Optimisation of HPLC-based methods for the separation and detection of herbicide glyphosate and its major metabolite in water

Submitted in partial compliance with the requirements for the Masters Degree in Technology: Chemistry, Durban University of Technology, Durban, South Africa, 2010.Water storage dams play an important part in the collection and purification of water destined for human consumption. However, the nutrient rich silt in these dams promotes rapid growth of aquatic plants which tend to block out light and air. Glyphosate is universally used as the effective non-selective herbicide for the control of aquatic plants in rivers and dams. Invariably there is residual glyphosate present in water after spraying of dams and rivers with glyphosate herbicide. The amount of residual glyphosate is difficult to determine on account of high solubility of glyphosate in water. Thus a method of sample preparation and a sensitive HPLC method for the detection of trace amounts of glyphosate and its major metabolite aminomethylphosphonic acid (AMPA) in water is required. A crucial step in sample preparation is pre-column derivitization of glyphosate with 9-fluorenylmethyl chloroformate (FMOC-Cl). For sample pretreatment, water samples were derivatized with FMOC-Cl at pH 9, extracted with ethyl acetate and sample clean-up was carried out by passing a sample through the SPE cartridge. For SPE, recovery studies were done to choose a suitable cartridge for glyphosate and AMPA analysis. The following cartridges were compared, namely, C18, Oasis HLB and Oasis MAX SPE cartridges. Best recoveries (101% for glyphosate and 90% for AMPA) were obtained using 500 mg of C18 solid-phase extraction cartridge. The eluent from SPE cartridge was injected into HPLC column. Three types of separation columns (namely; C18 column, silica based amino column and polymeric amino column) were compared for the separation of glyphosate and AMPA. The best separation of glyphosate and AMPA in water samples was achieved using a polymeric amino column and a mobile phase at pH 10 which contained a mixture of acetonitrile and 0.05 M phosphate buffer (pH 10) 55:45, (v/v) respectively. The method was validated by spiking tap water , deionized water and river water at a level of 100 μg/l. Recoveries were in the range of 77% -111% for both analytes. The method was also used in determining the levels of glyphosate and AMPA in environmental samples. This method gave detection limits of 3.2 μg/l and 0.23 μg/l for glyphosate and AMPA respectively. The limits of quantification obtained for this method were 10.5 μg/l and 3.2 μg/l for glyphosate and AMPA respectively.Eskom Tertiary Education Support Programme (TESP) Durban University of Technology.

Determination of selected acidic pharmaceutical compounds in wastewater treatment plants

