The shortcomings of current antidepressant agents prompts the design of novel multimodal antidepressants and the identification of new antidepressant targets, especially those located at sub-cellular level. Such antidepressants should possess improved response rates as well as safety profiles. Methylene blue (MB) is reported to possess diverse pharmacological actions and is attracting increasing attention for the treatment of a variety of disorders including Alzheimer’s disease, bipolar disorder, anxiety and depression. MB acts on both monoamine oxidase (MAO) and the nitric oxide (NO)-cGMP pathway, and possesses antidepressant activity in rodents. The principal goal of this study was to design a close structural analogue of MB and to evaluate the effects of these structural changes on MAO inhibition, a well-known antidepressant target. Furthermore, MAO inhibition is also responsible for cardiovascular toxicity in clinically used MAOI inhibitors. For this purpose we investigated the antidepressant properties of the synthetic MB analogue (ethyl-thioniniumchloride; ETC) as well as azure B, the major metabolite of MB, in the forced swim test (FST). ETC was synthesized with a high degree of purity from diethyl-p-phenylenediamine with 6% yield. ETC was firstly evaluated as a potential inhibitor of recombinant human MAO-A and MAO-B. Azure B and ETC were evaluated over a dosage range of 4-30 mg/kg for antidepressant-like activity in the acute FST in rats, and the results were compared to those obtained with saline, imipramine (15 mg/kg) and MB (15 mg/kg) treated rats. Locomotor activity was evaluated to ensure that changes in swim motivation are based on antidepressant response and not due to an indirect effect of the drug on locomotor activity. The results document that ETC inhibits MAO-A and MAO-B with IC50 values of 0.51 μM and 0.592 μM, respectively. Furthermore, ETC inhibits MAO-A and MAO-B reversibly, while the mode of inhibition is most likely competitive. In the acute FST, azure B and ETC were more effective than imipramine and MB in reversing immobility, without inducing locomotor effects. Azure B and ETC increased swimming behaviour during acute treatment, which is indicative of enhanced serotonergic neurotransmission. Azure B and ETC did not affect noradrenergicmediated climbing behaviour. These results suggest that azure B may be a contributor to the antidepressant effect of MB, and acts via increasing serotonergic transmission. Secondly, small structural changes made to MB do not abolish its antidepressant effect even though ETC is a less potent MAO-A inhibitor than MB. / MSc (Pharmaceutical Chemistry), North-West University, Potchefstroom Campus, 2014

The shortcomings of current antidepressant agents prompts the design of novel multimodal antidepressants and the identification of new antidepressant targets, especially those located at sub-cellular level. Such antidepressants should possess improved response rates as well as safety profiles. Methylene blue (MB) is reported to possess diverse pharmacological actions and is attracting increasing attention for the treatment of a variety of disorders including Alzheimer’s disease, bipolar disorder, anxiety and depression. MB acts on both monoamine oxidase (MAO) and the nitric oxide (NO)-cGMP pathway, and possesses antidepressant activity in rodents. The principal goal of this study was to design a close structural analogue of MB and to evaluate the effects of these structural changes on MAO inhibition, a well-known antidepressant target. Furthermore, MAO inhibition is also responsible for cardiovascular toxicity in clinically used MAOI inhibitors. For this purpose we investigated the antidepressant properties of the synthetic MB analogue (ethyl-thioniniumchloride; ETC) as well as azure B, the major metabolite of MB, in the forced swim test (FST). ETC was synthesized with a high degree of purity from diethyl-p-phenylenediamine with 6% yield. ETC was firstly evaluated as a potential inhibitor of recombinant human MAO-A and MAO-B. Azure B and ETC were evaluated over a dosage range of 4-30 mg/kg for antidepressant-like activity in the acute FST in rats, and the results were compared to those obtained with saline, imipramine (15 mg/kg) and MB (15 mg/kg) treated rats. Locomotor activity was evaluated to ensure that changes in swim motivation are based on antidepressant response and not due to an indirect effect of the drug on locomotor activity. The results document that ETC inhibits MAO-A and MAO-B with IC50 values of 0.51 μM and 0.592 μM, respectively. Furthermore, ETC inhibits MAO-A and MAO-B reversibly, while the mode of inhibition is most likely competitive. In the acute FST, azure B and ETC were more effective than imipramine and MB in reversing immobility, without inducing locomotor effects. Azure B and ETC increased swimming behaviour during acute treatment, which is indicative of enhanced serotonergic neurotransmission. Azure B and ETC did not affect noradrenergicmediated climbing behaviour. These results suggest that azure B may be a contributor to the antidepressant effect of MB, and acts via increasing serotonergic transmission. Secondly, small structural changes made to MB do not abolish its antidepressant effect even though ETC is a less potent MAO-A inhibitor than MB. / MSc (Pharmaceutical Chemistry), North-West University, Potchefstroom Campus, 2014

The thesis is focused on a determination of heparin and chondroitin sulfate, using flow injection analysis with spectrofluorimetric detection. The determination is based on the interaction of negatively charged heparin, chondroitin sulfate resp., with a cationic dye (azure B or phenosafranine) which is manifested by the decrease in fluorescence intensity of the dye in its emission maximum. The optimal conditions for the determination in static mode were found, and calibration dependencies were measured. The conditions of FIA were optimized and following parameters were established: the volume of dispensed sample of 100 ml, the length of the reaction coil 60 cm, the flow rate 0.7 ml min-1 , the concentration of azure B 1.6×10-5 mol dm-3 , the concentration of phenosafranine 3.5×10-5 mol dm-3 . For the determination of heparin using azure B it was found: LOD = 0.023 IU ml-1 , LOQ = 0.186 IU ml-1 , and linear dynamic range 0.19-1.43 IU ml-1 . For the determination of heparin using phenosafranine it was found: LOD = 0.102 IU ml-1 , LOQ = 0.192 IU ml-1 , and linear dynamic range 0.19-1.79 IU ml-1 . For the determination of chondroitin sulfate using azure B it was found: LOD = 0.58 mg dm-3 , LOQ = 2.37 mg dm-3 , and linear dynamic range 2.37-8.32 mg dm-3 . The developed determination was applied to the...