Determination of free D-proline and D-leucine in the brains of mutant mice lacking D-amino acid oxidase activity
Determination of free D-proline and D-leucine in the brains of mutant mice lacking D-amino acid oxidase activity
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DOI：
10.1006/abio.2001.5382
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发表时间：
2001-11-15
影响因子：
2.9
通讯作者：
Zaitsu, K
中科院分区：
文献类型：
--
作者：
Hamase, K;Inoue, T;Zaitsu, K
建立了一种准确测定生物样品中微量d -脯氨酸的新方法。该d -氨基酸由4-氟-7-硝基-2,1,3-苯并恶二唑衍生，采用微ods柱和手性柱相结合的柱切换高效液相色谱系统测定。d -脯氨酸在小鼠大脑样品中的检出限为1 fmol（注射量，S/N = 3）。加标d -脯氨酸（10 fmol）的日内精密度为2.14%，日精密度为5.35% （RSD）。采用新方法测定了缺乏d -氨基酸氧化酶活性的突变型ddY/DAO（-）小鼠和对照型ddY/DAO（+）小鼠8个脑区和血清中游离d -脯氨酸的含量。还研究了游离d -亮氨酸的含量。d -脯氨酸在ddY/DAO（+）小鼠和ddY/DAO-小鼠脑内的含量和分布几乎相同，在垂体和松果体中均有较高的d -脯氨酸含量。另一方面，d -亮氨酸的含量在两个菌株之间是不同的。在ddY/DAO（+）小鼠的大脑中，与其他区域相比，松果体中观察到相对较高的d -亮氨酸。在ddY/DAO（-）小鼠大脑中，d -亮氨酸含量约为ddY/DAO（+）小鼠的10倍，且未见区域差异，而l -脯氨酸和l -亮氨酸含量在两菌株之间无显著差异。在血清中，ddY/DAO-小鼠游离d -脯氨酸和d -亮氨酸的含量明显高于ddY/DAO（+）小鼠。(C) 2001学术出版社。
A new procedure to accurately measure a trace amount Of D-proline in biological samples has been developed. This D-amino acid was derivatized with 4-fluoro-7-nitro-2,1,3-benzoxadiazole and was determined by a column-switching HPLC system, a combination of a micro-ODS column and a chiral column. The detection limit for D-proline spiked in a mouse cerebrum sample is 1 fmol (injection amount, S/N = 3). Within-day precision and day-to-day precision obtained for spiked D-proline (10 fmol) are 2.14 and 5.35% (RSD), respectively. Using the new method, the amount of free D-proline in eight brain regions and sera of mutant ddY/DAO(-) mice, lacking D-amino acid oxidase activity, and control ddY/DAO(+) mice was determined. The amount of free D-leucine was also investigated. The amount and distribution Of D-proline in the brains of ddY/DAO(+) mice and ddY/DAO- mice are almost the same, and relatively high amounts Of D-proline have been observed in the pituitary gland and in the pineal gland. On the other hand, the amount Of D-leucine is different between the two strains. In the brains of ddY/DAO(+) mice, a relatively high amount Of D-leucine has been observed in the pineal gland compared with other regions. In the brains of ddY/DAO(-) mice, D-leucine amounts are approximately 10 times higher than those obtained in ddY/DAO(+) mice and regional difference has not been observed, while the amounts of L-proline and L-leucine are not significantly different between the two strains. In the serum, the amounts of both free D-proline and D-leucine are significantly higher in the ddY/DAO- mice than those obtained in ddY/DAO(+) mice. (C) 2001 Academic Press.