QUANTITATION OF HYDROXYPYRIDINIUM CROSSLINKS IN COLLAGEN BY HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY
QUANTITATION OF HYDROXYPYRIDINIUM CROSSLINKS IN COLLAGEN BY HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY
复制标题
DOI：
10.1016/0003-2697(84)90101-5
复制
发表时间：
1984-01-01
影响因子：
2.9
通讯作者：
VANNESS, KP
中科院分区：
文献类型：
--
作者：
EYRE, DR;KOOB, TJ;VANNESS, KP
描述了一种定量测定胶原中3-羟基吡啶交联的高效液相色谱方法。它可以应用于所有类型结缔组织的粗水解物。矿化组织可以直接水解和分析，而不受矿物离子的干扰。在反相C18色谱柱上，以乙腈水溶液为梯度，以0.01 M正七氟丁酸为离子配对剂，分离羟基吡啶残基（HP）和赖基（LP）形式。用荧光光谱仪检测交联氨基酸的自然荧光，精确定量至2pm （1ng）注射。羟基吡啶交联丰富的组织包括各种形式的软骨、骨、牙本质、韧带、肌腱、筋膜、椎间盘、肺、肠、子宫颈、主动脉和玻璃体。在正常组织中，相对于HP，交联的次要形式LP仅在骨和牙本质中大量存在。这两种交联在皮肤、角膜、大鼠尾腱和基底膜中基本缺失。
An HPLC [high performance liquid chromatography] method for quantifying the 3-hydroxypyridinium crosslinks of collagen is described. It can be applied to crude hydrolysates of all types of connective tissue. Mineralized tissues can be hydrolyzed directly and analyzed without interference from the mineral ions. The hydroxylysyl (HP) and lysyl (LP) forms of hydroxypyridinium residue were resolved on a reverse-phase C18 column using a gradient of acetonitrile in water and 0.01 M n-heptafluorobutyric acid as an ion-pairing agent. The crosslinking amino acids were accurately quantified down to 2 pM (1 ng) injected, by detecting their natural fluorescence with a spectrofluorometer. Tissues in which hydroxypyridinium crosslinks were plentiful included all forms of cartilage, bone, dentin, ligament, tendon, fascia, intervertebral disc, lung, gut, cervix, aorta and vitreous humor. Among normal tissues, LP, the minor form of the crosslink, was present in significant amounts relative to HP only in bone and dentin. Both crosslinks were essentially absent from skin, cornea, rat tail tendon and basement membranes.