Newly Developed Methods for the Determination of Selected Pharmaceuticals in Water and Vegetable Samples by HPLC and their Removal
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Date
2021-06-22
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Addis Ababa University
Abstract
Pharmaceutical consumptions are now increasing worldwide as it is essential to treat and prevent health issues but at the end of the day, they end up in the environment as a persistent pollutant. However, very little research has been carried out in Africa and there is no study in Ethiopia concerning method development and quantitative analysis of pharmaceuticals in the environment. This research work is aimed to develop analytical methods for the determination of selected pharmaceuticals from water and vegetable samples and it also presents their removal.
An effective liquid chromatography for the determination of thirteen pharmaceuticals (metformin, amoxicillin, chloroquine, theophylline, trimethoprim, caffeine, norfloxacin, ciprofloxacin, acetylsalicylic acid, doxycycline hyclate, metronidazole, albendazole and cloxacillin) was developed and validated. Kromasil C18 column, gradient elution with aqueous formic acid (0.1%), methanol and acetonitrile, a UV absorption wavelength of 250 nm and a mobile phase flow rate of 1 mL/min over a 22 min run time was optimized for complete separation. The method was validated and results for: linearity, precision, sensitivity, accuracy, specificity, suitability and method robustness were obtained and met the ICH guidelines. Using the validated chromatographic conditions, solid-phase extraction (SPE) for the extraction of 13 pharmaceutical compounds in water samples was developed. Different types of SPE sorbent, sample solution pH, type of elution solvent, sample load volume and addition of EDTA were evaluated. The LOD and LOQ of the optimized method were in the range 0.1 - 0.8 μg/L and 0.2 - 2.6 μg/L, respectively. The proposed method was applied for water samples from Addis Ababa, Ethiopia. Trimethoprim concentration of 7.8(1.1) μg/L, caffeine 3.2(0.4) μg/L, albendazole 2.1(0.1) μg/L and norfloxacin <LOQ were observed pharmaceuticals in a hospital water sample. Antibiotics trimethoprim and ciprofloxacin in the concentration of 0.5(0.02) μg/L and 0.3(0.01) μg/L, respectively, were found in the sewerage treatment plant influent. As pharmaceuticals are found in very small amounts in the environment, UHPLC-MS was optimized for selected ten pharmaceuticals. The linearity range and the calibration curve were obtained in the concentration range of 0.5 - 1000 μg/L. Limit of LOD and LOQ found ranged from 0.010 - 0.039 μg/L and 0.033 - 0.130 μg/L, respectively. Recovery on spiked river water resulted in > 65% for 70% of the compounds and the rest were found in > 55%.
Amoxicillin was not detected in all sampling points and caffeine was found in high concentration. The concentration ranged from not detected to 48.8 μg/L.
QuEChERS sample preparation was optimized for five selected pharmaceuticals and a combination of d-SPE sorbent, MgSO4, PSA, C18 and DE (clean-up sorbent), methanol (extraction solvent), MgSO4 with NaCl (extraction salt) and 5 min extraction time were obtained. The method was successfully applied to different vegetable samples collected from Addis Ababa, Ethiopia and Johannesburg, South Africa (carrot, cabbage and lettuce) and none of the target analytes were found in the sample investigated. The matrix effect study on vegetable samples collected was found to be very high that suppressed the signal during analysis.
For selective removal of pharmaceuticals, molecularly imprinted polymer (MIP) was synthesized and optimized for albendazole, ciprofloxacin, norfloxacin and venlafaxine in an aqueous solution. The polymer was characterized by FT-IR, TGA, XRD, SEM and BET. MIP mass 50 mg, sample pH of 9 and adsorption time of 20 min and 1 mL mixture of formic acid in water and methanol as elution solvent was optimized. The adsorption experimental data fitted with the Freundlich isotherm model and the pseudo-second-order kinetic model best fitted which implied adsorption through chemisorption. MIP reusability was demonstrated for six repeated cycles without significant loss in performance. Lastly, the efficiency of Moringa oleifera (seed cake, seed husk and water-soluble protein) was studied for removal of nine pharmaceuticals. The adsorbent was characterized by FT-IR, SEM and points zero charge. Sample solution pH of 5, a contact time of 80 min and 0.5 mg/L initial concentration of the analyte were optimized. Adsorption on the Moringa adsorbent was well defined by Freundlich adsorption isotherm model and the pseudo-second-order kinetic model. The results obtained showed that the use of this plant can be considered as one of the most promising, and low-cost adsorbents for the removal
of pharmaceuticals.
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Keywords
Pharmaceuticals, Water, Solid Phase Extraction, UHPLC-MS, Vegetable, Quecehers, Molecularly Imprinted Polymer, Moringa Oleifera