Targeted uptake of folic acid-functionalized polymeric nanoparticles loading glycoalkaloidic extract in vitro and in vivo assays.

Solanum lycocarpum fruits contain two major glycoalkaloids (GAs), solamargine (SM) and solasonine (SS). These compounds are reported as cytotoxic. However, they have poor water solubility and low bioavailability. To overcome these disadvantages and getting an efficient formulation the current study aimed to develop, characterize, and test the effectiveness of a nanotechnology-based strategy using poly(D,L-lactide) (PLA) nanoparticles functionalized with folate as delivery system of glycoalkaloidic extract (AE) for bladder cancer therapy. The strategic of adding folic acid into nanoformulations can increase the selectivity of the compounds to the cancer cells reducing the side effects. Our results revealed the successful preparation of AE-loaded folate-targeted nanoparticles (NP-F-AE) with particle size around 177 nm, negative zeta potential, polydispersity index <0.20, and higher efficiency of encapsulation for both GAs present in the extract (>85 %). To investigate the cellular uptake, the fluorescent dye coumarin-6 was encapsulated into the nanoparticle (NP-F-C6). The cell studies showed high uptake of nanoparticles by breast (MDA-MB-231) and bladder (RT4) cancer cells, but not for normal keratinocytes cells (HaCaT) indicating the target uptake to cancer cells. The cytotoxicity of nanoparticles was evaluated on RT4 2D culture model showing 2.16-fold lower IC50 than the free AE. Furthermore, the IC50 increased on the RT4 spheroids compared to 2D model. The nanoparticles penetrated homogeneously into the urotheliumof porcine bladder. These results showed that folate-conjugated polymeric nanoparticles are potential carriers for targeted glycoalkaloidic extract delivery to bladder cancer cells.

Quantification of 5-FU in skin samples for the development of new delivery systems for topical cancer treatment.

5-Fluorouracil (5-FU) is used for the topical treatment of pre-cancerous skin lesions. However, previous reports focus on 5-FU quantification in plasma samples, failing to provide information about drug quantification in the skin. Therefore, the aim of this work was to develop a simple and reliable analytical method employing HPLC-UV for 5-FU quantification in skin samples. Chromatographic separation was obtained using the commonly available Lichrospher RP-C18 endcapped column. Porcine ear skin matrix-based analytical curves with thymine, as internal standard, were used. 5-FU was eluted at 5.2 min and thymine at 13.0 min. The method was selective, precise and accurate in the linear range from 0.3 to 6 µg/mL. The samples were stable after three cycles of freeze/thawing and extraction efficiency rates above 95% were obtained. In vitro skin penetration studies of an aqueous solution and a commercial cream of 5-FU were performed. The cream improved 5-FU retention into the skin and permeation through the skin compared to the solution. Therefore, the method developed herein can be applied to the study of formulations for topical delivery of 5-FU.

Depletion study and estimation of the withdrawal period for enrofloxacin in pacu (Piaractus mesopotamicus).

Defining the pharmacokinetic parameters and depletion intervals for antimicrobials used in fish will help in the development of important guidelines for future regulations by Brazilian agencies on the use of these substances in fish farming. This paper presents a depletion study for enrofloxacin (ENR) and its main metabolite, ciprofloxacin (CIP), in pacu (Piaractus mesopotamicus) fillets. The depletion study was carried out under monitored environmental conditions, with the temperature controlled at 27 °C to mimic the fish farming conditions in Brazil. ENR was administered orally via medicated feed for 10 consecutive days at daily dosages of 10 mg/kg body weight (b.w.). The fish were slaughtered at 6, 12, and 24 h and 2, 3, 5, 8, 12, 17, and 24 days after the medication period. Considering a maximum residue limit of 100 µg/kg for the sum of the ENR and CIP residues in the fillet, the results obtained in the depletion study allowed the estimation of a half-life for ENR of 2.75 days and a withdrawal period of 23 days. The results obtained in this study are important for the farming of pacu in tropical regions.

Depletion study and estimation of the withdrawal period for oxytetracycline in tilapia cultured in Brazil.

Defining pharmacokinetic parameters and depletion intervals for antimicrobials used in fish represents important guidelines for future regulation by Brazilian agencies of the use of these substances in fish farming. This article presents a depletion study for oxytetracycline (OTC) in tilapias (Orechromis niloticus) farmed under tropical conditions during the winter season. High performance liquid chromatography, with fluorescence detection for the quantitation of OTC in tilapia fillets and medicated feed, was developed and validated. The depletion study with fish was carried out under monitored environmental conditions. OTC was administered in the feed for five consecutive days at daily dosages of 80 mg/kg body weight. Groups of ten fish were slaughtered at 1, 2, 3, 4, 5, 8, 10, 15, 20, and 25 days after medication. After the 8th day posttreatment, OTC concentrations in the tilapia fillets were below the limit of quantitation (13 ng/g) of the method. Linear regression of the mathematical model of data analysis presented a coefficient of 0.9962. The elimination half-life for OTC in tilapia fillet and the withdrawal period were 1.65 and 6 days, respectively, considering a percentile of 99% with 95% of confidence and a maximum residue limit of 100 ng/g. Even though the study was carried out in the winter under practical conditions where water temperature varied, the results obtained are similar to others from studies conducted under controlled temperature.

Determination of quinolone residues in tilapias (Orechromis niloticus) by HPLC-FLD and LC-MS/MS QToF.

A method for the simultaneous determination of flumequine, oxolinic acid, sarafloxacin, danofloxacin, enrofloxacin, and ciprofloxacin in tilapia (Orechromis niloticus) fillet, using high-performance liquid chromatography with fluorescence detection (HPLC-FLD) is presented. The quinolones were extracted from the food matrix with a solution of 10% trichloroacetic acid and methanol (80:20 v/v). Clean-up of the extract was performed using polymeric solid-phase extraction cartridges. Identification of the quinolones was confirmed by liquid chromatography-tandem mass spectrometry. The HPLC-FLD method was validated in-house and the following analytical parameters were obtained: linearity higher than 0.99 for all the quinolones; intra- and interassay precisions were lower than 3.5% and 10.9%, respectively; and recoveries ranged from 73% to 110%. The limit of quantification was below the maximum residue limit established by the Joint Expert Committee on Food Additives (JECFA), which indicates that the method is appropriate for the determination of quinolones in fish fillet.