The development, manufacture and evaluation of sustained release gastric-resistant isoniazid and gastroretentive microporous rifampicin microspheres
- Authors: Mwila, Chiluba
- Date: 2018
- Subjects: Biodegradation , Microspheres (Pharmacy) , Drug delivery systems , Rifampin , Isoniazid
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/63497 , vital:28421 , DOI 10.21504/10962/63497
- Description: According to the World Health Organization Global Tuberculosis (TB) 2017 Report, there were an estimated 10.4 million new TB cases worldwide of which, in 2016, 65 % occurred in men, 28.1 % in women and 6.9 % in children. TB is the ninth leading cause of death globally and is the leading cause due to an infectious organism surpassing HIV/AIDS. Treatment is long-term and the use of a combination of medicines is required for success. The concern related to the use of fixed dose combination products for the treatment of TB is the issue of low bioavailability of rifampicin observed from a number of fixed dose combination (FDC) formulations. The hydrolysis of rifampicin, in acidic media, to form insoluble 3-formyl rifamycin SV contributes to poor bioavailability of rifampicin. The degradation of rifampicin to form this poorly absorbed compound is accelerated in the presence of isoniazid via the reversible formation of isonicotinyl hydrazone is a further factor contributing to the poor bioavailability of rifampicin. Therefore, the development of a novel drug delivery technology that prevents interactions between rifampicin and isoniazid in an acidic medium is required. A Box Behnken design was successfully used for the optimisation of a rapid and accurate stability-indicating gradient elution RP-HPLC method for the simultaneous analysis of isoniazid, pyrazinamide and rifampicin. The method was validated using ICH guidelines and the results indicate it can be used for the rapid analysis of commercially available TB FDC formulations containing the active pharmaceutical ingredients, API. The method is precise, sensitive and has the necessary selectivity for use during formulation development and optimisation studies for a combination of rifampicin, isoniazid and pyrazinamide. Initially formulation activities were undertaken with rifampicin and isoniazid for the development of an approach to enhance the effective delivery of these compounds. The characterisation of rifampicin and isoniazid was undertaken using spectroscopic, thermal and microscopic analysis. The studies revealed that the compounds are crystalline and exhibit distinct characteristic sharp peaks in X-ray diffractograms and Differential Scanning Calorimetry thermograms. The thermograms, 13C Nuclear Magnetic Resonance and Fourier Transform Infrared spectroscopy results identified that rifampicin occurs as the form II polymorph however, as there are no significant biopharmaceutic differences between the polymorphic forms of rifampicin this information was used for identification purposes only. The results were used as baseline data for comparative purposes to monitor changes that may occur when rifampicin and isoniazid are used in formulation development, dosage form manufacture and characterisation activities for a FDC technology designed to deliver both compounds simultaneously. Hydroxypropylmethylcellulose acetate succinate (HPMC-AS) and Eudragit® L100 polymers were successfully used for manufacture of isoniazid loaded gastric-resistant sustained release microspheres using an o/o solvent emulsification and evaporation approach. A Hybrid experimental design was used to investigate the influence of input variables viz., homogenisation speed and amount of HPMC-AS and Eudragit® L100 on gastric-resistance, INH release and encapsulation efficiency. The approach of using coating polymers viz., HPMC-AS and Eudragit® L100, to manufacture gastric resistant sustained release microspheres of isoniazid is unique and was efficient for preventing the release of isoniazid in an acidic environment. Only 0.523 % isoniazid was released from the optimised formulation after 2 h exposure to pH 1.2 0.1 M HCl suggesting there is also the possibility of minimising the accelerated degradation of rifampicin that occurs in the presence of isoniazid in acidic media. The microspheres also exhibited sustained release properties without burst release in pH 6.8 0.1 M phosphate buffer as < 5 % isoniazid was released at 0.5 h and only 11 % isoniazid was released at 2 h. The release of isoniazid was sustained over the entire period of dissolution testing with > 85 % isoniazid released at 24 h, implying that the majority of encapsulated isoniazid would be available for absorption. The manufacturing process resulted in the production of hard spherical particles and particle size analysis revealed that the microspheres ranged between 