Ureal cobas c 311

On the last day of the trial period pigs were submitted to a 14 h fasting prior blood sampling. Blood was collected via venipuncture in the orbital sinus using needles (40 × 1.6 mm) and placed in 10 mL uncoated vacuum tubes. Samples were immediately sent to the Viçosa Clinical Laboratory (Viçosa, MG, Brazil) to measure the concentration of serum urea nitrogen (SUN), triglycerides (TG) and total cholesterol (CHO) (Ureal Cobas c 311, Roche Diagnostics GmbH, Basel, Switzerland).

At 110 days of gestation, sows were weighed and blood samples were collected in two 10 mL tubes (Becton Dickinson Vacutainer Systems), one with silicone and the other with EDTA (ethylenediamine tetraacetic acid) to obtain serum and plasma, respectively. One hour after collection, blood samples were centrifuged (Eppendorf^® 5702R, Eppendorf AG, Hamburg, Germany) at 3000× g for 10 min at 4 °C, allowing the removal of serum and plasma, which were transferred to 1.5 mL microtubes and stored at −20 °C until further analysis.
Serum samples were used to determine the concentrations of urea (Ureal Cobas c 311; Roche Diagnostics GmbH, Basel, Switzerland) and insulin (Kit Architect Insulina; Abbott Laboratórios do Brasil Ltd.a., São Paulo, Brazil). Plasma samples were used to determine estradiol concentration using microparticle chemiluminescence immunoassay (Architect estradiol assay, Abbott, IL, USA).