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High performance liquid chromatographic assay of amprolium and ethopabate in chicken feed using solid-phase extraction.

A method for the assay of mixtures of amprolium and ethopabate in chicken feed was developed utilizing reversed-phase high-performance liquid chromatography (HPLC) after sample clean-up of a methanolic extract by solid-phase extraction using CN cartridges. HPLC was done with benzocaine as internal standard on a C-8 column with methanol-water 40:60, containing octanesulfonic acid, triethylamine and acetic acid, as mobile phase. Eluate was monitored at 274 nm. Baseline separation was achieved with retention times of approximately 7.5, 9.4, and 10.4 min, for amprolium, benzocaine, and ethopabate respectively. Feed constituents did not give peaks after 6.5 min. Peak area ratios were linear over 10-180 ng of amprolium, and 2-18 ng of ethopabate injected. Limits of quantitation at AUFS 0.05 were 0.5 and 0.3 ng respectively. Recovery studies from spiked feed (n = 9), covering +/- 30% of usual doses in feed, gave percent recoveries (+/- SD) of 99.4 +/- 1.4% for amprolium and 100.5 +/- 2.6% for ethopabate. Applying the method to two different batches of commercial feed gave results which were comparable to those obtained by the AOAC spectrofluorometric methods.

Amprolium↗

Sensitive determination of ethopabate residues in chicken tissues by liquid chromatography with fluorometric detection.

A liquid chromatographic (LC) method is described for determination of ethopabate residues in chicken tissues. The drug is extracted from tissues with acetonitrile, and the extract is concentrated to 2-3 mL. This aqueous solution is rinsed with ethyl acetate and cleaned up by Florisil column chromatography. LC analysis is carried out on a Zorbax ODS column, and ethopabate is quantitated by using a fluorometric detector set at 306 nm (excitation) and 350 nm (emission). Recoveries of ethopabate added to chicken tissues at levels of 0.01 and 0.05 ppm were 87.8 and 92.7%, respectively. The detection limit was 100 pg for ethopabate standard, and 0.5 ppb in chicken tissues.

Aminobenzoates↗

High pressure liquid chromatographic determination of ethopabate in feeds.

A high pressure liquid chromatographic (HPLC) method is described for determining ethopabate in poultry feeds. Feed samples containing ethopabate are finely ground, extracted 30 min by sonification with methanol-water (80 + 20), and filtered. The extract is cleaned up on an alumina column, and the first 4 ml eluate collected is analyzed. The drug is eluted through a muBondapak C18 column with acetonitrile-water (30 + 70) at a flow rate of 1.4 ml/min and detected by ultraviolet absorption at 280 nm. Chromatography is complete in 7 min, and detector response is linear. The drug is quantitated by peak height ratios. The procedure described is reproducible and shows good agreement with the official AOAC colorimetric method. The lower limit of detection is 2 ng by HPLC, making the method applicable to residue analyses.

Aminobenzoates↗

Gas-liquid chromatographic determination of ethopabate in feed premixes.

A gas-liquid chromatographic method is presented for determining ethopabate in 0.8 and 8.0% premixes. A sample is extracted with tetrahydrofuran containing an internal standard, by sonication or overnight soaking. The extract is clarified by centrifugation, diluted if necessary, and injected into a gas chromatograph equipped with a flame ionization detector. Average per cent recoveries for spiked blank samples were 100.6 at the 0.8% level and 100.4 at the 8.0% level. Precision, as indicated by replicate analyses of several premixes, ranged from 0.5 to 1.7% relative standard deviation.

Aminobenzoates↗

Sample preparation of carbadox, furazolidone, nitrofurazone, and ethopabate in medicated feeds for high pressure liquid chromatography.

Medicated feeds (pelleted or mash) containing guarantees of carbadox, furazolidone, nitrofurazone, and ethopabate are pretreated with water, extracted with 95% dimethylformamide overnight at room temperature, cleaned up on a column of alumina, and injected into a high pressure liquid chromatograph for quantitative measurement. Carbadox, nitrofurazone, and furazolidone can be separated; chromatograms show excellent baseline resolution, and results are in good agreement with colorimetric methods. The same extraction and cleanup can be used to improve colorimetric methods for furazolidone and nitrofurazone.

