Pesticides are used worldwide to increase crop production and to remove unwanted weeds. In New Zealand, clopyralid (3,6 dichloro-2-pyridinecarboxylic acid) is one of the most widely used pesticides to control broadleaf weeds. There is very little research however reporting the removal of clopyralid in wastewaters by biological means; thus, the purpose of this research was to examine the potential biological degradation of industrial strength clopyralid wastewaters under aerobic conditions.

The study was divided into six phases, namely: Phase I (a Preliminary Phase), Phase II (an Acclimatization Phase), Phase III (a Batch Phase), Phase IV (a Kinetic Modelling Phase), Phase V (an Intermediate Identification Phase) and Phase VI (a Microbial Inhibition Phase). In the first phase, prior to the addition of clopyralid, stability parameters were assessed, these included suspended solids, COD removal, ORP and DO in a sequencing batch reactor operated under aerobic conditions. Once stability in the SBR was achieved, a commercial formulation of clopyralid at 50 mg/L of concentration was introduced. The first sign of degradation in the SBR was identified in the acclimatization phase and the results showed that 98 % of the clopyralid had been removed effectively during 5 days of acclimatization. After that, COD removal of 50 mg/L of clopyralid was measured and it was found that 70 % of the COD was removed. However, when the concentration of clopyralid was increased to 100 mg/L then the removal of clopyralid was 98 % but COD removal dropped to approximately 60 %. Due to this, batch reactors were used in phase III with a pure form of clopyralid since it was suspected that the large COD in the SBR effluent was due to additives in the commercial formulation that are not disclosed by the manufacturer due to proprietary interests.

Batch reactors were run for 24 h with initial clopyralid concentrations from 50 to 300 mg/L. It was observed that the biomass successfully treated the clopyralid up to the 250 mg/L concentration mark with 98 % removal at 100 mg/L. At the highest initial concentration of 300 mg/L, no further degradation was observed. In parallel with clopyralid degradation, COD removal was also observed; however results indicated that increasing concentrations of clopyralid created a potentially toxic effect on the biomass, with bacterial lysis most probably contributing to soluble microbial products which affected the effluent residual COD. It was also predicted that clopyralid

degradation may form different transformation products during the degradation which increased the effluent COD. Consequently, the COD removal efficiency decreased with increasing clopyralid concentration.

Phase IV checked the toxic effect of clopyralid on the biomass. It was identified that clopyralid did not have any inhibitory effect between 50 to 225 mg/L of clopyralid concentration; however, at 250 mg/L the clopyralid utilization rate dropped reaching zero at 300 mg/L, which reflects the fact that clopyralid at higher concentrations is toxic to biomass. For this experimental phase, a mathematical model was also investigated and the Luong Model (1985) proved to be the best fit.

In phase V, inorganic compounds (such as nitrate, nitrite and ammonia) and organic compounds (such as TOC), transformation products and SMPs were evaluated to test their possibility of contributing to the residual COD. It was determined that out of all these compounds, only SMPs were likely contributing to residual COD.

Finally, microbial growth inhibition was studied in the presence of clopyralid by two ways i.e. by measuring the Oxygen Uptake rate (OUR) for all species present in the sludge and microbiologically for the species cultured on media in laboratory environment. The results showed that some of the species were inhibited immediately upon contact with clopyralid as the OUR dropped suddenly to approximately half when clopyralid was introduced. However, some of the species were resistant at higher concentrations of clopyralid according to in vitro assays.

Overall, this study concludes that in batch tests (run for 24 hrs) with clopyralid injected having initial concentrations between 50 mg/L and 300 mg/L, the biomass successfully treated the clopyralid (98 % removal at 100 mg/L) up to the 250 mg/L concentration mark. At the highest initial concentration of 300 mg/L, no further degradation was observed. In parallel with clopyralid degradation, COD removal was also observed; however results indicated that increasing concentration of clopyralid created a potential toxic effect on the biomass, with bacterial lysis forming soluble microbial products that contribute to the effluent residual COD. Consequently, the removal efficiency of the COD decreased with increasing clopyralid concentrations.