Optimization, validation and application of an assay for the activity of HMG-CoA reductase in vitro by LC-MS/MS
10.1016/j.jpha.2015.06.002
- Author:
Jing WANG
;
Jiye SUN
;
Chunjie SHA
;
Yufeng SHAO
;
Yanhong LIU
;
Youxin LI
;
Zhenwen DUAN
;
Wanhui LIU
- Publication Type:Journal Article
- Keywords:
Xuezhikang;
LC-MS/MS;
HMG-CoA reductase;
Mevalonolactone;
Quality control
- From:
Journal of Pharmaceutical Analysis
2015;(6):383-388
- CountryChina
- Language:Chinese
-
Abstract:
A stable HMG-CoA reductase (HMGR) reaction in vitro was developed by a sensitive, selective and precise liquid chromatography–tandem mass spectrometry (LC–MS/MS) method. The optimized enzyme reac-tion condition contained 1.5μg of HMGR, 20 nM of NADPH with 50 min of reaction time. The method was validated by several intra-and inter-day assays. The production transitions of m/z 147.0/59.1 and m/z 154.0/59.1 were used to detect and quantify mevalonolactone (MVAL) and MVAL-D7, respectively. The accuracy and precision of the method were evaluated over the concentration range of 0.005–1.000μg/mL for MVAL and 0.010–0.500μg/mL for lovastatin acid in three validation batch runs. The lower limit of quantitation was found to be 0.005μg/mL for MVAL and 0.010μg/mL for lovastatin acid. Intra-day and inter-day precision ranged from 0.95%to 2.39%and 2.26%to 3.38%for MVAL, 1.46%to 3.89%and 0.57% to 5.10% for lovastatin acid, respectively. The results showed that the active ingredients in Xuezhikang capsules were 12.2 and 14.5 mg/g, respectively. This assay method could be successfully applied to the quality control study of Xuezhikang capsule for the first time.
