Determination of total cholesterol in serum by high-performance liquid chromatography with electrochemical detection
Determination of total cholesterol in serum by high-performance liquid chromatography with electrochemical detection
复制标题
DOI：
10.1016/j.chroma.2007.08.020
复制
发表时间：
2007-09-28
影响因子：
4.1
通讯作者：
Kusu, Fumiyo
中科院分区：
文献类型：
--
作者：
Hojo, Kazuhiro;Hakamata, Hideki;Kusu, Fumiyo
建立了一种不需衍生化、简便、灵敏的测定胆固醇的高效液相色谱法。胆固醇的伏安行为研究表明，胆固醇在非水溶剂中可在玻碳电极上被氧化。这是适用于开发的方法，通过电化学检测（HPLC-ED）的HPLC。优化了HPLC-ED分离胆固醇和氧化甾醇的条件，包括26-羟基胆固醇和24 S-羟基胆固醇。分离用SiO2 Osil C30-UG-3柱;含有50 mM LiClO 4的乙腈-2-丙醇（9：1，v/v）作为移动的相;和相对于Ag/AgCl的1.9 V的施加电位进行。在0.5 ~ 100 μ M范围内，电流峰高与胆固醇的进样量呈线性关系（r > 0.999）。胆固醇的检测限（S/N = 3）为0.36 μ M（1.8 pmol）。胆固醇在100 μ M直接检测的相对标准偏差（RSD）小于1.0%（n = 8）。用本法测定了正常人血清中的总胆固醇和游离胆固醇，回收率大于90%，相对标准偏差（n = 6）小于3.0%。(c)2007 Elsevier B. V.保留所有权利。
A simple and sensitive HPLC method that does not require derivatization for determining cholesterol has been developed. Investigation of voltammetric behavior of cholesterol showed that cholesterol could be oxidized on a glassy carbon electrode in non-aqueous solvents. This was applied to the development of a method by HPLC with electrochemical detection (HPLC-ED). The HPLC-ED was optimized using the separation of cholesterol and oxysterols including 26-hydroxycholesterol and 24S-hydroxycholesterol. The separation was carried out with a Develosil C30-UG-3 column; acetonitrile-2-propanol (9: 1, v/v) containing 50 mM LiClO4 as a mobile phase; and an applied potential at 1.9 V versus Ag/AgCl. The current peak height was linearly related to the amount of cholesterol injected from 0.5-100 mu M (r > 0.999). The detection limit (S/N = 3) of cholesterol was 0.36 mu M (1.8 pmol). Cholesterol at 100 mu M was directly detected with a relative standard deviation (RSD) of less than 1.0% (n = 8). Total cholesterol and free cholesterol in control human serum were determined by the present method with the recovery of more than 90% and the RSD (n = 6) of less than 3.0%. (c) 2007 Elsevier B.V. All rights reserved.