mean serum concentration-time data revealed that an open one compartment model with linear first order absorption and elimination best described the absorption and disposition of erythromycin, although poor computer fits for individual data sets were observed. Some evidence of non-linear elimination is presented utilizing both compartmental and non-compartmental pharmacokinetic techniques. Large intra-and inter-personal variability in erythromycin absorption and disposition was experienced which was evaluated in five subjects who each received one 500 mg erythromycin estolate tablet from the same batch, on three separate occasions. In addition. an HPLC method is described for the analysis of "total erythromycin" concentrations following erythromycin estolate administration which involves hydrolysis of the ester component prior to chromatography. as well as an HPLC method utilizing amperometric electrochemical detection capable of monitoring the stability of erythromycin base in stored biological fluids. These methods were uti I ized in various stability studies involving erythromycin base and propionate as well as for the analysis of erythromycin estolate dosage forms.
- Full Text:
- Date Issued: 1988
- Authors: Stubbs, Christopher
- Date: 1988
- Subjects: Erythromycin High performance liquid chromatography
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3809 , http://hdl.handle.net/10962/d1004372
- Description: Erythromycin is a macrolide antibiotic used mainly in the treatment of infections caused by gram-positive organisms. Erythromycin base is rap idly degraded in acidic media necessitating the use of structurally modified erythromycin derivatives or acid resistant dosage forms in order to decrease gastric inactivation of the drug. The majority of pharmacokinetic studies to-date have utilized relatively non-specific microbiological assay procedures which are unable to differentiate between concentrations of active erythromycin base and the inactive pro-drug derivatives. A high-performance liquid chromatographic (HPLC) technique is described for the simultaneous determination of erythromycin base and propionate (inactive pro-drug form) in human serum and urine following the oral administration of erythromycin estolate, an acid stable derivative of erythromycin. The method involves a solid-phase extraction step prior to chromatography on a C18 reversed-phase column with coulometric electrochemical detection. Sample handling and storage techniques are presented which minimize hydrolysis of the inactive ester moiety between sample collection and analysis, thereby more accurately reflecting the in vivo situation than in previously published studies. Results from single dose pharmacokinetic studies indicate that only 10-15% of the total erythromycin concentration in vivo is present as the active base component following oral administration of erythromycin estolate. This percentage increases to approximately 25% during multiple dose administration. Novel urinary excretion data are presented which reveal that approximately 40% and 55% of the total erythromycin excreted in urine is excreted as erythromycin base following single and multiple dosages respectively. Computer fitting of mean serum concentration-time data revealed that an open one compartment model with linear first order absorption and elimination best described the absorption and disposition of erythromycin, although poor computer fits for individual data sets were observed. Some evidence of non-linear elimination is presented utilizing both compartmental and non-compartmental pharmacokinetic techniques. Large intra-and inter-personal variability in erythromycin absorption and disposition was experienced which was evaluated in five subjects who each received one 500 mg erythromycin estolate tablet from the same batch, on three separate occasions. In addition. an HPLC method is described for the analysis of "total erythromycin" concentrations following erythromycin estolate administration which involves hydrolysis of the ester component prior to chromatography. as well as an HPLC method utilizing amperometric electrochemical detection capable of monitoring the stability of erythromycin base in stored biological fluids. These methods were uti I ized in various stability studies involving erythromycin base and propionate as well as for the analysis of erythromycin estolate dosage forms.
- Full Text:
- Date Issued: 1988
Aspects of prostacyclin in experimental hypertension
- Authors: Botha, Julia Hilary
- Date: 1983
- Subjects: Prostacyclin Prostaglandins Prostaglandin endoperoxides Thromboxanes Hypertension
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3825 , http://hdl.handle.net/10962/d1006109
- Description: A new prostaglandin - prostaglandin X (later renamed prostacyclin or prostaglandin I₂ (PGI₂)), was discovered by Moncada, Gryglewski, Bunting and Vane in 1976. This unstable substance was shown to be produced by vascular tissue and to be a vasodilator and the most potent endogenous inhibitor of platelet aggregation known. Because of its properties, it appeared that a lack of it may be related to the development and or maintenance of hypertension, a disorder featuring vasoconstriction and an increased tendency to arterial thrombosis. The present studies aimed to investigate this possibility using a rat model. A bioassay for prostacyclin was first perfected. This consisted of a modification of the method used by Moncada, Higgs and Vane (1977): PGI₂ released by rat aortic strips, during incubation in tris buffer, was measured by assessing the ability of the incubate to inhibit adenosine diphosphate induced aggregation of human platelets, as compared to the inhibitory effect of standard prostacyclin sodium salt. The specificity of the assay for the detection of PGI₂ was tested. The abil ity of hypertensive rat aorta to release prostacycl in was investigated in two studies. The first compared aortas of Wistar rats of the New Zealand genetically hypertensive strain (GH) with those of matched normotensive Wistar controls. In the second study, hypertension was induced by wrappi ng the ri ght kidney with surgical silk and removing the contralateral kidney. Ten weeks later, aortic generation of prostacyclin by these animals was compared with that of matched sham controls which had received identical surgical manipulation but for the application of silk to the right kidney. Contrary to expectation, in both forms of hypertension, aortas of the rats with elevated pressure produced consistently more prostacyclin than those of matched controls. In order to discover more about the relationship between elevated pressure and elevated PGI₂ production, the effect of pressure reduction with hypotensive agents on the ability of GH rat aortas to produce prostacyclin, was investigated. After pressure had been controlled within normal range for one week (achieved by oral administration of furosemide, dihydralazine and reserpine for one month), aortic PGI₂ was reduced in comparison with matched GH controls. However, the reduction was not consistent and statistical significance was not reached. Because it was subsequently reported by other workers, that some of the hypotensive agents which had been employed may effect prostaglandin levels per se, no conclusions could be drawn from this study as to any possible direct relationships between pressure and aortic prostacyclin generating capacity. A further means of reducing elevated pressure (which had no inherent effect on prostaglandin levels) was thus sought. A mechanical method was eventually selected, application of a silver clip to the aortas of GH rats, just below the diaphragm, producing an immediate reduction in pressure distal to the constriction. Eighteen hours with later, PGI₂ production by these distal aortas those of matched sham GH controls and was was compared found to be consistently reduced. These results indicate that the ability to produce PGI₂ may be influenced by prior local pressure changes and that the increased capacity of hypertensive rat aortas to generate prostacyclin may be related to the increased mechanical transmural stress consequent on elevated pressure. Since haemostatic balance must be influenced not only by vascular PGI₂ generation but also by platelet sensitivity to PGI₂, the response of GH platelets to the anti-aggregatory effect of prostacyc1in was also investigated. As it had been shown by Sinzinger, Si1berbauer, Horsch and Gall (1981) that intra-arterial infusion of PGI₂ in humans decreased platelet sensitivity to the substance, the possibility existed that platelet sensitivity in hypertension might be reduced. This hypothesis was, however, invalidated as the sensitivity of GH platelets to the anti-aggregatory effect of PGI₂ was almost identical to that of normotensive controls. The shortcomings of the methodology and the possible importance of these findings in the hypertensive animal are discussed. The idea that elevated PGI₂ in hypertension may play a protective role both with respect to platelet aggregation and in attenuating further pressure rises is considered. It is finally suggested that it will be possible to draw more accurate conclusions as to the meaning of the increased PGI₂ generation in hypertension (both in relation to vascular tone and platelet function) only when details of production of, and sensitivity to, thromboxane A₂ are known. Thromboxane A₂ (TXA₂) is a vasoconstrictor and promotor of aggregation (Hamberg, Svensson and Samuelson, 1975) and it may be that, despite elevated vascular PGI₂ generation, the TXA₂/PGI₂ balance is still tipped in favour of vasoconstriction and platelet aggregation in hypertension.