415.76 ± 76.93 μm and 903.35 ± 197.10 μm in diameter. The microspheres exhibited excellent flow properties attributed to the spherical nature of particles. Carr‟s index (CI) was 4.934 ± 0.775 % and the Hausner ratio (HR) was 1.148 ± 0.033 indicating good packability of the microspheres that would help in achieving weight and content uniformity of capsule dosage units. The manufacturing process however produced a low % yield suggesting that scale up difficulties may be encountered. However the high encapsulation efficiency observed may counter the challenges associated with the low yield. The DSC thermograms and FT Raman spectra of 1:1 mixtures of isoniazid, excipients and the microspheres did not reveal any potential detrimental interactions. Microporous floating sustained release microspheres for the delivery of rifampicin in the stomach have been successfully manufactured using emulsification and a diffusion/evaporation process. A novel approach using solvent mixture of acetone and dichloromethane that has not been reported for the manufacture of rifampicin microspheres was successfully used and resulted in the formation of a stable emulsion and the manufacture of rifampicin-loaded microspheres with uniform characteristics. In addition the manufacturing process was shorter than most other reported methods. A Box-Behnken experimental design was successfully used to study the influence of ethylcellulose, Eudragit® RLPO and d-glucose content on the floating properties, encapsulation efficiency and % yield of microspheres. The optimised formulation did not yield desired floating characteristics as the % buoyancy was low and floating lag times were high. The optimised formulation was modified by addition of NaHCO3 to increase the % buoyancy and reduce the floating lag time. Rifampicin release from the microspheres of the modified batch was 87.10 % at 12 h and the microspheres exhibited a % buoyancy of 87.66 ± 1.28 % (n = 6) and floating lag time of 15 ± 3.2 (n = 6) seconds. The microspheres remained buoyant for up to 12 h and an encapsulation efficiency of 88.26 ± 1.25 % was achieved. SEM images of microspheres following exposure to dissolution fluid revealed that the microspheres had numerous pores on their surface. The mean particle size distribution ranged between 423.19 ± 121.86 μm to 620.07 ± 102.67 μm. The microspheres exhibited similar flow characteristics to isoniazid microspheres with a CI of 1.422 ± 0.074 %, and HR of 1.034 ± 0.002. The excellent flow characteristics indicate that filling of the microspheres into hard gelatin capsules was unlikely to pose a challenge in respect of producing a product with uniform content. Rifampicin-excipient compatibility studies did not reveal any potential or significant interactions suggesting that the excipients used for the manufacture of the microspheres were compatible, although long term stability studies would be required to ascertain this is, indeed the case. The microporous floating sustained release microspheres manufactured in these studies has the potential to increase the bioavailability of rifampicin as they may be retained in the stomach where the solubility of rifampicin is high and from which absorption is best achieved. The degradation of rifampicin after 12 h dissolution testing in pH 1.2 0.1 M HCl in the presence of isoniazid gastric-resistant sustained release microspheres was only 4.44%. These results indicate that the degradation of rifampicin in the presence of isoniazid in acidic media can be overcome by encapsulation of both active pharmaceutical ingredients in a manner that ensure release in different segments of the gastrointestinal tract. The use of sustained release microporous gastroretentive rifampicin microspheres in combination with sustained release isoniazid gastric-resistant microspheres revealed that accelerated degradation of rifampicin in the presence of isoniazid is reduced significantly when using this approach and a FDC of rifampicin and isoniazid microspheres has the potential to improve the bioavailability of rifampicin thereby enhancing therapeutic outcomes. In vivo studies would be required to confirm the potential benefits of using this approach to deliver rifampicin in combination with isoniazid. , Thesis (PhD) -- Faculty of Pharmacy, Pharmacy, 2018
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- Date Issued: 2018
Optimization of the vermidegradation of cow dung – waste paper mixtures
- Authors: Unuofin, Frank Oshioname
- Date: 2014
- Subjects: Biodegradation , Vermicomposting , Waste paper , Earthworm culture , Phosphatic fertilizers
- Language: English
- Type: Thesis , Doctoral , PhD (Soil Science)
- Identifier: vital:11966 , http://hdl.handle.net/10353/d1021276 , Biodegradation , Vermicomposting , Waste paper , Earthworm culture , Phosphatic fertilizers
