Lithium heparin coated vacutainer tubes

Racemic (rac) PZQ was obtained from Sigma-Aldrich (Buchs, Switzerland). PZQ enantiomers as well as the metabolite trans-4-OH were donated by Merck Serono (Darmstadt, Germany). Eleven-fold deuterized PZQ (PZQd11, internal standard-IS) was acquired from Toronto Research Chemicals (Ontario, Canada). The chemical structures of PZQ, PZQd11 and trans-4-OH are depicted in Fig 1. Acetonitrile, ethanol and methanol of MS grade were purchased from Carl Roth GmBH (Allschwil, Switzerland), and ammonium formate, ammonium acetate and formic acid of MS grade from Sigma-Aldrich (Buchs, Switzerland). Ultrapure water was provided using a Millipore Milli-Q water purification system (Merck Millipore, Darmstadt, Germany). Blank human plasma and blood were supplied in lithium heparin-coated vacutainer tubes (BD, Allschwil, Switzerland) from the local blood donation centre (Basel, Switzerland).

Blood samples were collected at 1600 h on days four and nine post-estrus via jugular venipuncture into lithium heparin-coated vacutainer tubes (BD (Becton, Dickinson and Company) precision glide needle 18 g × 1.5 inch, BD, North Ryde, NSW, Australia; BD Vacutainer Lithium Heparin Blood Collection Tubes, 9 mL, BD, North Ryde, NSW, Australia) and stored on ice until processing. Samples were centrifuged at 3000 rpm for 10 min before the plasma was pipetted into Eppendorf tubes and stored at −20 °C. Blood plasma progesterone was determined in the Adelaide Research Assay Facility by coated tube radioimmunoassay (IM1188; Beckman Coulter, Brea, CA, USA). The limit of detection was 0.08 ng/mL and the mean intra-assay coefficient of variation was 6.9%. Pregnancy rate, which was the number of sows that were pregnant determined using ultrasonography at day 28 of gestation, was also recorded.