- Full Text:
- Date Issued: 1983
- Authors: Botha, Julia Hilary
- Date: 1983
- Subjects: Prostacyclin Prostaglandins Prostaglandin endoperoxides Thromboxanes Hypertension
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3825 , http://hdl.handle.net/10962/d1006109
- Description: A new prostaglandin - prostaglandin X (later renamed prostacyclin or prostaglandin I₂ (PGI₂)), was discovered by Moncada, Gryglewski, Bunting and Vane in 1976. This unstable substance was shown to be produced by vascular tissue and to be a vasodilator and the most potent endogenous inhibitor of platelet aggregation known. Because of its properties, it appeared that a lack of it may be related to the development and or maintenance of hypertension, a disorder featuring vasoconstriction and an increased tendency to arterial thrombosis. The present studies aimed to investigate this possibility using a rat model. A bioassay for prostacyclin was first perfected. This consisted of a modification of the method used by Moncada, Higgs and Vane (1977): PGI₂ released by rat aortic strips, during incubation in tris buffer, was measured by assessing the ability of the incubate to inhibit adenosine diphosphate induced aggregation of human platelets, as compared to the inhibitory effect of standard prostacyclin sodium salt. The specificity of the assay for the detection of PGI₂ was tested. The abil ity of hypertensive rat aorta to release prostacycl in was investigated in two studies. The first compared aortas of Wistar rats of the New Zealand genetically hypertensive strain (GH) with those of matched normotensive Wistar controls. In the second study, hypertension was induced by wrappi ng the ri ght kidney with surgical silk and removing the contralateral kidney. Ten weeks later, aortic generation of prostacyclin by these animals was compared with that of matched sham controls which had received identical surgical manipulation but for the application of silk to the right kidney. Contrary to expectation, in both forms of hypertension, aortas of the rats with elevated pressure produced consistently more prostacyclin than those of matched controls. In order to discover more about the relationship between elevated pressure and elevated PGI₂ production, the effect of pressure reduction with hypotensive agents on the ability of GH rat aortas to produce prostacyclin, was investigated. After pressure had been controlled within normal range for one week (achieved by oral administration of furosemide, dihydralazine and reserpine for one month), aortic PGI₂ was reduced in comparison with matched GH controls. However, the reduction was not consistent and statistical significance was not reached. Because it was subsequently reported by other workers, that some of the hypotensive agents which had been employed may effect prostaglandin levels per se, no conclusions could be drawn from this study as to any possible direct relationships between pressure and aortic prostacyclin generating capacity. A further means of reducing elevated pressure (which had no inherent effect on prostaglandin levels) was thus sought. A mechanical method was eventually selected, application of a silver clip to the aortas of GH rats, just below the diaphragm, producing an immediate reduction in pressure distal to the constriction. Eighteen hours with later, PGI₂ production by these distal aortas those of matched sham GH controls and was was compared found to be consistently reduced. These results indicate that the ability to produce PGI₂ may be influenced by prior local pressure changes and that the increased capacity of hypertensive rat aortas to generate prostacyclin may be related to the increased mechanical transmural stress consequent on elevated pressure. Since haemostatic balance must be influenced not only by vascular PGI₂ generation but also by platelet sensitivity to PGI₂, the response of GH platelets to the anti-aggregatory effect of prostacyc1in was also investigated. As it had been shown by Sinzinger, Si1berbauer, Horsch and Gall (1981) that intra-arterial infusion of PGI₂ in humans decreased platelet sensitivity to the substance, the possibility existed that platelet sensitivity in hypertension might be reduced. This hypothesis was, however, invalidated as the sensitivity of GH platelets to the anti-aggregatory effect of PGI₂ was almost identical to that of normotensive controls. The shortcomings of the methodology and the possible importance of these findings in the hypertensive animal are discussed. The idea that elevated PGI₂ in hypertension may play a protective role both with respect to platelet aggregation and in attenuating further pressure rises is considered. It is finally suggested that it will be possible to draw more accurate conclusions as to the meaning of the increased PGI₂ generation in hypertension (both in relation to vascular tone and platelet function) only when details of production of, and sensitivity to, thromboxane A₂ are known. Thromboxane A₂ (TXA₂) is a vasoconstrictor and promotor of aggregation (Hamberg, Svensson and Samuelson, 1975) and it may be that, despite elevated vascular PGI₂ generation, the TXA₂/PGI₂ balance is still tipped in favour of vasoconstriction and platelet aggregation in hypertension.
- Full Text:
- Date Issued: 1983
Aspects of the transdermal permeation and analysis of betamethasone 17-valerate
- Authors: Smith, Eric W
- Date: 1988
- Subjects: Transdermal medication Skin absorption Dermatologic agents
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3815 , http://hdl.handle.net/10962/d1004741
- Description: The current world-wide interest in transdermal drug delivery makes the prospect of valid in vitro diffusion cell methodology highly attractive. A new laboratory diffusion cell has been designed and constructed based on theoretical principles and practical permeation reports surveyed in recent literature, and has been applied to the monitoring of betamethasone 17-valerate permeation. The cell performance has been validated with respect to hydrodynamic mixing efficiency and temperature of the receptor phase. The steady-state permeation of this corticosteroid has been monitored through various synthetic and animal membranes in order to select the most appropriate media for in vitro study. The permeation of betamethasone 17-valerate has been monitored from various types of commercial and extemporaneously prepared semisolid topical formulation (cream, lotion, ointment and scalp application), through silicone membrane, human and weanling pig stratum corneum, and full thickness hairless mouse skin, and these in vitro results have been compared to data from in vivo blanching assays, using the same formulations, in an attempt to correlate the findings. This experimental methodology has necessitated the development of ancillary analytical techniques. A column-switching high-performance liquid chromatographic method has been developed for the rapid on-line clean-up and analysis of betamethasone 17-valerate contained in the various topical formulations, which minimizes sample handling and extraction procedures. The method has been modified for the analysis of this corticosteroid in the isopropyl myristate receptor phase used in the in vitro permeation experiments, and scintillation counting of tritium-labelled water has been used to verify the integrity of the animal membranes. The comparison of in vitro permeation and in vivo blanching results indicate good correlation of the data in certain instances. The closest correlations have been observed when the human stratum corneum has been used in vitro and these results are compared to data from the occluded mode of the blanching assay. The results of the porcine and murine media have also correlated with the human in vivo data, whereas the silicone membrane appears applicable only in certain in vitro experiments. The results indicate that valid, comparative percutaneous absorption data may be obtained in vitro by using a well designed, validated diffusion cell system.