Aminobenzoates↗

Effect of feed additive antibiotics on chickens infected with Eimeria tenella.

Six experiments were carried out to study the effect of feed additive antibiotics on body weight, mortality and oocyst production in chickens infected with coccidiostat-susceptible or -resistant strain of E. tenella. Levels of antibiotic and coccidiostat in fed (mg./kg.) were: thiopeptin, 2; zinc bacitracin, 20; penicillin, 12; chlortetracycline, 22; amprolium plus ethopabate, 125 plus 8; clopidol, 125. All experiments included 7 groups; 2 of these groups were uninfected and infected controls, and the 5 remaining groups were all infected and given diet containing antibiotic, amprolium plus ethopabate, antibiotic and amprolium plus ethopabate, clopidol, or antibiotic and clopidol. Chickens in each group were fed respective diet beginning one day prior to coccidia exposure. In two experiments, infection with a coccidiostat-susceptible strain resulted in severe clinical coccidiosis in chickens on the basal ration and on thiopeptin-diet, but dietary thiopeptin prompted recovery of body weight. In one experiment where chickens were infected with a strain resistant to amprolium plus ethopabate and clopidol, birds on dietary thiopeptin attained higher body weight than birds on the basal ration. In three experiments when a strain resistant to amprolium plus ethopabate was inoculated, birds given the basal ration, bacitracin, penicillin, chlortetracycline, or amprolium plus ethopabate diet developed cecal coccidiosis. Chickens on ration containing antibiotic alone attained higher body weight than chickens on the basal ration. Combination of antibiotic and amprolium plus ethopabate resulted in higher weight attained than amprolium plus ethopabate alone. Clopidol suppressed development of coccidiosis, and the combination of antibiotic and clopidol resulted in higher gains than in clopidol alone.

Aminobenzoates↗

[Effectiveness of a number of anticoccidial agents. A brief survey taken in the field (author's transl)].

A floor pen trial was carried out on broilers in 1975 to compare the anticoccidial efficacy of monensin1), 3,5 dinitro-o-toluamide2), clopidol3), amprolium + ethopabacte4), amprolium + ethopabate + sulfaquinoxaline + pyrimethamine5) and robenidine6) in the presence of untreated controls. 4,200 Hybro chicks were distributed over twenty-eight pens, each initially housing 150 birds. Four pen replicates were allocated to each of the seven treatments. With the exception of robenidine, all agents used resulted in a statistically significant (P less than or equal to 0,05) gain in final weight compared with the untreated controls. The feed conversion rate showed significant (P less than or equal to 0,05) improvement in the groups treated with monensin, robenidine, amprolium + ethopabate.

Amprolium↗

Effect of dietary antibiotics on chickens infected with Eimeria tenella.

Two experiments were performed to study the effect of dietary antibiotics on percent daily weight change, mortality and gross cecal pathology in chickens during the critical phase of Eimeria tenella infection. In the first experiment, chickens were continuously fed ration containing thiopeptin, 2 mg/kg.; bacitracin, 20 mg./kg.; penicillin, 12 mg./kg.; or chlortetracycline, 22 mg./kg. One day after antibiotic fed was given, each bird received an oral inoculation of 30,000 sporulated oocyts. In the second experiment, chickens were consecutively fed ration containing amprolium plus ethopabate, 125 plus 8 mg./kg., and a combination of the coccidiostat and one of 4 antibiotics; thiopeptin, bacitracin, penicillin, or chloretracycline. One day after medicated feed was given, birds were each given an oral inoculation of 30,000 amprolium plus ethopabate-resistant E. tenella oocysts. The experiments were terminated 7 days after coccidia exposure. In both experiments, E. tenella infection resulted in depression in all birds of infected groups. Average percent weight change of infected birds was significantly lower than that of uninfected unmedicated control between 4 and 5 days after infection. Significantly greater number of birds died of cecal coccidiosis in group fed dietary bacitracin than that of other infected groups. Dietary antibiotics did not reduce gross cecal lesions.

Administration, Oral↗