液相色谱-质谱法测定唾液内的褪黑素浓度

Determination concentration of melatonin in human saliva by HPLC-MS/MS

目的建立唾液中褪黑素(镇静催眠药)浓度的HPLC-MS/MS测定方法。方法唾液样品经碱化后,用二氯甲烷萃取,以伊托必利作内标进样。色谱柱用Sunfire C18,柱温35℃;流动相为0.1%甲酸水溶液-甲醇,梯度洗脱,流量为0.3mL·min-1;质谱用ESI离子源。结果本方法的定量下限是5.48pg·mL-1(S/N>10),线性范围为5.48～250.00pg·mL-1(γ2=0.9994);最低检测限为2pg·mL-1(S/N>3);本方法的日内、日间RSD均小于15%,提取回收率约为60%。结论本研究建立的褪黑素唾液浓度的测定方法快速、准确、灵敏度高。

Objective To establish a HPLC - MS/MS method for the determination of melatonin in human saliva. Methods Saliva samples and internal standard （itopride） were extracted by dichloromethane after alkalization. The analysis was conducted using a Sunfire C18 column with gradient elution and the column temperature is 35 ℃. The mobile phase consisted of methanol and water with 0. 1% formic acid at flow rate of 0.3 mL . min- 1 Analysis in mass was operated by the electrospray ionization. Results LLOQ of the method is 5.48 pg . mL-1 （ S/N 〉 10）. The standard curve is linear in the range of 5.48 -250.00 pg . mL-1 （y2 = 0. 9994）. LLOD is 2 pg . mL- 1 （ S/N 〉 3 ）. The RSDs of inter - day and intra - day are all less than 15%. The extraction recovery is closed to 60%. Conclusion The method is fast, precise and sensitive, and suitable for the determination of melatonin in human saliva.