- Full Text:
- Date Issued: 1988
- Authors: Smith, Eric W
- Date: 1988
- Subjects: Transdermal medication Skin absorption Dermatologic agents
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3815 , http://hdl.handle.net/10962/d1004741
- Description: The current world-wide interest in transdermal drug delivery makes the prospect of valid in vitro diffusion cell methodology highly attractive. A new laboratory diffusion cell has been designed and constructed based on theoretical principles and practical permeation reports surveyed in recent literature, and has been applied to the monitoring of betamethasone 17-valerate permeation. The cell performance has been validated with respect to hydrodynamic mixing efficiency and temperature of the receptor phase. The steady-state permeation of this corticosteroid has been monitored through various synthetic and animal membranes in order to select the most appropriate media for in vitro study. The permeation of betamethasone 17-valerate has been monitored from various types of commercial and extemporaneously prepared semisolid topical formulation (cream, lotion, ointment and scalp application), through silicone membrane, human and weanling pig stratum corneum, and full thickness hairless mouse skin, and these in vitro results have been compared to data from in vivo blanching assays, using the same formulations, in an attempt to correlate the findings. This experimental methodology has necessitated the development of ancillary analytical techniques. A column-switching high-performance liquid chromatographic method has been developed for the rapid on-line clean-up and analysis of betamethasone 17-valerate contained in the various topical formulations, which minimizes sample handling and extraction procedures. The method has been modified for the analysis of this corticosteroid in the isopropyl myristate receptor phase used in the in vitro permeation experiments, and scintillation counting of tritium-labelled water has been used to verify the integrity of the animal membranes. The comparison of in vitro permeation and in vivo blanching results indicate good correlation of the data in certain instances. The closest correlations have been observed when the human stratum corneum has been used in vitro and these results are compared to data from the occluded mode of the blanching assay. The results of the porcine and murine media have also correlated with the human in vivo data, whereas the silicone membrane appears applicable only in certain in vitro experiments. The results indicate that valid, comparative percutaneous absorption data may be obtained in vitro by using a well designed, validated diffusion cell system.
- Full Text:
- Date Issued: 1988
Biologically active natural products from South African marine invertebrates
- Authors: Hooper, Gregory John
- Date: 1997
- Subjects: Natural products -- South Africa Marine metabolites -- South Africa Marine invertebrates -- South Africa
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3761 , http://hdl.handle.net/10962/d1003239
- Description: This thesis describes the chemical and biological investigation of the extracts of six different marine invertebrate organisms collected along the South African coastline. The work on these extracts has resulted in the isolation and structural elucidation of twenty-one previously undescribed secondary metabolites; The history of marine natural product chemistry in South Africa has not previously been reviewed and so a comprehensive review covering the literature from the 1940's up until the end of 1995 is presented here. The marine ascidian Pseudodistoma species collected in the Tsitsikamma Marine Reserve was shown to contain four new unsaturated amino alcohols [47], [48], [49] and [50] which were isolated as their acetyl derivatives. These compounds exhibited strong antimicrobial activity. Four new pyrroloiminoquinone alkaloids, the tsitsikammamines A [90] to D [93],were isolated from a new genus of Latrunculid sponge collected in the Tsitsikamma Marine Reserve. These highly pigmented compounds also possessed strong antimicrobial activity. An investigation of two phenotypic colour variants of the soft coral Capnella thyrsoidea resulted in the isolation of the known steroid 5α-pregna-1, 20-dien-3-one [97] and an additional six new metabolites, 16β-hydroxy-5α-pregna-1 ,20-dien-3-one 16-acetate [98], 3α,16β-dihydroxy-5α-pregna-1, 20-diene 3,16-diacetate [99] and four xenicane diterpenes, the tsitsixenicins A [100] to D [103]. This is the first reported isolation of xenicane diterpenes from the soft coral family Nephtheiidae. Tsitsixenicin A and B showed good anti-inflammatory activity by inhibiting superoxide production in both rabbit and human cell neutrophils. A further four new metabolites were isolated from two soft corals which could only be identified to the genus level and were designated Alcyonium species A and species B. Alcyonium species A was collected in the Tsitsikamma Marine Reserve and yielded two new polyhydroxysterols, cholest-5-ene-3β, 7β, 19-triol 19-acetate [121] and cholest-5,24-diene-3β, 7β, 19-triol 19-acetate [122]. The soft coral Alcyonium species B was collected off Aliwal Shoal and was found to contain two known xenicane diterpenes, 9-deacetoxy-14, 15-deepoxyxeniculin [110] and zahavin A [16], and two new xenicane diterpenes, 7 -epoxyzahavin A [123] and xeniolide C [124]. Compounds [110], [16] and [123] exhibited strong anti-inflammatory activity and compounds [110] and [16] showed good antithrombotic activity. The endemic soft coral A/cyanium fauri collected at Riet Point near Port Alfred yielded the new sesquiterpene hydroquinone rietone [141] in high yierd, fogether with the minor compounds 8'-acetoxyrietone [142] and 8'-desoxyrietone [143]. Rietone exhibited moderate activity in the NCl's in-vitro anti-HIV bioassays.
- Full Text:
- Date Issued: 1997
- Authors: Hooper, Gregory John
- Date: 1997
- Subjects: Natural products -- South Africa Marine metabolites -- South Africa Marine invertebrates -- South Africa
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3761 , http://hdl.handle.net/10962/d1003239
- Description: This thesis describes the chemical and biological investigation of the extracts of six different marine invertebrate organisms collected along the South African coastline. The work on these extracts has resulted in the isolation and structural elucidation of twenty-one previously undescribed secondary metabolites; The history of marine natural product chemistry in South Africa has not previously been reviewed and so a comprehensive review covering the literature from the 1940's up until the end of 1995 is presented here. The marine ascidian Pseudodistoma species collected in the Tsitsikamma Marine Reserve was shown to contain four new unsaturated amino alcohols [47], [48], [49] and [50] which were isolated as their acetyl derivatives. These compounds exhibited strong antimicrobial activity. Four new pyrroloiminoquinone alkaloids, the tsitsikammamines A [90] to D [93],were isolated from a new genus of Latrunculid sponge collected in the Tsitsikamma Marine Reserve. These highly pigmented compounds also possessed strong antimicrobial activity. An investigation of two phenotypic colour variants of the soft coral Capnella thyrsoidea resulted in the isolation of the known steroid 5α-pregna-1, 20-dien-3-one [97] and an additional six new metabolites, 16β-hydroxy-5α-pregna-1 ,20-dien-3-one 16-acetate [98], 3α,16β-dihydroxy-5α-pregna-1, 20-diene 3,16-diacetate [99] and four xenicane diterpenes, the tsitsixenicins A [100] to D [103]. This is the first reported isolation of xenicane diterpenes from the soft coral family Nephtheiidae. Tsitsixenicin A and B showed good anti-inflammatory activity by inhibiting superoxide production in both rabbit and human cell neutrophils. A further four new metabolites were isolated from two soft corals which could only be identified to the genus level and were designated Alcyonium species A and species B. Alcyonium species A was collected in the Tsitsikamma Marine Reserve and yielded two new polyhydroxysterols, cholest-5-ene-3β, 7β, 19-triol 19-acetate [121] and cholest-5,24-diene-3β, 7β, 19-triol 19-acetate [122]. The soft coral Alcyonium species B was collected off Aliwal Shoal and was found to contain two known xenicane diterpenes, 9-deacetoxy-14, 15-deepoxyxeniculin [110] and zahavin A [16], and two new xenicane diterpenes, 7 -epoxyzahavin A [123] and xeniolide C [124]. Compounds [110], [16] and [123] exhibited strong anti-inflammatory activity and compounds [110] and [16] showed good antithrombotic activity. The endemic soft coral A/cyanium fauri collected at Riet Point near Port Alfred yielded the new sesquiterpene hydroquinone rietone [141] in high yierd, fogether with the minor compounds 8'-acetoxyrietone [142] and 8'-desoxyrietone [143]. Rietone exhibited moderate activity in the NCl's in-vitro anti-HIV bioassays.
