Maestri L, Ghittori S, Imbriani M, Capodaglio E
Centro Richerche di Fisiopatologia e Sicurezza del Lavoro, University of Pavia, Clinica del Lavoro Foundation, IRCCS, Italy.
J Chromatogr B Biomed Appl. 1994 Jul 1;657(1):111-7. doi: 10.1016/0378-4347(94)80076-6.
A sensitive method for the determination of free and total urinary 2,5-hexandione (2,5-HD) using high-performance liquid chromatography with fluorescence detection was developed. After purification of urine with a disposable C18 cartridge, 2,5-HD was derivatized with dansylhydrazine; 1,3-diacetyl benzene (1,3-DAB) was added to the samples, as internal standard, prior to extraction. The resulting fluorescent adducts were separated on a reversed-phase column with a gradient mobile phase of 25 mM phosphate buffer (pH 6.4) and acetonitrile. The retention times of the 2,5-HD and 1,3-DAB derivatives were 9.4 and 13.7 min, respectively. The derivatives were detected by a fluorescence detector (excitation 340 nm, emission 525 nm). The mean recoveries of 2,5-HD and 1,3-DAB were 92.0 and 94.0%, respectively; the detection limit of 2,5-HD (signal-to-noise ratio of 3) was 5 micrograms/l in urine without hydrolysis and ca. 12 micrograms/l in hydrolyzed samples. The method was applied to 39 urine samples from workers exposed to n-hexane; the mean values were 2.597 mg/l (S.D. = +/- 0.758) for total 2,5-HD and 0.179 mg/l (S.D. = +/- 0.086) for free 2,5-HD. Urine samples of 22 non-exposed subjects showed a mean concentration of 0.437 mg/l (S.D. = +/- 0.109) and 0.022 mg/l (S.D. = +/- 0.011) for total and free 2,5-HD, respectively.
建立了一种采用高效液相色谱-荧光检测法测定游离和总尿2,5-己二酮（2,5-HD）的灵敏方法。用一次性C18柱净化尿液后，2,5-HD用丹磺酰肼衍生化；在萃取前，向样品中加入1,3-二乙酰苯（1,3-DAB）作为内标。所得荧光加合物在反相柱上用25 mM磷酸盐缓冲液（pH 6.4）和乙腈的梯度流动相进行分离。2,5-HD和1,3-DAB衍生物的保留时间分别为9.4分钟和13.7分钟。衍生物通过荧光检测器（激发波长340 nm，发射波长525 nm）进行检测。2,5-HD和1,3-DAB的平均回收率分别为92.0%和94.0%；2,5-HD的检测限（信噪比为3）在未水解的尿液中为5微克/升，在水解样品中约为12微克/升。该方法应用于39名正己烷暴露工人的尿液样本；总2,5-HD的平均值为2.597毫克/升（标准差=±0.758），游离2,5-HD的平均值为0.179毫克/升（标准差=±0.086）。22名未暴露受试者的尿液样本中，总2,5-HD和游离2,5-HD的平均浓度分别为0.437毫克/升（标准差=±0.109）和0.022毫克/升（标准差=±0.011）。
