Systemic therapy with oxytetracycline is often used for treatment of clinical metritis although data about its penetration into the uterus and uterine secretion are lacking. Uterine secretions and milk from six cows with clinical metritis were collected for microbiological assay. The animals were treated intramuscularly with long-acting oxytetracycline (20 mg/kg) and samples of plasma, milk and uterine secretions were collected for determination of the antibiotic concentrations by HPLC-PDA analysis. Pharmacokinetics of the antibiotic and in silico prediction of its penetration into the uterus were described. Trueperella pyogenes with MIC values of 16–64 µg mL⁻¹ was isolated (n of cows = 4) from uterine secretions. Oxytetracycline showed fast absorption and penetration in the uterine secretions and milk. No change of withdrawal time for milk was necessitated in cows with clinical metritis. Maximum levels in uterine secretions and predicted concentrations of oxytetracycline in the uterus were lower than MIC values. Systemic administration of long-acting oxytetracycline did not guarantee clinical cure and was not a suitable choice for treatment of clinical metritis associated with Trueperella pyogenes. The appropriate approach to antibiotic treatment of uterine infections of cows requires knowledge on penetration of the antibiotics at the site of infection and sensitivity of pathogens.

Puerperal metritis in cows is often treated with antibacterial drugs. The prudent use of antibiotics in farm animals requires state-of-the-art knowledge of their pharmacokinetics and data from sensitivity tests of pathogenic bacteria. Changes in oxytetracycline levels in the uterine secretion over time after intrauterine administration in cows with metritis were evaluated in relation to the sensitivity of pathogenic bacterial isolates. Oxytetracycline levels in plasma, milk and uterine secretion were determined via LC–MS/MS analysis. Pathogenic bacteria were isolated and their sensitivity to oxytetracycline was determined. The concentrations of oxytetracycline in the uterine secretion were 433.79 (39.17–1668.76) µg·mL⁻¹ six hours after the third application at a dose of 8 mg/kg and 84.33 (1.58–467.55) µg·mL⁻¹ 96 h after the last treatment. These levels were higher than the minimum inhibitory concentrations—namely, between 4 and 64 µg·mL⁻¹—against pathogenic isolates Trueperella pyogenes, Streptococcus intermedius, Escherichia coli and Bacillus pumilis. Higher concentrations over time were measured in milk rather than in plasma, indicating the need to monitor the withdrawal time for milk. The intrauterine infusion of oxytetracycline for three consecutive days resulted in the successful treatment of metritis in terms alleviating inflammation and restoring the estrus cycle in cows.

A fast and sensitive HPLC method was validated in order to analyze doxycycline in plasma and milk of sheep and in plasma of rabbits. The samples were processed with trifluoroacetic acid (TFA). After the centrifugation step, a supernatant containing extracted doxycycline and internal standard oxytetracycline was injected into the HPLC system with PDA detection. The method showed linearity in the range of 0.125 - 2.5 µg/mL for ovine plasma, 0.125 – 5.0 µg/mL for ovine milk, and 0.125 – 1 µg/mL for rabbit plasma. The inter-assay precision varied between 5.69 – 13.55 %. Values for intra-assay precision were between 0.62 – 8.67 %. Accuracy was higher than 90% in all of the tested concentrations in the three types of biological matrices. The mean extraction recovery was higher than 90 % for all matrices. In order to handle only with free drug concentrations, microfiltration of standard solutions with low (0.25mg/mL), medium (0.5mg/mL) and high (1.0mg/mL) concentration was performed. A percentage for correction of the quantified doxycycline was calculated. The most significant adjustments should be made at the low concentrations. The correction for rabbit plasma is 24.63±5.03%, for ovine plasma is 20.10±8.01% and for milk–16.68±0.04 %. This method can be used for routine determination of doxycycline concentrations for pharmacokinetic studies and further dosage adjustment.

Doxycycline is a well-tolerated tetracycline antibiotic, registered for use in rabbits and administered for treatment of bacterial infections in this animal species. Nevertheless, the available pharmacokinetic data are limited and this study aimed to investigate the pharmacokinetics of orally administered doxycycline in mature and immature rabbits by application of the population approach. The rabbits were treated orally with doxycycline hyclate (5 mg/kg bw) in the form of a solid gelatin capsules. Free plasma concentrations were determined with HPLC analysis with Photodiode array detection. The estimated typical value of volume of distribution (tvV), total body clearance, and absorption rate constant were 4.429 L/kg, 1.473 L/kg/h, and 0.257 h⁻¹, respectively. The highest between-subject variability (BSV) of 69.30% was observed for tvV. Co-variates such as body weight, age, and biochemical parameters did not improve the tested model and did not contribute to explanation of the BSV. The population pharmacokinetic model of the orally administered doxycycline in rabbits should be further developed by addition of data from more animals treated with higher doses. An oral dose of 5 mg/kg could ensure percentage of the time from the dosing interval during which the concentration is above minimum inhibitory concentration (MIC) %fT > MIC of 35% if MIC of 0.18 μg·mL⁻¹ and a dosing interval of 12 h is assumed which does not cover criteria for rational use of antibiotics.

Orally administered tetracycline antibiotics interact with feed, which may impact their bioavailability and efficacy. Therefore, the pH-dependent adsorption of doxycycline and its interaction with feed for ruminants was studied in vitro. Adsorption experiments on animal feed (135 and 270 mg) with initial doxycycline concentrations of 35, 75, and 150 µg/mL were performed. Desorption experiments were conducted by agitation of a predetermined mass of doxycycline-loaded animal feed in PBS, at pH = 3.0, 6.0, and 7.4, to simulate changes in the gastrointestinal tract. Antibiotic concentrations were determined by LC-MS/MS analysis. The adsorption/desorption of doxycycline was described by mathematical models. Chemisorption with strong intermolecular interactions between the active functional groups of doxycycline and the organic biomass was found. The experimental release curve comprised three sections: initial prolonged 27–30% release (pH = 6.0), followed by moderate 56–59% release (pH = 3.0), and final 63–74% release (pH = 7.4). The sigmoidal model showed a considerable role of diffusion with an initial prevalence of desorption and a decreased desorption rate thereafter. The Weibull equation revealed an initial release stage followed by a lag time section and sustained release. The study of doxycycline adsorption by the animal feed proved a maximum 80% encapsulation efficiency and revealed initial diffusion followed by chemisorption. The highest release efficiency of 74% suggests high bioavailability of doxycycline after oral administration in ruminants.