- Full Text:
- Date Issued: 1997
Biopharmaceutics and pharmacokinetics of the macrolide antibiotic Josamycin
- Authors: Skinner, Michael Fredrick
- Date: 1992
- Subjects: Antibiotics -- Bioavailability , Antibiotics -- Pharmacokinetics
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3791 , http://hdl.handle.net/10962/d1003269
- Description: The investigations detailed herein have been conducted to address various aspects of the biopharmaceutics and pharmacokinetics of josamycin which to-date, have received little or no attention in the literature. Areas of investigation have included the selective determination of josamycin in serum and urine samples, the stability of josamycin in stored biological samples, intrinsic dissolution rates, solubility, acid and alkali stability and bioavailability and pharmacokinetics after dosing with a solution, powder and tablets. High performance liquid chromatography (HPLC) was used as the main analytical tool throughout these studies and proved to be highly versatile for the determination of josamycin in a number of different media. HPLC analysis afforded simple yet accurate determination of josamycin in samples from dissolution, solubility, tablet content and stability studies. Furthermore, the specificity afforded by HPLC was particularly useful for the separation of josamycin from degradation products formed in acid and alkali media. Since metabolites of josamycin are microbiologically active, microbiological assays do not determine the concentration solely of josamycin. An analytical method capable of the selective determination of josamycin in serum and urine samples is therefore required for the procurement of reliable bioavailability and pharmacokinetic data. HPLC affords this selectivity and a method for the selective determination of josamycin in serum and urine was successfully developed. The assay was simple yet precise, accurate and sensitive. Furthermore, it was well suited to the determination of josamycin in a large number of biological samples. Its success was largely due to the use of a solid phase extraction step using C₁₈ extraction columns, with a highly specific wash sequence followed by a phase separation step after elution from the extraction column. Chromatography was performed on a C₁₈ reversed-phase analytical column with UV detection of josamycin and internal standard at 231 nm and at 204 nm respectively using a programmable multi-wavelength detector. Only slight modification of the assay described should enable the selective determination of the metabolites of josamycin. This assay, therefore, lays the groundwork for future investigations into the pharmacokinetics of these metabolites. The re-usability of extraction columns was assessed in an attempt to reduce the cost of sample analysis. It was found that extraction columns could be used twice for the extraction of serum samples and up to four times for the extraction of urine samples. The difference between the re-usability of extraction columns for serum and urine samples was ascribed to various differences in the composition of the sample matrix. The stability of josamycin in stored serum and urine samples was also assessed.
- Full Text:
- Date Issued: 1992
- Authors: Skinner, Michael Fredrick
- Date: 1992
- Subjects: Antibiotics -- Bioavailability , Antibiotics -- Pharmacokinetics
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3791 , http://hdl.handle.net/10962/d1003269
- Description: The investigations detailed herein have been conducted to address various aspects of the biopharmaceutics and pharmacokinetics of josamycin which to-date, have received little or no attention in the literature. Areas of investigation have included the selective determination of josamycin in serum and urine samples, the stability of josamycin in stored biological samples, intrinsic dissolution rates, solubility, acid and alkali stability and bioavailability and pharmacokinetics after dosing with a solution, powder and tablets. High performance liquid chromatography (HPLC) was used as the main analytical tool throughout these studies and proved to be highly versatile for the determination of josamycin in a number of different media. HPLC analysis afforded simple yet accurate determination of josamycin in samples from dissolution, solubility, tablet content and stability studies. Furthermore, the specificity afforded by HPLC was particularly useful for the separation of josamycin from degradation products formed in acid and alkali media. Since metabolites of josamycin are microbiologically active, microbiological assays do not determine the concentration solely of josamycin. An analytical method capable of the selective determination of josamycin in serum and urine samples is therefore required for the procurement of reliable bioavailability and pharmacokinetic data. HPLC affords this selectivity and a method for the selective determination of josamycin in serum and urine was successfully developed. The assay was simple yet precise, accurate and sensitive. Furthermore, it was well suited to the determination of josamycin in a large number of biological samples. Its success was largely due to the use of a solid phase extraction step using C₁₈ extraction columns, with a highly specific wash sequence followed by a phase separation step after elution from the extraction column. Chromatography was performed on a C₁₈ reversed-phase analytical column with UV detection of josamycin and internal standard at 231 nm and at 204 nm respectively using a programmable multi-wavelength detector. Only slight modification of the assay described should enable the selective determination of the metabolites of josamycin. This assay, therefore, lays the groundwork for future investigations into the pharmacokinetics of these metabolites. The re-usability of extraction columns was assessed in an attempt to reduce the cost of sample analysis. It was found that extraction columns could be used twice for the extraction of serum samples and up to four times for the extraction of urine samples. The difference between the re-usability of extraction columns for serum and urine samples was ascribed to various differences in the composition of the sample matrix. The stability of josamycin in stored serum and urine samples was also assessed.
- Full Text:
- Date Issued: 1992
Biopharmaceutics of phenylpropanolamine
- Authors: Dowse, Roslind
- Date: 1984
- Subjects: Biopharmaceutics Pharmacokinetics Phenylpropanolamine Pharmacology High performance liquid chromatography
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3818 , http://hdl.handle.net/10962/d1004915
- Description: Phenylpropanolamine (PPA), a sympathomimetic amine, has been widely used over the past 40 years as a decongestant and, in much larger dosages, as an appetite suppressant. Considerable interest has recently been shown in this drug due to its increasing popularity as an over-the-counter anorectic agent. Much controversy exists concerning the unfavourable side-effects of PPA resulting from the higher doses required for appetite suppression and the potential of this drug for abuse. A literature search revealed a paucity of information concerning the determination of PPA in biological fluids and, most noticeably, on the pharmacokinetics of this drug. An original method for determining PPA in serum and urine using high performance liquid chromatography (HPLC) which has increased sensitivity over other published HPLC methods is presented here. The simplicity of the extraction from biological fluids and subsequent determination by HPLC, enables concentrations of PPA to be monitored after a single dose of the drug. This method is therefore readily applicable to bioavailability and pharmacokinetic studies. The dissolution profiles of 4 sustained-release formulations of PPA were determined in a modified USP rotating paddle apparatus and the samples analysed using HPLC. A mathematical equation was applied to these data which are expressed in terms of dissolution parameters. Oral test dosage forms and solutions of PPA were investigated in bioavailability trials using the developed HPLC method to analyse the urine and serum samples. Linear one body compartment kinetics were assumed and the WagnerNelson method used to transform in vivo serum data to absorption plots which were then fitted to the well known Weibull equation. In order to more appropriately characterize the kinetic processes of absorption, distribution and elimination, a more complex model was utilized which involved numerical integration of a series of differential equations. The data were fitted to these models using nonlinear regression techniques. The pharmacokinetics of PPA are shown to exhibit some evidence of nonlinearity. The absorption of the drug appears to be di scontinuous and PPA seems to favour a two body compartment model.