A thesis submitted to the Faculty of Science, University of the Witwatersrand in fulfilment of the requirements for the degree of Doctor of Philosophy. November 2016.This research was directed towards the environmental monitoring and assessment of the most used non-steroidal anti-inflammatory drugs (NSAIDs) which are naproxen, ibuprofen and diclofenac. The work involved the development and application of sensitive techniques for the quantification of naproxen, ibuprofen and diclofenac in the South African aquatic environment. Based on this information, a multi-templates molecularly imprinted polymer (MIP) was synthesized and applied alongside the commercial available sorbent (Oasis MAX) in the solid-phase extraction (SPE) of target compounds from water samples. The extracted compounds were then quantified using high performance liquid chromatography (HPLC). MIP was synthesized by applying a bulk polymerization approach at 70 ͦ C where all target compounds were used as multi-templates. Other reagents used in synthesis were 2-vinyl pyridine, 1,1’-azobis-(cyclohexanecarbonitrile), ethylene glycol dimethacrylate and toluene as functional monomer, initiator, cross-linker and porogenic solvent, respectively. Synthesis of a non-imprinted polymer (NIP) under similar reaction conditions as MIP was carried out with the omission of templates. Techniques employed in characterization of MIP and NIP were Fourier transform infrared spectroscopy (FTIR), Brunauer, Emmett and Teller (BET) method, CHNS analyzer, zeta potential, cross-polarization/magic angle spinning nuclear magnetic resonance spectroscopy, thermogravimetric analysis, differential scanning calorimetry and x-ray diffraction. Monomer-template interactions were investigated using molecular dynamics. The performance of the MIP was evaluated based on its ability to selectively extract target compounds in organic (acetonitrile, acetone, chloroform and toluene) and aqueous media. The extraction capacity of the MIP in organic solvents for naproxen, ibuprofen and diclofenac increased from high polarity to low polarity solvents. In a low polarity solvent (toluene), the extraction capacity achieved for naproxen, ibuprofen and diclofenac were 14.4, 11.0 and 14.0 mg/g, respectively. In this case, the selectivity of the MIP where gemfibrozil was employed as the competing species was evident. Selectivity of the MIP collapsed during the adsorption of naproxen, ibuprofen and diclofenac from water using gemfibrozil and fenoprofen as competitors. This resulted in high extraction efficiencies for target compounds and competitors, however, both gemfibrozil and fenoprofen were easily desorbed from the MIP using weak organic solvent due to lack of molecular recognition. During the binding sites characterization, the best fit of pseudo-second-order implied a chemisorption of all target compounds onto MIP sorbent. The data also fitted well in Langmuir isotherm which meant that the adsorption of target pharmaceuticals occurred on the homogeneous binding sites of the MIP. Optimized adsorption conditions in water such as MIP amount of 50 mg, extraction time of 10 min, sample pH of 2.5 and sample volume of 10 mL were applied for the selective adsorption of naproxen, ibuprofen and diclofenac in contaminated wastewater and river water. In WWTP influent, naproxen recovery was 38%, whereas ibuprofen and diclofenac were 69% and 87%, respectively. MIP was further used as a selective adsorbent in solid-phase extraction (SPE) of three drugs from environmental samples. The selectivity of the MIP in environmental samples was compared to that of the commercially available Oasis MAX sorbent. The application of molecularly imprinted solid-phase extraction (MISPE) reduced matrix effects and improved the sensitivity of the analytical method. In this case, the detection limits for naproxen, ibuprofen and diclofenac were 0.2, 1 and 0.6 μg/L, respectively. When deionized water was spiked with 5 and 50 μg/L of target compounds, recoveries greater than 80% were obtained. Thereafter, the developed MISPE was applied for selected acidic drugs from environmental samples. Environmental samples were collected from urban (Durban) and semi-urban/rural areas (Ladysmith) of KwaZulu-Natal Province in South Africa. The most abundant compound in the environment was ibuprofen. In river water samples from Durban, the maximum concentrations found for naproxen, ibuprofen and diclofenac were 6.8, 19 and 9.7 μg/L, respectively. The maximum amounts found for the same drugs in Ladysmith river samples were generally lower with naproxen, ibuprofen and diclofenac detected at 2.8, 6.7 and 2.6 μg/L, respectively. The same trend was observed in wastewater. Further work on the monitoring of acidic compounds in wastewater was conducted using Oasis MAX as the SPE sorbent prior to HPLC analysis. All target compounds were detected in Kingsburg and Umbilo WWTPs located in Durban surroundings. The influent and effluent concentrations detected were in the ranges of 6.4 to 69 μg/L and 0.6 to 4.2 μg/L, respectively. Further to this, the removal efficiency of the target compounds during the WWTP process in Kingsburg and Umbilo was in the range of 69 to 97%. The extent of pollution in the environment was further assessed by the monitoring of ketoprofen and triclosan in wastewater and river water using SPE with Oasis HLB sorbent and HPLC. Traces of both compounds ranging from 1.2 to 9.0 μg/L were detected in wastewater. The maximum concentrations found in river water were 2.0 and 0.9 μg/L for ketprofen and triclosan, respectively. Overall, the analytical methods implemented in this work were highly accurate, precise and sensitive. The synthesized MIP was highly selective and its application in environmental studies led to the development of a less expensive analytical method. This work also gives the overview of the extent of water pollution caused by acidic pharmaceuticals in various water matrices.MT201

Simultaneous determination of naproxen, ibuprofen and diclofenac in wastewater using solid-phase extraction with high performance liquid chromatography

The occurrence and removal efficiency for naproxen, ibuprofen and diclofenac in two of eThekwini Municipality’s wastewater treatment plants (WWTPs), Kingsburgh and Umbilo, were investigated. This paper describes a simple method that can be used routinely for the simultaneous determination of such compounds in the influent and effluent of the WWTPs. Target compounds were extracted from wastewater and pre-concentrated using the optimized Oasis MAX solid-phase extraction (SPE) method. During SPE, the pH of wastewater samples was adjusted to 2.5; then 100 mL of each sample was loaded onto a pre-conditioned cartridge. The SPE cartridge was rinsed with methanol:water (10:90%, v:v) prior to sequential elution of retained analytes with 2 mL methanol, followed by 2 mL methanol and acetic acid (90:10, v:v) and 2 mL of 2% (v:v) formic acid diluted using a mixture of methanol and acetic acid (40:60, v:v). The eluted analytes from the SPE cartridge were quantified using high performance liquid chromatography (HPLC) equipped with photo diode array detection. The analytical method was validated by spiking deionized water with 5 and 50 μg·L-1 of target compounds, for which the recovery range of 76 to 98% was achieved with good precision. The instrument quantification limits obtained were 0.1 μg·L-1, for naproxen and 0.4 μg·L-1 for both ibuprofen and diclofenac. The detected concentrations for naproxen, ibuprofen and diclofenac in the influent of both WWTPs were in the ranges of 15–20 μg·L-1, 55–69 μg·L-1 and 6.4–16 μg·L-1, respectively. In effluent, the detected concentrations for naproxen, ibuprofen and diclofenac were in the ranges of 0.6–1.1, 2.1–4.2 and 1.4–2.0 μg·L-1, respectively. Overall, the employed SPE-HPLC method led to rapid pre-concentration of target compounds prior to their trace quantification in wastewater samples