Use of sulfo-phospho-vanillin to quantitate unsaturated neutral lipids in thin-layer chromatography.
Use of sulfo-phospho-vanillin to quantitate unsaturated neutral lipids in thin-layer chromatography.
复制标题
使用磺基-磷酸-香草醛在薄层色谱中定量不饱和中性脂质。
DOI：
10.1016/0021-9673(75)80023-9
复制
发表时间：
1975
期刊：
影响因子：
--
通讯作者：
G. Dohm
中科院分区：
文献类型：
--
作者：
E. Tapscott;G. Dohm
材料和方法脂质标准品得自Sigma（St. Louis，MO，USA）点在通过将Adsorbosil I（CaS 〇 4粘合剂，Applied Science Labs.，宾夕法尼亚州州立学院，USA）至0.25 mm厚度。将制备的板风干并在100”下活化1.5小时。将样品交替点样在IO-cm泳道上，泳道之间留出1.5cm。首先将板在乙醚-苯-乙酸（40：50：1）中展开至原点以上7 cm，风干并在己烷-乙醚（9 G：4）中重新展开至顶部。这些溶剂系统提供了胆固醇、油酸、三油酸甘油酯、油酸甲酯和油酸胆固醇酯以该顺序的良好分离（图I）。将每个斑点及其相应的空白从板上刮入试管中，并加入1.0 ml浓硫酸。将混合物在沸水浴中加热10分钟，并在冷水中冷却5分钟。向样品中加入2.0 ml如前所述制备的磷酸-香草醛试剂。然后将混合物在37 ° C下孵育15分钟，并在800 g下离心5分钟以分离凝胶。在540 nm处读取吸光度。
MATERIALS AND METHODSLipid standards obtained from Sigma (St. Louis, MO., USA) were spotted on 20 x 20 cm plates prepared by spreading Adsorbosil I (CaS04 binder, Applied Science Labs., State College, Pa., USA) to 0.25-mm thickness. The prepared plates were air dried and activated at 100” for 1.5 h. Samples were spotted alternately on IO-cm lanes leaving 1.5 cm between lanes. Plates were first developed in diethyl ether-benzene-acetic acid (40: 50: 1) to 7 cm above the origin, air dried and redeveloped to the top in hexane-diethyl ether (9G: 4). These solvent systems afforded good separation of cholesterol, oleic acid, triolein, methyl oleate, and cholesteryl oleate in that order (Fig. I), Spots were visualized by spraying with Rhodamine 6G (0.001 y<:. w/v) in methanol.Each spot and its corresponding blank were scraped from the plate into test tubes and 1.0 ml concentrated sulfuric acid was added. The mixture was heated for IO min in a boiling wateribath and cooled for 5 min in cold water. To the sample were added 2.0 ml of phospho-vanillin reagent prepared as previously described’. The mixture was then incubated for 15 min at 37” and centrifuged for 5 min at 800 g to separate the gel. The absorbance was read at 540 nm.