Suspect screening of antimicrobial agents transformation products in environmental samples development of LC-QTrap method running in pseudo MRM transitions.

The aim of the work was to develop a new HPLC-MS/MS method that allows for the simultaneous detection of antimicrobials agents (targeted analysis) and their transformation products (non-targeted analysis), which enabled the elucidation of their transformation pathways in the environment. Targeted analysis was performed for 16 selected antimicrobials agents (AMs) in wastewater collected at different stages of the treatment process and river water from sections before and after wastewater discharge. The samples were collected in the Lyna sewage treatment plant (Olsztyn, Poland) in three measuring periods at different seasons. Analytes were selected from tetracyclines, fluoroquinolones, ß-lactams, macrolides, glycopeptides, lincosamides and synthetic antibiotics. As a part of the targeted analysis, 13 AMs were detected in wastewater samples, and 7 of them in river water samples. However, their presence and concentrations were closely related to the type of the sample and the season in which the sample was taken. The highest concentrations of AMs were detected in samples collected in September (max. 1643.7 ng L-1 TRI), while the lowest AMs concentrations were found in samples collected in June (max. 136.1 ng L-1 CLR). The total content of AMs in untreated wastewater was in the range of 1.42-1644 ng L-1, while in the river water was for upstream 1.22-48.73 ng L-1 and for downstream 2.24-149 ng L-1. In the non-target analysis, 33 degradation products of the selected AMs were identified, and the transformation pathways of their degradation were speculated. In the course of the research, it was found that as a result of the processes taking place in wastewater treatment plant, the parent substances are transformed into a number of stable transformation products. Transformation products resulted from hydroxylation, ring opening, oxidation, methylation or demethylation, carboxylation, or cleavage of the CN bond of the parent AMs.

Application of UHPLC-MS/MS method to study occurrence and fate of sulfonamide antibiotics and their transformation products in surface water in highly urbanized areas.

Sulfonamide antibiotics (SAs) are used on a large scale in human and veterinary medicine. The main goal of this study was to develop a method for the detection of selected SAs (sulfamethoxazole, sulfadiazine, sulfamethazine, sulfathiazole, sulfapyridine, sulfamerazine, sulfamethiazole, and sulfisoxazole) in aqueous samples (targeted analysis), and then conduct a non-targeted analysis to determine the transformation products to elucidate their degradation pathways. These analyses were performed using ultra-high-performance liquid chromatography coupled to tandem mass spectrometry. The procedure was used to detect selected antibiotics in water samples collected throughout a highly urbanized area. Among the studied compounds, sulfamethoxazole (max. 78.88 ng L-1) and sulfapyridine (max. 38.88 ng L -1) were the most common pollutants identified in surface waters. Trace amounts of sulfadiazine (below LOQ = 0.40 ng L-1) were also detected. Next, the samples were screened to detect the transformation products. Several sulfadiazine and sulfamethoxazole transformation products were detected and confirmed in the environmental samples.

Microbial and chemical pollutants on the manure-crops pathway in the perspective of "One Health" holistic approach.

This study determined the impact of poultry and bovine manure fertilization on the content of antibiotics, heavy metals (HMs), and the quantitative and qualitative composition of integrase and antibiotic resistance genes in soil, groundwater, and crops cultivated on manure-amended plots. Antibiotic concentration levels were analyzed using the HPLC-MS/MS, heavy metal concentration level were measured by HGAAS and ICP-OES, while the integrase genes and ARGs were quantified using Real-Time PCR (qPCR) method. Manure, soil, and crops samples contained the highest concentration of Zn (104-105 ng gdm-1) and Cu (103-105 ng gdm-1) of all HMs tested. Manure-supplemented soil was characterised by a high concentration of doxycycline and enrofloxacin. A high abundance of integrase genes was noted in samples of manure (109-1010 copies gdm-1) and soil (107-108 copies gdm-1). Among all the analyzed genes, sul1, sul2, blaTEM, and integrase genes were the most common. Results of the study demonstrate the selective character of ARGs transfer from poultry and bovine manure to plants. The only gene to occur in all studied environmental compartments was sul1 (from 102 - groundwater to 1011 - poultry manure). It was also found that animal manure may cause an increase in the HMs concentration in soil and their accumulation in crops, which may influence the health of humans and animals consuming crops grown on manure-amended soil. The high abundance of integrase genes and ARGs and their reciprocal correlations with HMs pose a serious risk of the rapid spread of antibiotic resistance in the environment. Moreover, unusual dependencies between integrase genes and selected ARGs indicate the possibility of changes in the mobility nature of genetic elements.