- Full Text:
- Date Issued: 1984
- Authors: Dowse, Roslind
- Date: 1984
- Subjects: Biopharmaceutics Pharmacokinetics Phenylpropanolamine Pharmacology High performance liquid chromatography
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3818 , http://hdl.handle.net/10962/d1004915
- Description: Phenylpropanolamine (PPA), a sympathomimetic amine, has been widely used over the past 40 years as a decongestant and, in much larger dosages, as an appetite suppressant. Considerable interest has recently been shown in this drug due to its increasing popularity as an over-the-counter anorectic agent. Much controversy exists concerning the unfavourable side-effects of PPA resulting from the higher doses required for appetite suppression and the potential of this drug for abuse. A literature search revealed a paucity of information concerning the determination of PPA in biological fluids and, most noticeably, on the pharmacokinetics of this drug. An original method for determining PPA in serum and urine using high performance liquid chromatography (HPLC) which has increased sensitivity over other published HPLC methods is presented here. The simplicity of the extraction from biological fluids and subsequent determination by HPLC, enables concentrations of PPA to be monitored after a single dose of the drug. This method is therefore readily applicable to bioavailability and pharmacokinetic studies. The dissolution profiles of 4 sustained-release formulations of PPA were determined in a modified USP rotating paddle apparatus and the samples analysed using HPLC. A mathematical equation was applied to these data which are expressed in terms of dissolution parameters. Oral test dosage forms and solutions of PPA were investigated in bioavailability trials using the developed HPLC method to analyse the urine and serum samples. Linear one body compartment kinetics were assumed and the WagnerNelson method used to transform in vivo serum data to absorption plots which were then fitted to the well known Weibull equation. In order to more appropriately characterize the kinetic processes of absorption, distribution and elimination, a more complex model was utilized which involved numerical integration of a series of differential equations. The data were fitted to these models using nonlinear regression techniques. The pharmacokinetics of PPA are shown to exhibit some evidence of nonlinearity. The absorption of the drug appears to be di scontinuous and PPA seems to favour a two body compartment model.
- Full Text:
- Date Issued: 1984
Chemical and spectroscopic studies of the capsular polysaccharides of some klebsiella and escherichia coli serotypes
- Authors: Stanley, Shawn Mark Ross
- Date: 1990
- Subjects: Polysaccharides , Klebsiella , Escherichia
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3736 , http://hdl.handle.net/10962/d1001525
- Description: The work described in this thesis forms part of an international programme concerned with the structure elucidation of the capsular antigens of some Enterobacteriaceae. Many of the Klebsiella and some of the Escherichia coli are pathogenic to man and, hence, they are of interest. The virulence of bacteria is a multifactorial phenomenon, in which characteristic traits of bacteria and their hosts play comparable and complementary roles. It is accepted that pathogens are more virulent when encapsulated, because, nearly all disease causing bacteria have a capsule when freshly isolated from the host. This increase in pathogenicity is related, in part, to the capsular polysaccharides' ability to avoid or attenuate the host defence mechanisms. In the majority of cases the protective aspects of the capsule are overcome in the latter stages of infection when the formation of specific antibodies by the host has occurred. However there are situations in which an immune state of the infected host is virtually never reached, and susceptiblity to the infecting bacteria is maintained even in the advanced stage of an infection. Explanation of this phenomenon becomes possible by analysing the structure of the polysaccharides. The inability of the host to raise an immune response to the capsule may be because the structure of the polysaccharide is similar or identical to the host's carbohydrates. The serological and pathogenic relatedness of encapsulated E. coli and Klebsiella, to the encapsulated strains of other genera, is based on structural identity or similarity of the respective capsules. Capsular polysaccharides are analysed by both chemical and instrumental methods, and, at present, nuclear magnetic resonance spectroscopy is the most important analytical technique
- Full Text:
- Date Issued: 1990
- Authors: Stanley, Shawn Mark Ross
- Date: 1990
- Subjects: Polysaccharides , Klebsiella , Escherichia
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3736 , http://hdl.handle.net/10962/d1001525
- Description: The work described in this thesis forms part of an international programme concerned with the structure elucidation of the capsular antigens of some Enterobacteriaceae. Many of the Klebsiella and some of the Escherichia coli are pathogenic to man and, hence, they are of interest. The virulence of bacteria is a multifactorial phenomenon, in which characteristic traits of bacteria and their hosts play comparable and complementary roles. It is accepted that pathogens are more virulent when encapsulated, because, nearly all disease causing bacteria have a capsule when freshly isolated from the host. This increase in pathogenicity is related, in part, to the capsular polysaccharides' ability to avoid or attenuate the host defence mechanisms. In the majority of cases the protective aspects of the capsule are overcome in the latter stages of infection when the formation of specific antibodies by the host has occurred. However there are situations in which an immune state of the infected host is virtually never reached, and susceptiblity to the infecting bacteria is maintained even in the advanced stage of an infection. Explanation of this phenomenon becomes possible by analysing the structure of the polysaccharides. The inability of the host to raise an immune response to the capsule may be because the structure of the polysaccharide is similar or identical to the host's carbohydrates. The serological and pathogenic relatedness of encapsulated E. coli and Klebsiella, to the encapsulated strains of other genera, is based on structural identity or similarity of the respective capsules. Capsular polysaccharides are analysed by both chemical and instrumental methods, and, at present, nuclear magnetic resonance spectroscopy is the most important analytical technique
- Full Text:
- Date Issued: 1990
Design, development and evaluation of encapsulated oral controlled release theophylline mini-tablets
- Authors: Munday, Dale Leslie
- Date: 1991
- Subjects: Drugs -- Administration , Drugs -- Bioavailability , Drugs -- Controlled release , Drugs -- Dosage forms , Tablets (Medicine) , Biopharmaceutics , Drugs -- Testing
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3777 , http://hdl.handle.net/10962/d1003255 , Drugs -- Administration , Drugs -- Bioavailability , Drugs -- Controlled release , Drugs -- Dosage forms , Tablets (Medicine) , Biopharmaceutics , Drugs -- Testing
- Description: Conventional solid dosage forms often lead to fluctuations which exceed the maximum safe therapeutic level and/or decline below the minimum effective level. It is recognised that many drugs for chronic administration should be administered on a schedule that maintains plasma drug concentration within the therapeutic window. Research in controlled release dosage forms aims at designing a system with a zero-order input (eg, ideally to deliver 8.33% of the dose per hour over a 12 hour duration), producing steady state plasma drug levels. Oral dministration of drugs prepared as a controlled release formulation is extremely popular, and has attracted the attention of pharmaceutical scientists during the last decade. This has been due to the simultaneous convergence of various factors (eg, discovery of novel polymers and devices, better understanding of formulation and physiological constraints, expiration of existing patents, prohibitive cost of developing new drug entities), involved in the development of these delivery systems. Controlled release oral products can be formulated as single or multiple unit dosage forms and the relative merits of multiple unit forms with their own rate controlling systems are well established. This work describes the development of a relatively inexpensive multiple-unit capsule dosage form of theophylline containing coated mini-tablets for drug delivery throughout the gastrointestinal tract. Preformulation studies on theophylline anhydrous included solubility and dissolution rate determinations. Techniques including X-ray powder diffraction, differential scanning colorimetry and infrared spectroscopy provided no evidence of true polymorphism after recrystallisation from various solvents. However, scanning