Imai K, Matsunaga Y, Tsukamoto Y, Nishitani A
University of Tokyo, Branch Hospital Pharmacy, Tokyo, Japan.
J Chromatogr. 1987 Jul 29;400:169-76. doi: 10.1016/s0021-9673(01)81610-1.
Bis[4-nitro-2-(3,6,9-trioxadecyloxycarbonyl)phenyl] oxalate (TDPO) was used to examine the peroxyoxalate chemiluminescence (CL) reaction for the detection of fluorescent compounds. Some fluorescent compounds (perylene, eosine, rhodamine, Rose Bengal, fluorescein and umbelliferone) gave higher CL intensities as the proportion of water in the reaction medium increased to ca. 40%, whereas dansylalanine, 8-anilinonaphthalene-1-sulphonic acid, 7-nitrobenzo-2-oxa-1,3-diazol-4-ylproline and dihydronicotinamide adenine dinucleotide gave opposite results. The effects of temperature and time on the post-column reaction in reversed-phase high-performance liquid chromatography (HPLC) were investigated. Under the optimal conditions, the detection limit for dansylamino acids was at the sub-femtomole level. The advantage of using TDPO in HPLC is its stability in the presence of hydrogen peroxide [ca. 10% loss of activity per 8 h vs. 60% per 8 h for bis(2,4,6-trichlorophenyl) oxalate].
双[4-硝基-2-(3,6,9-三氧杂癸氧基羰基)苯基]草酸酯(TDPO)用于检测荧光化合物的过氧草酸酯化学发光(CL)反应。随着反应介质中水的比例增加到约40%，一些荧光化合物(苝、曙红、罗丹明、孟加拉玫瑰红、荧光素和伞形酮)产生更高的CL强度，而丹磺酰丙氨酸、8-苯胺基萘-1-磺酸、7-硝基苯并-2-恶唑-1,3-二氮杂-4-基脯氨酸和二氢烟酰胺腺嘌呤二核苷酸则给出相反的结果。研究了温度和时间对反相高效液相色谱(HPLC)柱后反应的影响。在最佳条件下，丹磺酰氨基酸的检测限在亚飞摩尔水平。在HPLC中使用TDPO的优点是其在过氧化氢存在下的稳定性[每8小时活性损失约10%，而双(2,4,6-三氯苯基)草酸酯每8小时损失60%]。
