Faculty of Pharmacy and Pharmaceutical Sciences, Fukuyama University, Fukuyama, Hiroshima 729-0292, Japan.
J Chromatogr B Analyt Technol Biomed Life Sci. 2010 Feb 1;878(3-4):327-32. doi: 10.1016/j.jchromb.2009.11.038. Epub 2009 Dec 3.
A simple and highly sensitive high-performance liquid chromatography procedure was developed for the determination of carnosine in urine. Carnosine was derivatized with 4-(5,6-dimethoxy-2-phthalimidinyl)-2-methoxyphenylsulfonyl chloride at 70 degrees C for 15 min in borate buffer (20 mmol l(-1), pH 9.0) to produce fluorescent sulfonamides. After hydrolysis of the reaction mixture with formic acid at 100 degrees C for 15 min, the fluorescent derivative of carnosine was separated on a reversed-phase column with a linear gradient elution using solvents of (A) acetate buffer (0.1 mmol l(-1), pH 7.0) and (B) acetonitrile at a flow-rate of 1.0 ml/min and was detected at excitation and emission wavelengths of 318 and 400 nm, respectively. The detection limit of carnosine was 4 fmol at a signal-to-noise ratio of 3. The within-day and day-to-day relative standard deviations were 2.7-4.6% and 0.4-5.2%, respectively. The concentration of carnosine in normal human urine was found to be 4.6-125 nmol (mg creatinine)(-1) (mean+/-SD: 21.6+/-26.6 nmol (mg creatinine)(-1), n=20).
建立了一种简单、灵敏的高效液相色谱法，用于测定尿中的肌肽。肌肽在硼酸缓冲液（20mmol/L，pH9.0）中于 70°C 下与 4-(5,6-二甲氧基-2-邻苯二甲酰亚氨基) -2-甲氧基苯磺酰氯衍生化 15 分钟，生成荧光磺酰胺。将反应混合物在 100°C 下用甲酸水解 15 分钟后，用线性梯度洗脱，用（A）乙酸盐缓冲液（0.1mmol/L，pH7.0）和（B）乙腈作为溶剂，在 1.0ml/min 的流速下分离肌肽的荧光衍生物，并分别在激发和发射波长为 318nm 和 400nm 处检测。肌肽的检测限为 4fmol，信噪比为 3。日内和日间相对标准偏差分别为 2.7-4.6%和 0.4-5.2%。正常人体尿液中的肌肽浓度为 4.6-125nmol（mg 肌酐）（-1）（均值+/-SD：21.6+/-26.6nmol（mg 肌酐）（-1），n=20）。