electron micrographs showed the effects of solvent polarity and cooling rate on the size and shape of recrystallised particles. Theophylline granules were manufactured by using various binders and were film coated by fluidised bed technology with various proportions of ethylcellulose, containing varying amounts of PEG 1540. In vitro release rates were dependent upon coating thickness and the proportion of PEG, which, being water soluble, created pores in the coating during dissolution studies as observed by a scanning electron microscope. However, substantial proportions of the drug remained unreleased from the granules. In order to overcome the problems of drug retention, plain granules were used and theophylline mini-tablets (3 mm diameter, weighing 15 - 20 mg) were manufactured and film coated with various Eudragits ® and other polymeric mixtures (soluble and insoluble). In vitro dissolution profiles from samples enclosed in hard gelatin capsules were determined using the USPXXI paddle apparatus in test media at pH 1.2 (HCI), pH 5.4 and 7.4 (phosphate buffers) at 37'C. Monitoring of in vitro theophylline release over 12 h, under identical hydrodynamic conditions, showed that the dissolution rate at pH 1.2 is substantially greater (95% of total drug content released in < 10 h) than that in phosphate buffers. The maximum release after 12 h was approximately 20 and 30% of total drug content of the tablet at pH 5.4 and 7.4, respectively. However, in vivo bioavailability after oral administration of tablets to rabbits corresponded to over 95% of total drug, compared with the same dose administered intravenously. The retarded drug release during in vitro dissolution in phosphate buffer was attributed to a possible interaction of phosphate ions with theophylline molecules at the tablet core-coat interface. These findings indicate that both rate and extent of theophylline release from the slow release coated mini-tablets are highly sensitive to phosphate buffers. The data also emphasise the usefulness of an animal model for assessment of in vivo drug release and subsequent absorption during the development of modified release dosage forms. Mini-tablets were subjected to isothermal and cyclic stresses to reach conditions for up to 6 months at different temperatures and relative humidity. The film integrity was maintained but ageing of the coating occurred which impeded dissolution. Reduced drug release was temperature related while the effect of relative humidi% was insignific~t. Encapsulated mini-tablets (uncoated and coated with Eudragit RL and RS 2% w/w) equivalent to a 300 mg dose, were evaluated both in vitro and in vivo using beagle dogs. The pharmacokinetic parameters from single and multiple dose studies showed several advantages over Theo-Dur® 300 mg tablets. Precise dosage titration is possible by careful adjustment of the number of encapsulated mini-tablets. This multiple unit mini-tablet delivery system will allow for greater flexibility in dosage adjustment compared to the currently available preparations, allowing individualised fine dose titration in those patients requiring therapeutic drug monitoring. The developmentof the multiple unit mini-tablet formulation appears to provide an optimal dosage form with maximum flexibility in respect of dose, duration range and ease of production.
- Full Text:
- Date Issued: 1991
Design, development and evaluation of encapsulated oral controlled release theophylline mini-tablets
- Authors: Munday, Dale Leslie
- Date: 1991
- Subjects: Drugs -- Administration , Drugs -- Bioavailability , Drugs -- Controlled release , Drugs -- Dosage forms , Tablets (Medicine) , Biopharmaceutics , Drugs -- Testing
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3777 , http://hdl.handle.net/10962/d1003255 , Drugs -- Administration , Drugs -- Bioavailability , Drugs -- Controlled release , Drugs -- Dosage forms , Tablets (Medicine) , Biopharmaceutics , Drugs -- Testing
- Description: Conventional solid dosage forms often lead to fluctuations which exceed the maximum safe therapeutic level and/or decline below the minimum effective level. It is recognised that many drugs for chronic administration should be administered on a schedule that maintains plasma drug concentration within the therapeutic window. Research in controlled release dosage forms aims at designing a system with a zero-order input (eg, ideally to deliver 8.33% of the dose per hour over a 12 hour duration), producing steady state plasma drug levels. Oral dministration of drugs prepared as a controlled release formulation is extremely popular, and has attracted the attention of pharmaceutical scientists during the last decade. This has been due to the simultaneous convergence of various factors (eg, discovery of novel polymers and devices, better understanding of formulation and physiological constraints, expiration of existing patents, prohibitive cost of developing new drug entities), involved in the development of these delivery systems. Controlled release oral products can be formulated as single or multiple unit dosage forms and the relative merits of multiple unit forms with their own rate controlling systems are well established. This work describes the development of a relatively inexpensive multiple-unit capsule dosage form of theophylline containing coated mini-tablets for drug delivery throughout the gastrointestinal tract. Preformulation studies on theophylline anhydrous included solubility and dissolution rate determinations. Techniques including X-ray powder diffraction, differential scanning colorimetry and infrared spectroscopy provided no evidence of true polymorphism after recrystallisation from various solvents. However, scanning electron micrographs showed the effects of solvent polarity and cooling rate on the size and shape of recrystallised particles. Theophylline granules were manufactured by using various binders and were film coated by fluidised bed technology with various proportions of ethylcellulose, containing varying amounts of PEG 1540. In vitro release rates were dependent upon coating thickness and the proportion of PEG, which, being water soluble, created pores in the coating during dissolution studies as observed by a scanning electron microscope. However, substantial proportions of the drug remained unreleased from the granules. In order to overcome the problems of drug retention, plain granules were used and theophylline mini-tablets (3 mm diameter, weighing 15 - 20 mg) were manufactured and film coated with various Eudragits ® and other polymeric mixtures (soluble and insoluble). In vitro dissolution profiles from samples enclosed in hard gelatin capsules were determined using the USPXXI paddle apparatus in test media at pH 1.2 (HCI), pH 5.4 and 7.4 (phosphate buffers) at 37'C. Monitoring of in vitro theophylline release over 12 h, under identical hydrodynamic conditions, showed that the dissolution rate at pH 1.2 is substantially greater (95% of total drug content released in < 10 h) than that in phosphate buffers. The maximum release after 12 h was approximately 20 and 30% of total drug content of the tablet at pH 5.4 and 7.4, respectively. However, in vivo bioavailability after oral administration of tablets to rabbits corresponded to over 95% of total drug, compared with the same dose administered intravenously. The retarded drug release during in vitro dissolution in phosphate buffer was attributed to a possible interaction of phosphate ions with theophylline molecules at the tablet core-coat interface. These findings indicate that both rate and extent of theophylline release from the slow release coated mini-tablets are highly sensitive to phosphate buffers. The data also emphasise the usefulness of an animal model for assessment of in vivo drug release and subsequent absorption during the development of modified release dosage forms. Mini-tablets were subjected to isothermal and cyclic stresses to reach conditions for up to 6 months at different temperatures and relative humidity. The film integrity was maintained but ageing of the coating occurred which impeded dissolution. Reduced drug release was temperature related while the effect of relative humidi% was insignific~t. Encapsulated mini-tablets (uncoated and coated with Eudragit RL and RS 2% w/w) equivalent to a 300 mg dose, were evaluated both in vitro and in vivo using beagle dogs. The pharmacokinetic parameters from single and multiple dose studies showed several advantages over Theo-Dur® 300 mg tablets. Precise dosage titration is possible by careful adjustment of the number of encapsulated mini-tablets. This multiple unit mini-tablet delivery system will allow for greater flexibility in dosage adjustment compared to the currently available preparations, allowing individualised fine dose titration in those patients requiring therapeutic drug monitoring. The developmentof the multiple unit mini-tablet formulation appears to provide an optimal dosage form with maximum flexibility in respect of dose, duration range and ease of production.
