Ming-Huang Chang

Liquid Chromatographic Determination of Amoxicillin Residues in Grouper Muscle Following Oral Administration of the Veterinary Drug

Improper application of antibiotic chemicals to livestock, poultry and aquaculture species may lead to the occurrence of residues in food supplies. An appropriate depletion is needed after the administration of drugs to animals for ensuring that residues in edible tissues are below established tolerance levels. The study was investigated to determine incurred amoxicillin residues in grouper (Epinephelus malabaricus) muscle following oral administration. Dosed fish were harvested after five depletion periods, and muscle fillets were analyzed for amoxicillin residues using an HPLC method with solid phase extraction and UV detection. The residue levels in grouper after a 6-h depletion (T(subscript max)) were at the maximum 0.91 and 1.31µg/g (C(subscript max)) toward 20 and 40 mg/kg b.w. dosage. Residue levels after a 72-h depletion decreased to below the methods limit of quantitation after last oral treatment. At the dosage (40 mg/kg b.w.), we suggested that the recommended withdraw period was 5 days.

Detection of Residues by Use of Solid-Phase Extraction and Reversed Phase High-Performance Liquid Chromatography after Oral Administration of Amoxicillin in Bass Muscle

An experimental trial was performed to establish an adequate depletion period of amoxicillin in bass. The amount of amoxicillin was applied by oral administration at the dose of 20 and 40 mg/kg body weight, and was given at a daily dose for five consecutive days. Amoxicillin levels were determined using a solid-phase extraction and high-performance liquid chromatography (HPLC) method. The drug was eluted from Mightysil RP-18 GP 250-4.6 mm φ5 m column with a mobile phase consisting of 0.005 M KH2PO4 buffer with pH 6.8-MeOH (90:10, v/v), at a flow rate of 1 mL/min. The effluent was monitored using a UV detector set at 210 nm. Each analysis required no longer than 15 mm. The validated concentration ranges of this method were 0.01 μg/mL to 10 μg/mL for working standard solutions, and 0.04 to 10 μg/g for muscle. Mean absolute recoveries were 78%. The limits of detection (LCD) and limits of quantitation (LOQ) for bass muscle were 0.04 and 0.1 quantitation g/g, respectively. After 5 consecutive oral doses of amoxicillin, residues in 6-h-postdosing (T(subscript max)) bass were at the maximum 0.84 and 1.18 μg/g (C(subscript max)) toward 20 and 40 mg/kg b.w. dosage, residue levels after a 72-h depletion decreased to below the methods limit of quantitation after last oral treatment. At the dosage (40 mg/kg b.w.), we suggested that the recommended withdraw period was 5 days.

Effects of Lepidium meyenii Walp (Maca) on Fecundity and Puppy Growth in Mice

Maca (Lepidium meyenii Walp) is a medicinal herb belonging to Aburana family native to Andes mountains in the Latin America. This herb has been reported with various reproductive efficacy and natives of Peru have been using it as a medicinal plant. In this study, we attempted to investigate the fecundity and other reproductive traits along with puppy weights in an in vivo experiment on laboratory mice. 15 female mice and 5 male mice were raised in the animal experiment center, college of medical science, National Taiwan University. They were mated as mentioned as in the methodology and from the second generation, more 15 females were selected to be mated. The feed of the experiment group was while the control group had normal chow. The fecundity rate, body weights were monitored between the control group and test groups added with 30% Maca. There was no significant change in body weight (P＞0.05) of the selected mice for mating before pregnancy and it was also similar in the selected offspring (P＞0.05). However, with regard to litter size in the first litter (FI), the test group (TFI) had 171 pups, showing significant difference to the control group (CF1) also with 128 pups (P＞0.01). In the second litter (FII), also the number of 188 pups from the test group (TFII) was significantly higher than the control group (CFII) (P＜0.05). The total offspring of the first and the second litter in both test and control groups were 359 and 283 respectively, presenting a significant difference between them (P＜0.01). There was no significant difference in the post weaning in both groups (P＞0.05). In the first litter (FI), the weight of pups in TFI was significantly heavier than the control group (CFI) during t he first and the second week (P＜0.05); in the second litter (FII), TFII was found to be significantly heavier than CFII only in the first week (P＜0.05).

ESTIMATION OF THE WITHDRAWAL PERIOD FOR AMOXICILLIN IN POMPANO (TRACHINOTUS BLOCHII) SERUM AND MUSCLE

Amoxicillin is one of the most popular antibiotics used in aquaculture. To ensure the residue of amoxicillin in the serum and the muscle of pompano (Trachinotus blochii) is below the established tolerance levels; we used reversed phase high-performance liquid chromatography (HPLC) to measure the amoxicillin in pompano after oral administration. The pharmacokinetics of amoxicillin in the serum and the muscle of pompano were established by single oral administration or administrations for five consecutive days at the dose of 40 mg/kg. After different depletion periods, amoxicillin residue was analyzed in both the serum and the muscle fillets by HPLC. The verified concentrations of this method were 0.01 to 10 μg/mL; the limit of detection (LOD) and the limit of quantitation (LOQ) were 0.04 and 0.1 μg/g. The concentration of amoxicillin in the serum of pompano reached the maximum threshold of 7.36 μg/g after 0.5 h depletion in the experiment with a single dose of amoxicillin (40 mg/kg of body weight). In addition, the maximal concentration (1.96 μg/g) of amoxicillin was detected in the muscle 2 h after depletion. Furthermore, the residue levels had decreased below the limit of detection after 48 h and 72 h in the serum and the muscle. Here we also suggested that 5 days of withdrawal were required after the multiple dosing of amoxicillin.

