Gupta R N, Whelton C
Department of Laboratory Medicine, St. Joseph's Hospital, Hamilton, Ontario, Canada.
J Chromatogr. 1992 Nov 6;582(1-2):236-41. doi: 10.1016/0378-4347(92)80325-k.
We describe a simple method for extracting homovanillic acid (HVA) from plasma. An aliquot of 0.5 ml of the internal standard solution (3-hydroxy-4-methoxycinnamic acid in 0.2 mol/l phosphoric acid) and 0.5 ml of the sample are applied to a 1-ml Bond Elut C18 column prewashed with methanol and 0.2 mol/l phosphoric acid. The sample is drawn through the column at low speed. The column is washed with water and eluted with dichloromethane. The eluate is evaporated under vacuum at ambient temperature and the residue reconstituted with 250 microliters of the mobile phase. A 10-microliters aliquot of the resulting solution is injected onto a 150 mm x 4.6 mm I.D. column packed with 5-microns octadecylsilyl silica particles (Beckman). Peaks are detected coulometrically in the screening-oxidation mode with E1 = +0.25 V and E2 = +0.38 V. In the resulting chromatogram, HVA and the internal standard give sharp peaks and are well separated from solvent and other endogenous electroactive acids. The extraction recovery is 90-95% which allows the determination of 0.5 microgram/l analyte.
我们描述了一种从血浆中提取高香草酸（HVA）的简单方法。将0.5 ml内标溶液（0.2 mol/l磷酸中的3 - 羟基 - 4 - 甲氧基肉桂酸）和0.5 ml样品加入到预先用甲醇和0.2 mol/l磷酸洗涤过的1 - ml Bond Elut C18柱中。样品以低速通过柱子。柱子用水洗涤，然后用二氯甲烷洗脱。洗脱液在室温下真空蒸发，残留物用250微升流动相复溶。取10微升所得溶液注入内径为150 mm×4.6 mm、填充有5微米十八烷基硅烷硅胶颗粒（贝克曼）的柱子中。在筛选氧化模式下，以E1 = +0.25 V和E2 = +0.38 V进行库仑检测。在所得色谱图中，HVA和内标给出尖锐的峰，并且与溶剂和其他内源性电活性酸很好地分离。提取回收率为90 - 95%，这使得能够测定0.5微克/升的分析物。