- Full Text:
- Date Issued: 1991
Development and assessment of ketoconazole intravaginal thermosetting hydrogel formulations
- Authors: Ramanah, Ashmita
- Date: 2017
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/5198 , vital:20785
- Description: Imidazole compounds are commonly used as antifungal therapies and ketoconazole was the first broad spectrum orally active azole identified and registered. However, the risks of hepatotoxicity and drug interactions following systemic delivery and absorption of ketoconazole outweigh the therapeutic benefits and ketoconazole was therefore discontinued as first line systemic antifungal therapy in many countries. Although not yet banned in South Africa, the South African Medicine Formulary has ceased to recommend the use of ketoconazole for systemic treatment. Topical use of ketoconazole is, however, regarded as safe following extensive human use as low systemic absorption occurs following topical administration. Vulvo-vaginal candidiasis is a yeast infection that affects a large number of women, some of whom present with several infections annually. The topical treatment options for vulvo-vaginal candidiasis include the use of vaginal tablets, capsules, ovules and creams administered as a single dose or one to three times daily for three to fourteen days either alone or in combination with another dosage form depending on the regimen. Administration of the dose nightly is recommended for most vaginal creams and ovule formulation due to leakage and the uncomfortable feel of the dosage form if administered during the day. A thermosetting gel that remains in the vagina following administration and prolongs the release of ketoconazole from a once daily dose would be a useful addition to the arsenal for intra-vaginal antifungal therapy. Thermosetting gels would be more comfortable to administer as the gel would set in a form similar to naturally occurring mucous in the vagina and, if formulated with a low pH, irritation of the sensitive and fissured tissue would be minimised. A further benefit would be that once set the gel would loosely take on the anatomical shape of the vagina. A simple, precise, accurate, reproducible and sensitive stability-indicating reversed phase-high performance liquid chromatographic method using ultraviolet detection for the quantitation of ketoconazole was developed and validated. The method was specific and was applied to the determination of ketoconazole in commercial and experimental formulations in addition to samples from degradation studies and in vitro release testing. Product performance characteristics of commercial products were investigated with the goal to provide a strategy for the development of a novel intra vaginal gel in the shortest possible time. Characterisation of Xolegel®, Kez® shampoo and Ketazol® cream included an evaluation of pH, viscosity and assay, in addition to spectroscopic and thermal analysis, to identify ideal characteristics of topical products that could be used as targets during formulation development of the gel. An in vitro release method was developed and validated for precision and accuracy and the in vitro release profiles of commercial ketoconazole products were compared using analysis of variance, model dependent and independent approaches. Ketoconazole release data from test gel manufactured during formulation development were investigated to obtain information about the relationship between formulation content and drug release. Poloxamers marketed as Pluronic® and Lutrol® are synthetic non-ionic tri-block copolymers that consist of hydrophobic propylene oxide and hydrophilic polyethylene oxide blocks, which in solution interact to exhibit thermo-reversible behaviour. In situ forming hydrogels consisting of poloxamers, more specifically poloxamer 407, are activated following a temperature stimulus and undergo a sol to gel transition. This approach was used to produce a thermosetting vaginal gel that would exhibit a long residence time in the vagina with an associated enhancement of therapeutic efficacy. Ketoconazole- excipient compatibility was investigated during preformulation studies using spectroscopic and thermal analysis to enable the selection of excipients best suited for the production of a novel dosage form prior to formulation development activities. No obvious interactions between ketoconazole and excipient were observed and ketoconazole was found in an amorphous form when in combination with polysorbate 80 and poloxamers. A two-level factorial design was used to produce solvent systems with different amounts of polysorbate 80, citric acid and ethanol to identify a vehicle in which ketoconazole exhibited optimum solubility and at a pH that would be least irritating to the vaginal mucosa with a low content of excipients. The optimised vehicle consisted of 4% m/v citric acid, 1.5% v/v polysorbate 80 and 9.5% v/v ethanol made up to 50 g with citrate-phosphate buffer adjusted to pH 5.0, resulted in a vehicle of pH of 3.5 in which 71.41 mg of ketoconazole was dissolved per mL. A Central Composite Design was used to evaluate compositions for the modulation of viscosity of the thermosetting dosage form such that it was a liquid at 22 °C that rapidly formed a stiff gel when heated to 37 °C (intra-vaginal temperature) using different amounts of the poloxamer grades 407, 188 and 237. Thermosetting gels containing 2% m/v ketoconazole were manufactured using specifications generated using the Central Composite Design and the viscosity at 22 °C and 37 °C, solution to gel transition time, potency and ketoconazole release at 24, 48 and 72 hours investigated. Contour and three-dimensional response surface plots and mathematical relationships with target ranges set for responses were identified and with the aid of Central Composite Design the optimisation of a desirable thermosetting gel was achieved. The optimised composition included 16% m/v poloxamer 407, 10% m/v poloxamer 188 and 6% m/v poloxamer 237 in the gel that was used as the basis for further optimisation studies. The low ketoconazole release for ketoconazole observed indicated that the poloxamers had formed a gel matrix that sustained the release of ketoconazole and would therefore ensure that once daily administration of the gel was possible. The sol-gel transition test may be used as a simple and cheap alternative to viscosity testing for thermosetting formulations when expensive viscometers and rheometers are unavailable and was successfully used for this purpose.Ketoconazole is photolabile and is prone to degradation in aqueous solutions. The hydrophobic core of micelles formed in these dosage forms are believed to shield ketoconazole molecules and improve stability in aqueous solutions and acidic gels. The thermosetting gel optimised for poloxamer content was subjected to a further Central Composite Design in which sodium metabisulphite content and vehicle pH were investigated. The length of storage was used as a numeric variable and storage condition as a categoric variable at two levels to monitor the stability of the gels. The formulations were investigated at sample times of 0, 1, 2, 4 and 8 weeks at 5 °C, 25 °C and 40 °C. The use of a Central Composite Design facilitated an understanding of the interactions between input variables and their impact on the responses analysed including ketoconazole content, release at 24, 48 and 72 hours, gel pH and viscosity at 22 °C and 37 °C. Design of Experiments may be used as a rapid cost effective tool for an overall assessment of the stability of novel topical dosage forms. However, a more thorough assessment of stability may be required for product registration. Ketoconazole was found to be unstable in the acidic thermosetting gels despite the addition of antioxidant. The gels in liquid form at 5 °C and 25 °C have a low number of micelles for ketoconazole incorporation and therefore additional optimisation studies would be required to enhance the shelf-life of this product.