Structural Changes of the Gastrocnemius Muscle of Emasculated Native Taiwanese Chickens Using Succinate Dehydrogenase Localization

The aim of this study was to compare the muscle fiber structural changes of the medial head and lateral head of the gastrocnemius muscle of intact cocks (Group 1), emasculated cocks with testicular tissue remaining (Group 2), emasculated cocks (Group 3), and intact hens (Group 4) of native Taiwanese DL2 chicken using results of succinate dehydrogenase localization studies. The results showed that the gastrocnemius muscle of each group of chickens contained Type-Ⅰ red muscle fibers, Type-ⅡA white muscle fibers and Type-ⅡB white muscle fibers. However, the percentages in each group varied. For the medial head of the gastrocnemius muscle, the ratio of white muscle fibers was higher than that of red muscle fibers. For the lateral head of the gastrocnemius muscle in the group 3, the ratio of white muscle fibers was higher than that of red muscle fibers. After comparing the muscle fiber diameters with the muscle fibers of each type in the medial head of the gastrocnemius muscle, there were no significant differences (P ＞ 0.01) among the four groups. But compared the muscle fiber diameters of the lateral head of the gastrocnemius muscle, type-Ⅰ red muscle fibers and type-ⅡA white muscle fiber of group 2 and group4 were significant differences (P ＜ 0.01). However merely the Type-ⅡB white muscle fiber of the group 2 were significant differences (P ＞ 0.01).

EVALUATION OF AMOXICILLIN WITHDRAWAL PERIOD TIME IN POMPANO LIVER AND KIDNEY

Amoxicillin is an antibiotic which belongs to the group of penicillins. It is approved in Taiwan for treating bacterial infections caused by Streptococcus sp. and Photobacterium sp. in anguilliformes, perciformes and salmoniformes. The pharmacokinetics of amoxicillin were determined in pompano following oral administration of a single dose of 40 mg/kg. Residue studies were performed to determine residues in liver and kidney tissues of healthy fish after oral gavage of amoxicillin at a daily dose of 40 mg/kg for five consecutive days. Amoxicillin residues were analyzed by HPLC using Hypersil-100 C18 column (150 mm × 4.6 mm i.d.) and mobile phase consists of 10 mM K2HPO4 (pH 8.5) with acetonitrile (80:20, v/v), at a flow rate of 1 mL/min. The effluent was monitored using a fluorescence detector set as 358 and 440 nm as excitation and emission wavelengths. Following a single oral dose, amoxicillin residues in 0.5 h post-dosing pompano were at a maximum of 6.17 μg/g in liver and 4.27 μg/g in kidney; the concentration of amoxicillin in liver and kidney declined with half-lives of 18.3 and 12.0 h. Amoxicillin residues in pompano liver and kidney tissues were proved to be under the MRL, 0.5 ug/g (liver and kidney) after a withdrawal period of five days.

CRITICAL-SIZED OSTEOCHONDRAL DEFECTS OF YOUNG MINIATURE PIGS AS A PRECLINICAL MODEL FOR ARTICULAR CARTILAGE REPAIR

Purpose: Evaluation by animal models is essential for tissue engineering-based articular cartilage repair techniques. Larger animals are considered to more closely approximate the clinical situations in translational medicine. Therefore, we used miniature pigs and induced full-thickness and osteochondral defects in them. We also studied if there were instances of spontaneous repair for providing baseline data for further cartilage regeneration study. Methods: A total of 12 miniature pigs with average age of 7.4 months were used in this study. Full-thickness and osteochondral defects with 2.7, 4.5 or 8.0 mm diameter were created at medial femoral condyles in the same pig, respectively. The pigs were sacrificed at 8, 16, 24 and 48 weeks. Gross appearances of defects were observed at aforementioned time points, and the histological analyses, including H&E and alcian blue staining, were performed for consequent evaluation as well. Results: The results showed that defects created in the center of the femoral condoyle migrated to periphery, and it implied that the pigs were still growing after 7–8 months of age. However, spontaneous repair was observed in 2.7 mm diameter defects but rarely seen in 4.5 and 8 mm diameter osteochondral defects. On the other hand, osteochondral defects repaired better than defects of full-thickness in the same 2.7 mm diameter. Conclusions: In order to prevent spontaneous repair of osteochondral defect in a young miniature pig animal model, a critical-sized osteochondral defect larger than 40% width of femoral medial condyle (4.5 mm in miniature pig) and observation period for more than 48 weeks are suggested by this study.