- Description: Vermicomposting is an eco-friendly waste management strategy. Its successful performance necessitate that key functioning parameters like earthworm stocking density, nutrient enrichment be established for each target waste/waste mixture. One main target waste mixture in South Africa, and in the University of Fort Hare in particular is waste paper mixed with cow dung and rock phosphate (RP) for phosphorus (P) enrichment. This study was carried out to address the following specific objectives, to determine (i) the effect of Eisenia fetida stocking density on the bioconversion of cow dung waste paper mixtures enriched with rock phosphate, (ii) an optimum application rate of low grade South African Rock Phosphate and time required for efficient vermicomposting of cow dung-waste paper mixtures, and (iii) to determine if the phosphorus in RP is responsible for improved biodegradation during the vermicomposting of cow dung-waste paper mixtures. Results of this study revealed that bioconversion of cow dung waste paper mixtures enriched with RP was highly dependent on E. fetida stocking density and time. The stocking density of 12.5 g-worms kg-1 feedstock of the mixtures resulted in highest earthworm growth rate and humification of the waste mixture as reflected by a C: N ratio of < 12, polymerization index (PI) or humic acid/fulvic acid ratio of > 1.9, and a humification index of >13 for the cow dung waste paper mixtures. A germination test carried out also revealed that the resultant vermicompost had no inhibitory effect on the germination of tomato, carrot, and radish. Extractable P increased with stocking density up to 22.5 g-worm kg-1 feedstock, suggesting that for maximum P release from RP enriched wastes, a high stocking density should be considered. Informed by an earlier study which demonstrated that RP improved vermidegration, a follow up study was done to determine the optimum amount of rock phosphate necessary for efficient vermidegradation of cow dung waste paper mixtures while ensuring a phosphorus rich vermicompost. The results showed that addition of RP at rates ≤ 1% P as RP efficiently enhanced the bioconversion of cow dung waste paper mixtures as reflected by low C: N ratio, high polymerization index (PI), HI and HR used as maturity indicators for matured compost. Final vermicompost products obtained at minimum amounts of RP application rates resulted highly humified vermicompost with finer morphological structure, with no inhibitory effect on the germination of tomato, carrot, and radish similar to the ones obtained at higher RP rates. The findings suggest that 1%P as RP application rate is optimum for efficient vermidegradation of cow dung waste paper mixtures. Since P or Ca happen to be the most prevalent elements in most rock phosphate used for compost enrichment, a study was carried out to determine if P or Ca in RP is predominantly responsible for the improved biodegradation of cow-dung waste paper mixture observed during vermicomposting. Phosphorus sources in form of triple superphosphate (TSP), phosphoric acid (PHA) and Ca in form of calcium chloride (CaCl2) salt were compared with rock phosphate. The results from the study indicated that TSP, a water soluble P source, resulted in greater and faster degradation of the waste mixtures than RP while the Ca source had the least effect. With TSP incorporation the compost maturity C: N ratio of 12 was reached within 28 days while RP, PHA and CaCl2 needed 42, 56 and more than 56 days, respectively. The results indicated that P was largely responsible for the enhanced bioconversion of the waste mixtures. This appeared linked to the effect of P to stimulate microbial growth as reflected by higher microbial biomass carbon levels where water soluble P sources were applied. The C: N ratios of the final vermicomposts at day 56 were 10, 11.5, 13, 14, and 23 for TSP, RP,PHA, Control (No P added) and CaCl2 treatments, respectively. Although TSP gave superior superior performance, RP may still be the preferred additive in the vermicomposting of cow dung waste paper mixtures as it is cheaper and produces mature compost in a shorter period of 8 weeks. Generally, the results of this study have shown that the vemidegradation of cow dung waste paper mixtures can be optimized through adoption of an E. fetida stocking density of 12.5g- worm kg-1 and an RP incorporation rate of 1% P as RP. However, higher rates of RPincorporation may be adopted where final vermicomposts with higher P fertilizer value are desired. Phosphorus appears to be the RP constituent responsible for its ability to enhance the vermidegradation of cow dung waste paper mixtures. Future studies should explore the effectiveness of other P-bearing minerals for their effectiveness in enhancing vemidegradation.