Optimization of a Method for Extraction and Determination of Residues of Selected Antimicrobials in Soil and Plant Samples Using HPLC-UV-MS/MS.

The residues of antimicrobials used in human and veterinary medicine are popular pollutants of anthropogenic origin. The main sources of introducing antimicrobials into the environment are sewage treatment plants and the agricultural industry. Antimicrobials in animal manure contaminate the surrounding soil as well as groundwater, and can be absorbed by plants. The presence of antimicrobials in food of plant origin may pose a threat to human health due to their high biological activity. As part of the research, a procedure was developed for the extraction and determination of ciprofloxacin, enrofloxacin, cefuroxime, nalidixic acid and metronidazole in environmental samples (soil and parsley root). An optimized solid-liquid extraction (SLE) method was used to separate antimicrobials from the solid samples and a mixture of citrate buffer (pH = 4): methanol (1:1; v/v) was used as the extraction solvent. Solid phase extraction (SPE) with OASIS® HLB cartridges was used to purify and pre-concentrate the sample. The recovery of the developed method was in the range of 55-108%. Analytes were determined by high-performance liquid chromatography coupled with an ultraviolet (UV) detector and a tandem mass spectrometer (HPLC-UV-MS/MS). The procedure was validated and applied to the determination of selected antimicrobials in soil and parsley root samples. Five types of soil and five types of parsley roots of different origins were analyzed. The presence of nalidixic acid in the parsley root samples was found in the concentration range of 0.14-0.72 ng g-1. It has been shown that antimicrobials are absorbed by the plant and can accumulate antimicrobials in its edible parts.

Supramolecular Solvent-Based Microextraction of Selected Anticonvulsant and Nonsteroidal Anti-Inflammatory Drugs from Sediment Samples.

The increase in the production and consumption of pharmaceuticals increases their presence in the global environment, which may result in direct threats to living organisms. For this reason, there is a need for new methods to analyze drugs in environmental samples. Here, a new procedure for separating and determining selected drugs (diclofenac, ibuprofen, and carbamazepine) from bottom sediment and water samples was developed. Drugs were determined by ultra-high performance liquid chromatography coupled with an ultraviolet detector (UHPLC-UV). In this work, a universal and single-step sample treatment, based on supramolecular solvents (SUPRAS), was proposed to isolate selected anticonvulsants and nonsteroidal anti-inflammatory drugs (NSAIDs) from sediment samples. The following parameters were experimentally selected: composition of the supramolecular solvent (composition THF:H2O (v/v), amount of decanoic acid), volume of extractant, sample mass, extraction time, centrifugation time, and centrifugation speed. Finally, the developed procedure was validated. A Speedisk procedure was also developed to extract selected drugs from water samples. The recovery of analytes using the SUPRAS procedure was in the range of 88.8-115%, while the recoveries of the Speedisk solid-phase extraction procedure ranged from 81.0-106%. The effectiveness of the sorption of the tested drugs by sediment was also examined.

Metabolic profiles in the course of the shikimic acid pathway of Raphanus sativus var. longipinnatus exposed to mesotrione and its degradation products.

The influence of pesticides on the metabolism of edible plants has not been fully investigated. Moreover, once introduced into the environment, pesticides are degraded to many compounds with undefined bioactivity. In presented work, under experimental conditions, model edible plant (Raphanus sativus var. longipinnatus) was exposed to herbicide stress by application of a herbicide (mesotrione, 2-(4-methanesulfonyl-2-nitrobenzoyl)cyclohexane-1,3-dione, MES) or its degradation products (amino-4-(methylsulfonyl)benzoic acid, AMBA; 4-(methylsulfonyl)-2-nitrobenzoic acid MNBA; cyclohexane-1,3-dione, CHD). Metabolic profiles of plants were employed to estimate the plant's defence response to MES and its metabolites. The intensity of herbicide stress was determined by measuring the changes in chlorophyll and catecholamines concentration formed in the shikimic acid pathway. Non-target analysis was conducted by LC-MS/MS, determination of catecholamines by LC-FL, chlorophyll by spectrophotometry. The highest phytotoxicity is characterized by MES (2000%-fold increase in the content of herbicide stress marker (normetanephrine) compared to a blank), followed by CHD (500%) combined with 15% increase in chlorophyll concentration. AMBA and MNBA as stress factors caused the increase in the content of catecholamines in the plant (86-160%). Simultaneously, an increase in chlorophyll content was observed (26-50%). Such diversity of the organism's defence response, also visible on metabolic profiles, can be associated with the chemical structure of compounds that are stress factors. MES and CHD, in contrast to AMBA and MNBA, have cyclohexano-1,3-moiety in their structure, which seems to be responsible for herbicidal properties.