Ozone, being an unstable molecule, is believed to be one of the strongest oxidant agents known to man. Rapid growth in the application of ozone — both as disinfectant and as form of alternative medicine — led to questions about the effects of uncontrolled ozone exposure and inhalation, whether intentional or unintentional, on the human body. This study specifically focussed on examining, identifying and substantiating the respiratory effect of acute exposure (10 min or less) to considerably higher ozone concentrations than reported on before (19.5 ± 0.5 ppm). Respiratory tissue of rodents (Duncan–Hartley guinea pigs of both sexes and Male Wistar rats) was subjected to ozone by utilising three distinctly diverse models of ozone introduction: (a) in vitro exposure, (b) in vivo exposure, and (c) ex vivo by employing an isolated lung perfusion model which allows for real–time, breath–by–breath data acquisition of ozone’s effect on respiratory mechanics. The effect of ozone on the isolated trachea in the presence of various drugs with well–known effects, including methacholine, isoproterenol and ascorbic acid was also examined. The results found in this study identified two direct effects on the isolated trachea due to ozone exposure: (1) a definite contraction of the isolated trachea immediately after exposure to ozone, and (2) a clearly visible and significant hyper responsiveness of the isolated trachea to irritants, e.g. methacholine. Although ozone has a negative effect on the trachea, it was concluded that ozone has no adverse effect on muscarinic acetylcholine receptors. An apparent EC50 value of ozone on the trachea was established by two different methods as (2.77 ± 0.02) x 10–3 M and (2.10 ± 0.03) x 10–3 M, respectively. Ozone furthermore displayed an attenuation of the beneficial pharmacological response of –sympathomimetic drugs (i.e. isoproterenol), while isoproterenol itself has a relaxing effect on the ozone–induced contraction of the isolated trachea. Indomethacin pre–treatment of isolated tracheal tissue significantly (77%) reduced the ozone–induced contraction of tracheal smooth muscle, suggesting that COXproducts of arachidonic acid play a prominent role in the development of pulmonary function decrements consequent to acute high–dose ozone exposure. Ascorbic acid exhibited a meaningful prophylactic effect on ozone–induced contraction of both isolated tracheal tissue and in the isolated lung perfusion model, emphasising the major role antioxidants play in both the epithelium lining fluid (ELF) of the respiratory system and in plasma throughout the body in protecting against the destructive effects of ozone. Surprisingly, pre–treatment with ascorbic acid did not prevent hyper responsiveness of isolated tracheal preparations to methacholine after a 10 min ozone (19.5 ± 0.5 ppm) exposure. In the lung perfusion model, the presence of ascorbic acid in the perfusion medium did, however, significantly reduce the magnitude and rate of decline in lung compliance after ozone exposure (46% decline with ascorbic acid versus 96% in the control study without ascorbic acid). Examination of a lung perfusion model exposed to ozone (19.5 ± 0.5 ppm O3; 5 seconds) presented a significant decline in lung compliance (95.6% within 2 min), tidal volume (70%) and maximum inspiratory flow (71.2%), with an ensuing reduction in lung elasticity and severely hampered breathing pattern. Microscopic examination after acute high–dose inhalation studies did not display any significant cellular damage, oedema or inflammation after acute high–dose ozone exposure. This suggests that significant cellular injury and inflammation is possibly not the causative factor of early breathing difficulty experienced after acute high–dose ozone inhalation, as these symptoms and particularly the result of inflammatory precursors, is believed to probably only set in at a later stage. Although the potential advantages of ozone in certain fields of medicine are not disputed, ozone, depending on its concentration and cumulative dose, can be either therapeutic or toxic. Observations in this study emphasised that even short bursts of high–dose ozone inhalation have deleterious effects on respiratory health and care should be taken not to jump to conclusions regarding ozone’s medical application without relevant scientific evidence. It must be stressed that high–dose inhalation of ozone should be avoided at all cost – especially by those with existing airway diseases. / Thesis (Ph.D. (Pharmacology))--North-West University, Potchefstroom Campus, 2011.