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- Date Issued: 2014
Improving sanitization and fertiliser value of dairy manure and waste paper mixtures enriched with rock phosphate through combined thermophilic composting and vermicomposting
- Authors: Mupondi, Lushian Tapiwa
- Date: 2010
- Subjects: Compost , Vermicomposting , Manures , Biodegradation , Waste paper -- South Africa -- Eastern Cape , Thermophilic bacteria , Fertilizers
- Language: English
- Type: Thesis , Doctoral , PhD (Soil Science)
- Identifier: vital:11961 , http://hdl.handle.net/10353/411 , Compost , Vermicomposting , Manures , Biodegradation , Waste paper -- South Africa -- Eastern Cape , Thermophilic bacteria , Fertilizers
- Description: Thermophilic composting (TC) and vermicomposting (V) are the two most common methods used for biological stabilization of solid organic wastes. Both have their advantages and disadvantages but the proposed method of combining composting and vermicomposting (CV) borrows pertinent attributes from each of the two methods and combines them to enhance overall process and product qualities. Dairy manure and waste paper are two wastes produced in large quantities at the University of Fort Hare. The study was carried out to address the following specific objectives, to determine (i) the effectiveness of combined thermophilic composting and vermicomposting on the biodegradation and sanitization of mixtures of dairy manure and paper waste, (ii) an optimum precomposting period for dairy manure paper waste mixtures that results in vermicomposts of good nutritional quality and whose use will not jeopardize human health, (iii) the effectiveness of phosphate rock (PR) in increasing available P and degradation and nutrient content of dairy manure-paper vermicomposts, (iv) the physicochemical properties of vermicompost substituted pine bark compost and performance of resultant growing medium on plant growth and nutrient uptake. Results of this study revealed that wastes with a C: N ratio of 30 were more suitable for both V and CV as their composts were more stabilized and with higher nutrient contents than composts made from wastes with a C: N ratio of 45. Both V and CV were effective methods for the biodegradation of dairy manure and paper waste mixtures with C: N ratio of 30 but the latter was more effective in the biodegradation of waste mixtures with a C: N ratio of 45. The combinination of composting and vermicomposting eliminated the indicator pathogen E. coli 0157 from the final composts whereas V only managed to reduce the pathogen population. iv A follow up study was done to determine the effects of precomposting on pathogen numbers so as to come up with a suitable precomposting period to use when combine composting dairy manure-waste paper mixtures. Results of this study showed that over 95% of fecal coliforms, E. coli and of E. coli 0157 were eliminated from the wastes within one week of precomposting and total elimination of these and protozoan (oo)cysts achieved after 3 weeks of precomposting. The vermicomposts pathogen content was related to the waste’s precomposting period. Final vermicomposts pathogen content was reduced and varied according to precomposting period. Vermicomposts from wastes precomposted for over two weeks were less stabilized, less humified and had less nutrient contents compared to vermicomposts from wastes that were precomposted for one week or less. The findings suggest that a precomposting period of one week is ideal for the effective vermicomposting of dairy manure-waste paper mixtures. Results of the P enrichment study indicated an increase in the inorganic phosphate and a reduction in the organic phosphate fractions of dairy manure-waste paper vermicompost that were enriched with PR. This implied an increase in mineralization of organic matter and or solubilization of PR with vermicomposting time. Applying PR to dairy manure-waste paper mixtures also enhanced degradation and had increased N and P contents of dairy manure-waste paper vermicomposts. Earthworms accumulated heavy metals in their bodies and reduced heavy metal contents of vermicomposts. A study to determine the physicochemical properties of vermicompost substituted pine bark compost and performance of resultant growing medium on plant growth and nutrient uptake was done. Results obtained revealed that increasing proportions of dairy manure vermicomposts in pine bark compost improved tomato plant height, stem girth, shoot and root dry weights. v Tomatoes grew best in the 40 to 60% CV substituted pine bark and application of Horticote (7:2:1 (22)) fertilizer significantly increased plant growth in all media. Progressive substitution pine bark with dairy manure vermicomposts resulted in a decrease in the percentage total porosity, percentage air space whilst bulk density, water holding capacity, particle density, pH, electrical conductivity and N and P levels increased. Precomposting wastes not only reduced and or eliminated pathogens but also improved the stabilisation and nutrient content of dairy manure waste paper mixtures. The application of PR to dairy manure waste paper mixtures improved the chemical and physical properties of vermicomposts. Earthworms bio-accumulated the heavy metals Cd, Cr, Cu, Pb and Zn whilst the contents of these in the vermicomposts declined. It is, therefore, recommended that dairy manure waste paper mixtures be precomposted for one week for sanitization followed by PR application and vermicomposting for stabilization and improved nutrients contents of resultant vermicomposts. Substitution of pine bark compost with 40 to 60 % PR-enriched vermicompost produced a growing medium with superior physical and chemical properties which supported good seedling growth. However, for optimum seedling growth, supplementation with mineral fertilizer was found to be necessary.
