Nitrofurantoin plasma- and urine exposure in eight healthy beagle dogs following standard nitrofurantoin dosing regimen

Bacterial cystitis is common in dogs and is usually treated with antibiotics. Nitrofurantoin is used for treatment of bacterial cystitis in humans and might provide a feasible treatment option in dogs. The aim of this study was to investigate the nitrofurantoin plasma concentration-time course and potential adverse effects in dogs. Nitro-furantoin (4.4-5.0 mg/kg) was administered orally to eight healthy beagles every 8 h for five days before repeated plasma and urine samples were collected. An additional four beagles served as untreated controls. The nitrofurantoin plasma and urine concentrations were measured using ultra high precision liquid chromatography coupled to tandem mass-spectrometry and further analysed using a non-compartmental pharmacokinetic model. In plasma, the median C-max was 2.1 mu g/mL, t(max) was 2 h, the terminal rate constant was 0.9 per h and the terminal half-life was 0.8 h. In urine, median C-max was 56 mu g/mL, t(max) was 1 h and the terminal half-life was 4.3 h. No adverse effects were observed clinically or in haematology or biochemistry. The data presented in this study combined with in vitro sensitivity data from common urine pathogens and the lack of observed adverse effects suggest that nitrofurantoin in a standard dosing regimen could be effective in sporadic bacterial cystitis treatment in dogs. Further clinical studies are highly warranted to verify the effectiveness in clinical cases

Chemical Derivatization in Combination with Liquid Chromatography Tandem Mass Spectrometry for Detection and Structural Investigation of Glucuronides

This thesis presents novel approaches for structural investigation of glucuronides using chemical derivatization in combination with liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MSn). Today, LC-ESI-MSn is the dominant technique for quantitative as well as qualitative analyses of metabolites, due to its high sensitivity and selectivity. However, for compounds without an easily ionizable group, e.g., steroids, the sensitivity is limited. In the work presented in this thesis, a derivatization procedure forming a basic oxime significantly increased the detection sensitivity for the altrenogest glucuronide. Furthermore, in structural evaluations of glucuronides, the limitation of LC-MSn becomes evident due to the initial neutral loss of 176 u, i.e. monodehydrated glucuronic acid, which often makes it impossible to elucidate the structures of the conjugates. To solve this problem, the main part of the work described in this thesis was devoted to chemical derivatization as a means of facilitating the determination of the site of conjugation. For the first time, the isomeric estriol glucuronides were evaluated using a combination of three reagents 2-chloro-1-methylpyridinium iodide (CMPI), 1-ethyl-3-(3-dimethyl- aminopropyl)-carbodiimide (EDC), and 2-picolylamine (PA). Interestingly, the derivatization gave a selective fragmentation pattern leading to differentiation of the isomers. Another derivatization reagent, 1,2-dimethylimidazole-4-sulfonyl chloride (DMISC), was also tested for the first time in structural investigations. The isomeric glucuronides of morphine, formoterol, and hydroxypropranolol were evaluated. They can all be conjugated in aliphatic as well as aromatic positions. DMISC was proven to be useful in two ways. Firstly, the morphine and formoterol glucuronides that contained a free phenol could be differentiated from those that were conjugated in the aromatic position based on different reactivity. Secondly, for the aromatic O-glucuronide of 4’-hydroxypropranolol, DMISC was proven to react with the amine. This product gave a different fragmentation pattern compared to the corresponding derivative of the aliphatic glucuronide

Benzylpenicillin-producing Trichophyton erinacei and methicillin resistant Staphylococcus aureus carrying the mecC gene on European hedgehogs - A pilot-study

Background A high carriage rate of methicillin-resistant Staphylococcus aureus with the mecC gene (mecC-MRSA) has been described among Wild European hedgehogs (Europeaus erineaus). Due to this frequent occurrence, it has been suggested that hedgehogs could be a natural reservoir for mecC-MRSA. However, the reason why hedgehogs carry mecC-MRSA remains unknown, but it has been hypothesized that mecC-MRSA could have evolved on the skin of hedgehogs due to the co-occurrence with antibiotic producing dermatophytes. The aim of this pilot-study was therefore to investigate if hedgehogs in Sweden carry Trichophyton spp. and to provide evidence that these dermatophytes are able to produce penicillin or similar substances. In addition, the study aimed to identify if dermatophytes co-occurred with mecC-MRSA. Methods Samples were collected from hedgehogs (Europeaus erineaus) that were euthanized or died of natural causes. All samples were screened for dermatophytes and mecC-MRSA using selective cultivation methods. Suspected isolates were characterized using PCR-based methods, genome sequencing and bioinformatic analyses. Identification of penicillin was performed by ultra-high-performance liquid chromatography-tandem mass spectrometry. Results In total 23 hedgehogs were investigated, and it was shown that two carried Trichophyton erinacei producing benzyl-penicillin, and that these hedgehogs also carried mecC-MRSA. The study also showed that 60% of the hedgehogs carried mecC-MRSA. Conclusion The pilot-study demonstrated that Trichophyton erinacei, isolated from Swedish hedgehogs, can produce benzylpenicillin and that these benzylpenicillin-producing T. erinacei co-occurred with mecC-MRSA. The study also reconfirmed the high occurrence of mecC-MRSA among hedgehogs

Seafood sold in Sweden contains BMAA: A study of free and total concentrations with UHPLC–MS/MS and dansyl chloride derivatization

β-N-Methylamino-l-alanine (BMAA) is a potential neurotoxin associated with the aquatic environment. Validated analytical methods for the quantification of both free and total concentrations of BMAA were used in an investigation of seafood purchased from different grocery stores in Uppsala, Sweden. The analysis was performed using ultra high performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UHPLC-ESI–MS/MS) and detection of BMAA as a dansyl derivate. The determined concentrations of free BMAA (after a simple trichloroacetic acid extraction) in mussels and scallops were up to 0.46 μg g−1 wet homogenate. The total BMAA (after hydrochloric acid hydrolysis) levels were between 0.29 and 7.08 μg g−1 wet mussel homogenate. The highest concentration of total BMAA was found in imported cooked and canned mussels which contained about ten times the quantity of BMAA measured in domestic cooked and frozen mussels. In this study it was also concluded that BMAA could be detected in seafood origin from four different continents. The risks associated with human exposure to BMAA through food are unknown today. However, the results of this study show that imported seafood in Sweden contain BMAA, indicating that this area needs more investigation, including a risk assessment regarding the consumption of e.g., mussels, scallops and crab