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目的建立同时测定血浆腐胺、精胺、亚精胺含量的柱前衍生化-HPLC法,并用于系统性红斑狼疮患者和正常人群血浆浓度测定,指导临床实践。方法 32例系统性红斑狼疮患者和30例健康志愿者血浆样品离心后,经丹磺酰氯衍生化和甲苯萃取,以C_(18)(250 mm×4.6 mm,5μm)柱为固定相,乙腈-0.2 mol·L~(-1)甲酸铵为流动相,经梯度洗脱,于272 mm波长处检测。结果腐胺、亚精胺质量浓度在0.2~40 mg·L~(-1)、精胺在0.2~30 mg·L~(-1)内呈现良好线性关系,腐胺、亚精胺最低定量限均为0.1 mg·L~(-1)、精胺为0.2 mg·L~(-1),腐胺、亚精胺最低检测限均为0.05 mg·L~(-1)、精胺为0.1 mg·L~(-1)。正常人群血浆腐胺浓度平均为1.570 mg·L~(-1)、精胺0.603 mg·L~(-1)、亚精胺1.710 mg·L~(-1)。结论建立了同时快速测定血浆中腐胺、精胺、亚精胺的柱前衍生化-HPLC法,方法灵敏、稳定、重现性好、选择性好、准确度高,可用于准确测定人血浆中多胺含量,并作为相关疾病诊断和治疗的重要参考指标。
OBJECTIVE To establish a pre-column derivatization-HPLC method for the simultaneous determination of plasma putrescine, spermine and spermidine in patients with systemic lupus erythematosus and normal subjects, and to guide the clinical practice. Methods Thirty - two patients with systemic lupus erythematosus (SLE) and 30 healthy volunteers were centrifuged and derivatized with Dansyl chloride and extracted with toluene. The stationary phase was C 18 (250 mm × 4.6 mm, 5 μm) 0.2 mol·L -1 ammonium formate as mobile phase, gradient elution, detected at 272 mm wavelength. Results The concentrations of putrescine and spermidine were in the range of 0.2-40 mg · L -1, and spermine showed a good linearity in the range of 0.2-30 mg · L -1. The contents of putrescine and spermidine were the lowest The limits of detection were 0.1 mg · L -1 and 0.2 mg · L -1 spermine respectively. The detection limits of putrescine and spermidine were both 0.05 mg · L -1, 0.1 mg · L -1. The average plasma putrescine concentration in the normal population was 1.570 mg · L -1, spermine 0.603 mg · L -1, and spermidine 1.710 mg · L -1. Conclusion The method for rapid and simultaneous determination of putrescine, spermine and spermidine in plasma by HPLC was established. The proposed method was sensitive, stable and reproducible, with good selectivity and high accuracy. It could be used to determine the plasma concentration of human plasma Polyamine content, and as an important reference diagnosis and treatment of related diseases.