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- Date Issued: 2010
Voltammetric analysis of pesticides and their degradation: A case study of Amitraz and its degradants
- Authors: Brimecombe, Rory Dennis
- Date: 2006
- Subjects: Hydrolysis , Biodegradation , Voltammetry , Pesticides -- Biodegradation , Pesticides -- Environmental aspects , Acaricides , Acaricides -- Physiological effect
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4131 , http://hdl.handle.net/10962/d1015724
- Description: Amitraz is a formamide acaricide used predominantly in the control of ectoparasites in livestock and honeybees. Amitraz hydrolysis is rapid and occurs under acidic conditions, exposure to sunlight and biodegradation by microorganisms. The main hydrolysis product of amitraz, 2,4-dimethylaniline, is recalcitrant in the environment and toxic to humans. An electrochemical method for the determination of total amitraz residues and its final breakdown product, 2,4-dimethylaniline, in spent cattle dip, is presented. Cyclic voltammetry at a glassy carbon electrode showed the irreversible oxidation of amitraz and 2,4-dimethylaniline. A limit of detection in the range of 8.5 x 10⁻⁸ M for amitraz and 2 x 10⁻⁸ M for 2,4-dimethylaniline was determined using differential pulse voltammetry. Feasibility studies in which the effect of supporting electrolyte type and pH had on electroanalysis of amitraz and its degradants, showed that pH affects current response as well as the potential at which amitraz and its degradants are oxidised. Britton-Robinson buffer was found to be the most suitable supporting electrolyte for detection of amitraz and its degradants in terms of sensitivity and reproducibility. Studies performed using environmental samples showed that the sensitivity and reproducibility of amitraz and 2,4-dimethylaniline analyses in spent cattle dip were comparable to analyses of amitraz and 2,4-dimethylaniline performed in Britton-Robinson buffer. In addition, the feasibility qf measuring amitraz and 2,4-dimethylaniline in environmental samples was assessed and compared to amitraz and 2,4-dimethylaniline analyses in Britton-Robinson buffer. Amitraz and 2,4-dimethylaniline were readily detectable in milk and honey. Furthermore, it was elucidated that 2,4-dimethylaniline can be metabolised to 3-methylcatechol by Pseudomonas species and the proposed breakdown pathway is presented. The biological degradation of amitraz and subsequent formation of 2,4-dimethylaniline was readily monitored in spent cattle dip. The breakdown of amitraz to 2,4-dimethylaniline and then to 3-MC was monitored using cyclic voltammetry.
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- Date Issued: 2006
Bacterial degradation of the acaricide amitraz
- Authors: Baker, Penelope Bridget
- Date: 1976
- Subjects: Acaricides , Biodegradation , Gas chromatography , Bacteriology -- Cultures and culture media
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4099 , http://hdl.handle.net/10962/d1009498
- Description: This thesis describes dip tank field trials and laboratory investigations on the acaricide Amitraz. Amitraz is a triazapenta- diene compound which is relatively unstable in fouled dip washes. The field trials were conducted on the farm Sea View according to the "Total Replacement Method" and on the farm Sea Ways according to the "Lime Stabilization Method" of dipping. The results of these trials showed that Amitraz was stable in clean dip washes, and under conditions of high pH resulting from the addition of slaked lime to the dip wash. Using mixed bacterial populations optimum conditions for degradation of Amitraz in the laboratory were determined. Bacterial cultures degraded Amitraz most efficiently in media supplemented with yeast extract or with a high content of sterile cattle faeces. Amitraz concentrations were determined by gas chromatography. A culture. efficient at degrading Amitraz was enriched from a dip tank sludge inoculum. From this culture ten bacterial isolates were identified; nine of these were of the genus Pseudomonas and one was an Achromobacter sp. Experiments with both mixed and pure cultures demonstrated that bacterial degradation of Amitraz was by the process of co-metabolism. The existence of four degradation products was shown using thin layer chromatography. Tentative identification of two of the products was made.
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- Date Issued: 1976