Ozone, being an unstable molecule, is believed to be one of the strongest oxidant agents known to man. Rapid growth in the application of ozone — both as disinfectant and as form of alternative medicine — led to questions about the effects of uncontrolled ozone exposure and inhalation, whether intentional or unintentional, on the human body. This study specifically focussed on examining, identifying and substantiating the respiratory effect of acute exposure (10 min or less) to considerably higher ozone concentrations than reported on before (19.5 ± 0.5 ppm). Respiratory tissue of rodents (Duncan–Hartley guinea pigs of both sexes and Male Wistar rats) was subjected to ozone by utilising three distinctly diverse models of ozone introduction: (a) in vitro exposure, (b) in vivo exposure, and (c) ex vivo by employing an isolated lung perfusion model which allows for real–time, breath–by–breath data acquisition of ozone’s effect on respiratory mechanics. The effect of ozone on the isolated trachea in the presence of various drugs with well–known effects, including methacholine, isoproterenol and ascorbic acid was also examined. The results found in this study identified two direct effects on the isolated trachea due to ozone exposure: (1) a definite contraction of the isolated trachea immediately after exposure to ozone, and (2) a clearly visible and significant hyper responsiveness of the isolated trachea to irritants, e.g. methacholine. Although ozone has a negative effect on the trachea, it was concluded that ozone has no adverse effect on muscarinic acetylcholine receptors. An apparent EC50 value of ozone on the trachea was established by two different methods as (2.77 ± 0.02) x 10–3 M and (2.10 ± 0.03) x 10–3 M, respectively. Ozone furthermore displayed an attenuation of the beneficial pharmacological response of –sympathomimetic drugs (i.e. isoproterenol), while isoproterenol itself has a relaxing effect on the ozone–induced contraction of the isolated trachea. Indomethacin pre–treatment of isolated tracheal tissue significantly (77%) reduced the ozone–induced contraction of tracheal smooth muscle, suggesting that COXproducts of arachidonic acid play a prominent role in the development of pulmonary function decrements consequent to acute high–dose ozone exposure. Ascorbic acid exhibited a meaningful prophylactic effect on ozone–induced contraction of both isolated tracheal tissue and in the isolated lung perfusion model, emphasising the major role antioxidants play in both the epithelium lining fluid (ELF) of the respiratory system and in plasma throughout the body in protecting against the destructive effects of ozone. Surprisingly, pre–treatment with ascorbic acid did not prevent hyper responsiveness of isolated tracheal preparations to methacholine after a 10 min ozone (19.5 ± 0.5 ppm) exposure. In the lung perfusion model, the presence of ascorbic acid in the perfusion medium did, however, significantly reduce the magnitude and rate of decline in lung compliance after ozone exposure (46% decline with ascorbic acid versus 96% in the control study without ascorbic acid). Examination of a lung perfusion model exposed to ozone (19.5 ± 0.5 ppm O3; 5 seconds) presented a significant decline in lung compliance (95.6% within 2 min), tidal volume (70%) and maximum inspiratory flow (71.2%), with an ensuing reduction in lung elasticity and severely hampered breathing pattern. Microscopic examination after acute high–dose inhalation studies did not display any significant cellular damage, oedema or inflammation after acute high–dose ozone exposure. This suggests that significant cellular injury and inflammation is possibly not the causative factor of early breathing difficulty experienced after acute high–dose ozone inhalation, as these symptoms and particularly the result of inflammatory precursors, is believed to probably only set in at a later stage. Although the potential advantages of ozone in certain fields of medicine are not disputed, ozone, depending on its concentration and cumulative dose, can be either therapeutic or toxic. Observations in this study emphasised that even short bursts of high–dose ozone inhalation have deleterious effects on respiratory health and care should be taken not to jump to conclusions regarding ozone’s medical application without relevant scientific evidence. It must be stressed that high–dose inhalation of ozone should be avoided at all cost – especially by those with existing airway diseases. / Thesis (Ph.D. (Pharmacology))--North-West University, Potchefstroom Campus, 2011.