Chemical degradation of 3H-labeled substance P in tris buffer solution.
Chemical degradation of 3H-labeled substance P in tris buffer solution.
复制标题
Tris 缓冲溶液中 3H 标记的 P 物质的化学降解。
DOI：
10.1016/0003-2697(88)90215-1
复制
发表时间：
1988
影响因子：
2.9
通讯作者：
Desiderio,DM
中科院分区：
文献类型：
--
作者：
Higa,T;Desiderio,DM
Substance P (SP) is an important neuropeptide that has been implicated in several physiological processes, and it is necessary to devise an analytical procedure to measure endogenous SP with a combination of high sensitivity and maximum molecular specificity. However, the unique chemical nature of SP (polarity, chemical stability, ease of oxidation, peptide bond lability) plays a significant role in its analysis, such as in receptor assays, immunoassays, chromatography, and mass spectrometry. In this study, we evaluated in polypropylene and glass assay tubes the effects on the recovery and stability of tritiated SP ([3H]SP) of several pertinent experimental parameters such as buffer, pH, multiple freeze-thaw cycles, and incubation temperature and time. Bovine serum albumin (BSA) effectively reduced the absorption of [3H]SP to polypropylene and glass tube surfaces. Following multiple (6×) freeze-thaw cycles of solutions in BSA-precoated tubes, the recovery of radioactive [3H]SP remained high (>75%) after the last cycle, whereas recovery was minimal in uncoated or siliconized glass tubes. A high level of radioactivity recovery was maintained for 14 days of storage of [3H]SP in triethylamine formate (TEAF) solution in BSA-precoated tubes at 4 and −20°C, but decreased at 37°C to less than 80% in only 3 h. Following storage in Tris-HCl (pH 7.4) buffer, a combination of HPLC and mass spectrometric analyses revealed that a significant amount of peptide bond cleavage occurred to produce the two peptides ArgProLys (RPK) and ArgProLysProGlnGln (RPKPQQ), with only a small amount of remaining intact SP. That decomposition was not observed in triethylamine formate TEAF (pH 3.14) buffer solutions.
