2007
DOI: 10.1194/jlr.d600049-jlr200
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Highly sensitive assay of HMG-CoA reductase activity by LC-ESI-MS/MS

Abstract: We have developed a new sensitive and specific nonradioisotope assay method to measure the activity of HMG-CoA reductase, the rate-controlling enzyme in the cholesterol biosynthetic pathway. This method was based upon a stable isotope dilution technique by liquid chromatographytandem mass spectrometry using electrospray ionization in positive mode. Mevalonic acid, the product of HMG-CoA reductase, was converted to mevalonolactone (MVL) in an incubation mixture, extracted by a salting-out procedure, derivatized… Show more

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Cited by 17 publications
(11 citation statements)
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“…15 The method for the measurement of HMGCR activity was carried out as described previously. 16 Determination of Intracellular cAMP and TC. See Supporting Materials and Methods.…”
Section: Methodsmentioning
confidence: 99%
“…15 The method for the measurement of HMGCR activity was carried out as described previously. 16 Determination of Intracellular cAMP and TC. See Supporting Materials and Methods.…”
Section: Methodsmentioning
confidence: 99%
“…2, the sensitivity of N-ESI was not suffi cient to quantify MA concentrations in a small volume of normal human serum. Recently, we derivatized another organic acid, mevalonate, into mevalonyl-(2-pyrrolidin-1-yl-ethyl)-amide and measured it using LC-P-ESI-MS/MS ( 12 ). In this method, mevalonate was lactonized into mevalonolactone and then a tertiary amine moiety was introduced by a characteristic amidation reaction with a primary alkylamine.…”
Section: Comparison Of the Sensitivities Between P-esi And N-esi Methodsmentioning
confidence: 99%
“…Hepatocellular microsomes for assay of HMGCR activity were prepared as described by Honda et al ( 23 ). The HepG2 cells stimulated by different concentrations of bTSH (0, 1, and 4 M) were homogenized with a loose-fi tting Tefl on pestle in 4 vol of 3 mM TrisHCl buffer (pH 7.4) containing 0.25 M sucrose, 0.1 mM EDTA, and Subclinical hypothyroidism is a type of thyroid function abnormality; its characteristics are elevated serum thyroidstimulating hormone (TSH) and normal serum thyroid hormone levels, as well as an increased serum cholesterol level ( 10 ).…”
Section: Hepatocellular Microsome Hmgcr Activity Assaymentioning
confidence: 99%