Amlodipine besylate

From 6 weeks of age, rats were weighed once a week and the average water intake was measured per cage. As from this age, blood pressure and heart rate were measured biweekly in conscious rats using a non-invasive tail-cuff system (Kent Scientific). Rats were first accustomed to this procedure by placing them in the restrainer on 4 consecutive days. Ten blood pressure and heart rate measurements were recorded from each rat per measurement session and averaged. From 9 weeks of age, treatment with the antihypertensive agents was initiated. Amlodipine besylate (5 mg/kg/day, Pfizer) and atenolol (50 mg/kg/day, Sigma-Aldrich) were dissolved in the drinking water. As a stable effect of the antihypertensive agents was observed after 7 weeks of treatment, blood pressure and heart rate were measured monthly from this time point on. Animals were treated until 55.1 ± 0.2 weeks of age.

Amlodipine besylate was purchased from Pfizer (Karlsruhe, Germany); canrenone from the European Directorate for the Quality of Medicines & HealthCare (Strasbourg, France); bisoprolol fumarate, metoprolol tartrate, carvedilol, candesartan, valsartan, olmesartan, torasemide, and a diazepam-d5 solution (1 mg/mL in methanol) from LGC Standards (Wesel, Germany); atorvastatin-d5 calcium salt from Alsachim (Shimadzu Group, Illkirch Graffenstaden, France); and methanol, acetonitrile (ACN), diethyl ether, ethyl acetate, and natrium sulfate from VWR International GmbH (Darmstadt, Germany), as well as ammonium formate and formic acid from Merck KGaA (Darmstadt, Germany). All chemicals were of analytical grade. Water was purified with a Millipore filtration unit (18.2 Ω × cm water resistance) from Merck (Darmstadt, Germany). Drug-free pooled human blank plasma was obtained from a local blood bank. The analysis of human samples was approved by the local ethics committee to assess medication adherence using toxicological analyses objectively and complied with the Declaration of Helsinki. All patients provided written informed consent.

In this study, the following major chemicals were used: L-NAME was obtained from Meilun Biotechnology Co., Ltd, Dalian, China, while amlodipine besylate was purchased from Pfizer, Dalian, China.