pyrimidinones and 4-4--dinitrocarbanilide

The development, optimization and validation of an ion-pairing high performance liquid chromatography method for the simultaneous determination of both nicarbazin (NIC) components: 4,4'-dinitrocarbanilide (DNC) and 2-hydroxy-4,6-dimethylpyrimidine (HDP) in bulk materials and feed additives are described. An experimental design was used for the optimization of the chromatographic system. Four variables, including mobile phase composition and oven temperature, were analyzed through a central composite design exploring their contribution to analyte separation. Five responses: peak resolutions, HDP capacity factor, HDP tailing and analysis time, were modelled by using the response surface methodology and were optimized simultaneously by implementing the desirability function. The optimum conditions resulted in a mobile phase consisting of 10.0 mmol L(-1) of 1-heptanesulfonate, 20.0 mmol L(-1) of sodium acetate, pH=3.30 buffer and acetonitrile in a gradient system at a flow rate of 1.00 mL min(-1). Column was an INERSTIL ODS-3 (4.6 mm×150 mm, 5 μm particle size) at 40.0°C. Detection was performed at 300 nm by a diode array detector. The validation results of the method indicated a high selectivity and good precision characteristics, with RSD less than 1.0% for both components, both in intra and inter-assay precision studies. Linearity was proved for a range of 32.0-50.0 μg mL(-1) of NIC in sample solution. The recovery, studied at three different fortification levels, varied from 98.0 to 101.4 for HDP and from 99.1 to 100.2 for DNC. The applicability of the method was demonstrated by determining DNC and HDP content in raw materials and commercial formulations used for coccidiosis prevention. Assays results on real samples showed that considerable differences in molecular ratio DNC:HDP exist among them.

Topics: Animal Feed; Animals; Carbanilides; Chromatography, High Pressure Liquid; Coccidiosis; Coccidiostats; Nicarbazin; Pyrimidinones

Nicarbazin is being investigated as an infertility agent for the control of non-migratory Canada geese (Branta canadensis L) populations. Nicarbazin is presently registered for use as a coccidiostat for poultry. Geese fed sufficient quantities of nicarbazin will lay non-viable eggs. We established nicarbazin consumption by measuring the concentration of a component of the formulation, 4,4'-dinitrocarbanilide (DNC) in the egg contents (yolk, albumin) in non-viable eggs. To estimate the nicarbazin consumption of birds that laid viable eggs (eggs that hatched or contained an embryo), a high-performance liquid chromatography method was developed to measure the concentration of DNC in egg shells. A statistically significant correlation was established using linear regression between the mean concentrations of DNC in the egg shell and in the egg contents in non-viable eggs. Viable eggs were estimated to contain lower levels of DNC than non-viable eggs. DNC concentrations in both the egg contents and the egg shell increased with increases in nicarbazin dose in feed. Our method allows for the estimation of nicarbazin consumption and DNC dose in eggs under field conditions, which is important in developing an effective infertility agent for over-abundant non-migratory goose populations.

Topics: Animal Feed; Animals; Carbanilides; Dose-Response Relationship, Drug; Egg Shell; Fertility; Geese; Nicarbazin; Pyrimidinones