Abstract: Objective To establish a liquid chromatography tandem mass spectrometry (LC-MS/MS) method to determine cortisol levels in the plasma of cynomolgus monkeys and evaluate the influence of transportation, fasting and chair restraint on its levels in plasma. Methods Seven male monkeys were transported for eight hours by airplane and truck, and the blood samples were collected on day 1, 7, 14, 20 and 28 after transportation. The blood samples in six male monkeys were collected after the animals were fasted for 0, 8, 16 and 24 hours, and after one or five times of two-hour chair restraint. Cortisol was determined by using LC-MS/MS. Results The cortisol levels in plasma samples increased significantly after transportation, and then decreased gradually, and reached a stable level three weeks later. The cortisol levels in plasma samples were significantly increased after 8, 16 and 24 hours of fasting, but no differences were observed among the three types of fasting. The cortisol levels in plasma samples increased significantly after one and five times of chair restraint, but the increased extent of the latter was lower than the former. The calibration curves were linear over the concentration range of 20.0-2000.0 ng/ml. The intra- and inter-day precision was less than 4.94% and 5.69%, respectively, and the accuracy (RE) was less than 10.04% and 5.69%, respectively. The extraction recovery was between 70.3% and 74.7%. The analyte proved to be stable within three freeze-thaw tests and in the autosampler for ten hours at 20℃. Conclusion This method is rapid and specific and suited to cortisol determination in plasma. All of the stimuluses such as transportation, fasting and chair restraint were stressful for the monkeys.

Key words: Cynomolgus monkey, Stress, Cortisol, LC-MS/MS