- Full Text:
- Date Issued: 2017
- Authors: Ramanah, Ashmita
- Date: 2017
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/5198 , vital:20785
- Description: Imidazole compounds are commonly used as antifungal therapies and ketoconazole was the first broad spectrum orally active azole identified and registered. However, the risks of hepatotoxicity and drug interactions following systemic delivery and absorption of ketoconazole outweigh the therapeutic benefits and ketoconazole was therefore discontinued as first line systemic antifungal therapy in many countries. Although not yet banned in South Africa, the South African Medicine Formulary has ceased to recommend the use of ketoconazole for systemic treatment. Topical use of ketoconazole is, however, regarded as safe following extensive human use as low systemic absorption occurs following topical administration. Vulvo-vaginal candidiasis is a yeast infection that affects a large number of women, some of whom present with several infections annually. The topical treatment options for vulvo-vaginal candidiasis include the use of vaginal tablets, capsules, ovules and creams administered as a single dose or one to three times daily for three to fourteen days either alone or in combination with another dosage form depending on the regimen. Administration of the dose nightly is recommended for most vaginal creams and ovule formulation due to leakage and the uncomfortable feel of the dosage form if administered during the day. A thermosetting gel that remains in the vagina following administration and prolongs the release of ketoconazole from a once daily dose would be a useful addition to the arsenal for intra-vaginal antifungal therapy. Thermosetting gels would be more comfortable to administer as the gel would set in a form similar to naturally occurring mucous in the vagina and, if formulated with a low pH, irritation of the sensitive and fissured tissue would be minimised. A further benefit would be that once set the gel would loosely take on the anatomical shape of the vagina. A simple, precise, accurate, reproducible and sensitive stability-indicating reversed phase-high performance liquid chromatographic method using ultraviolet detection for the quantitation of ketoconazole was developed and validated. The method was specific and was applied to the determination of ketoconazole in commercial and experimental formulations in addition to samples from degradation studies and in vitro release testing. Product performance characteristics of commercial products were investigated with the goal to provide a strategy for the development of a novel intra vaginal gel in the shortest possible time. Characterisation of Xolegel®, Kez® shampoo and Ketazol® cream included an evaluation of pH, viscosity and assay, in addition to spectroscopic and thermal analysis, to identify ideal characteristics of topical products that could be used as targets during formulation development of the gel. An in vitro release method was developed and validated for precision and accuracy and the in vitro release profiles of commercial ketoconazole products were compared using analysis of variance, model dependent and independent approaches. Ketoconazole release data from test gel manufactured during formulation development were investigated to obtain information about the relationship between formulation content and drug release. Poloxamers marketed as Pluronic® and Lutrol® are synthetic non-ionic tri-block copolymers that consist of hydrophobic propylene oxide and hydrophilic polyethylene oxide blocks, which in solution interact to exhibit thermo-reversible behaviour. In situ forming hydrogels consisting of poloxamers, more specifically poloxamer 407, are activated following a temperature stimulus and undergo a sol to gel transition. This approach was used to produce a thermosetting vaginal gel that would exhibit a long residence time in the vagina with an associated enhancement of therapeutic efficacy. Ketoconazole- excipient compatibility was investigated during preformulation studies using spectroscopic and thermal analysis to enable the selection of excipients best suited for the production of a novel dosage form prior to formulation development activities. No obvious interactions between ketoconazole and excipient were observed and ketoconazole was found in an amorphous form when in combination with polysorbate 80 and poloxamers. A two-level factorial design was used to produce solvent systems with different amounts of polysorbate 80, citric acid and ethanol to identify a vehicle in which ketoconazole exhibited optimum solubility and at a pH that would be least irritating to the vaginal mucosa with a low content of excipients. The optimised vehicle consisted of 4% m/v citric acid, 1.5% v/v polysorbate 80 and 9.5% v/v ethanol made up to 50 g with citrate-phosphate buffer adjusted to pH 5.0, resulted in a vehicle of pH of 3.5 in which 71.41 mg of ketoconazole was dissolved per mL. A Central Composite Design was used to evaluate compositions for the modulation of viscosity of the thermosetting dosage form such that it was a liquid at 22 °C that rapidly formed a stiff gel when heated to 37 °C (intra-vaginal temperature) using different amounts of the poloxamer grades 407, 188 and 237. Thermosetting gels containing 2% m/v ketoconazole were manufactured using specifications generated using the Central Composite Design and the viscosity at 22 °C and 37 °C, solution to gel transition time, potency and ketoconazole release at 24, 48 and 72 hours investigated. Contour and three-dimensional response surface plots and mathematical relationships with target ranges set for responses were identified and with the aid of Central Composite Design the optimisation of a desirable thermosetting gel was achieved. The optimised composition included 16% m/v poloxamer 407, 10% m/v poloxamer 188 and 6% m/v poloxamer 237 in the gel that was used as the basis for further optimisation studies. The low ketoconazole release for ketoconazole observed indicated that the poloxamers had formed a gel matrix that sustained the release of ketoconazole and would therefore ensure that once daily administration of the gel was possible. The sol-gel transition test may be used as a simple and cheap alternative to viscosity testing for thermosetting formulations when expensive viscometers and rheometers are unavailable and was successfully used for this purpose.Ketoconazole is photolabile and is prone to degradation in aqueous solutions. The hydrophobic core of micelles formed in these dosage forms are believed to shield ketoconazole molecules and improve stability in aqueous solutions and acidic gels. The thermosetting gel optimised for poloxamer content was subjected to a further Central Composite Design in which sodium metabisulphite content and vehicle pH were investigated. The length of storage was used as a numeric variable and storage condition as a categoric variable at two levels to monitor the stability of the gels. The formulations were investigated at sample times of 0, 1, 2, 4 and 8 weeks at 5 °C, 25 °C and 40 °C. The use of a Central Composite Design facilitated an understanding of the interactions between input variables and their impact on the responses analysed including ketoconazole content, release at 24, 48 and 72 hours, gel pH and viscosity at 22 °C and 37 °C. Design of Experiments may be used as a rapid cost effective tool for an overall assessment of the stability of novel topical dosage forms. However, a more thorough assessment of stability may be required for product registration. Ketoconazole was found to be unstable in the acidic thermosetting gels despite the addition of antioxidant. The gels in liquid form at 5 °C and 25 °C have a low number of micelles for ketoconazole incorporation and therefore additional optimisation studies would be required to enhance the shelf-life of this product.
- Full Text:
- Date Issued: 2017