Determination of 24S- and 27-hydroxycholesterol in plasma by high-performance liquid chromatography-mass spectrometry

Burkard, Ines; Rentsch, Katharina; Eckardstein, Arnold von

doi:10.1194/jlr.d300036-jlr200

Cited by 80 publications

(71 citation statements)

References 23 publications

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“…Recently, liquid chromatography-atmospheric pressure chemical ionization-mass spectrometry (LC-APCI-MS) was introduced as a sensitive, specific, and rapid method for the quantification of oxysterols (20,21). In addition, LCtandem mass spectrometry (LC-MS/MS) using electrospray ionization (ESI) has also been applied to the analysis of oxysterols (22).…”

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confidence: 99%

Highly sensitive quantification of key regulatory oxysterols in biological samples by LC-ESI-MS/MS

Honda

Yamashita

Hara

et al. 2009

Journal of Lipid Research

177

154

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We describe a highly sensitive and specific method for the quantification of key regulatory oxysterols in biological samples. This method is based upon a stable isotope dilution technique by liquid chromatography-tandem mass spectrometry (LC-MS/MS). After alkaline hydrolysis of human serum (5 ml) or rat liver microsomes (1 mg protein), oxysterols were extracted, derivatized into picolinyl esters, and analyzed by LC-MS/MS using the electrospray ionization mode. The detection limits of the picolinyl esters of 4b-hydroxycholesterol, 7a-hydroxycholesterol, 22R-hydroxycholesterol, 24S-hydroxycholesterol, 25-hydroxycholesterol, 27-hydroxycholesterol, and 24S,25-epoxycholesterol were 2-10 fg (5-25 amol) on-column (signal-to-noise ratio 5 3). Reproducibilities and recoveries of these oxysterols were validated according to one-way layout and polynomial equation, respectively. The variances between sample preparations and between measurements by this method were calculated to be 1.8% to 12.7% and 2.9% to 11.9%, respectively. The recovery experiments were performed using rat liver microsomes spiked with 0.05 ng to 12 ng of oxysterols, and recoveries of the oxysterols ranged from 86.7% to 107.3%, with a mean recovery of 100.6%. This method provides reproducible and reliable results for the quantification of oxysterols in small amounts of biological samples. Supplementary key words liquid chromatography-tandem mass spec-Biological samples contain a large number of oxysterols (1), and most of them are formed from cholesterol by enzymatic oxidation (2 -6) ( Fig. 1) or autoxidation (7). By contrast, the oxysterol 24S,25-epoxycholesterol is not derived from cholesterol but is produced de novo from acetyl-CoA via a shunt in the mevalonate pathway (8).These oxysterols are important molecules for preserving lipid homeostasis in the body. 7a-Hydroxycholesterol is a product of CYP7A1, which is the rate-limiting enzyme in the classic bile acid biosynthetic pathway. 27-Hydroxycholesterol, 24S-hydroxycholesterol, 4b-hydroxycholesterol, 22R-hydroxycholesterol, and 24S,25-epoxycholesterol are effective endogenous ligands of the nuclear receptors liver X receptor a (LXRa) and LXRb (9-11). In addition, 27-hydroxycholesterol (12), 25-hydroxycholesterol (13), and 24S,25-epoxycholesterol (14) are known to downregulate the cholesterol biosynthetic pathway, presumably by blocking the processing of the sterol-regulatory element binding protein.GC-MS has historically been used for the analyses of oxysterols in serum and tissues (1, 15) because the sensitivity and specificity of conventional GC with flame ionization detector is not sufficient to quantify oxysterols in biological samples. However, GC-MS is still not an ideal method, especially for the analysis of 24S,25-epoxycholesterol, because this epoxycholesterol does not survive the temperature required for GC analysis (16). Another approach to quantifying oxysterols in biological samples was HPLC with ultraviolet (UV) detection after derivatization to the D 4 -3-ketones (16-19). Thi...

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mentioning

confidence: 99%

Highly sensitive quantification of key regulatory oxysterols in biological samples by LC-ESI-MS/MS

Honda

Yamashita

Hara

et al. 2009

Journal of Lipid Research

177

154

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“…The concentration of 27-OHC in the plasma of the CTX patient was reduced (3.8 vs 90-140 ng/ml normal range in controls) (Burkard et al 2004), reflecting the decreased activity of CYP27A1 (Table 1). To establish whether this decreased concentration of 27-OHC was mirrored by changes in glucocorticoid ratios reflecting the activity of HSD11B1, the concentrations of glucocorticoid metabolites in the plasma and urine were determined by GC-MS.…”

Section: Analysis Of 27-ohc and Glucocorticoid Metabolites In The Ctxmentioning

confidence: 99%

“…F and E and their corresponding tetrahydrometabolites THF, 5a-THF and THE, a-cortol, b-cortol, a-cortolone and b-cortolone in the urine were quantified by GC-MS. Control values were as those reported in previous studies , N'Gankam et al 2002, Burkard et al 2004 …”

Section: Rna Extraction and Real-time Pcrmentioning

confidence: 99%

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Evidence for a role of sterol 27-hydroxylase in glucocorticoid metabolism in vivo

Vögeli¹,

Jung²,

Dick³

et al. 2013

Journal of Endocrinology

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The intracellular availability of glucocorticoids is regulated by the enzymes 11b-hydroxysteroid dehydrogenase 1 (HSD11B1) and 11b-hydroxysteroid dehydrogenase 2 (HSD11B2). The activity of HSD11B1 is measured in the urine based on the (tetrahydrocortisolC5a-tetrahydrocortisol)/tetrahydrocortisone ((THFC5a-THF)/THE) ratio in humans and the (tetrahydrocorticosteroneC5a-tetrahydrocorticosterone)/tetrahydrodehydrocorticosterone ((THBC5a-THB)/THA) ratio in mice. The cortisol/cortisone (F/E) ratio in humans and the corticosterone/11-dehydrocorticosterone (B/A) ratio in mice are markers of the activity of HSD11B2. In vitro agonist treatment of liver X receptor (LXR) downregulates the activity of HSD11B1. Sterol 27-hydroxylase (CYP27A1) catalyses the first step in the alternative pathway of bile acid synthesis by hydroxylating cholesterol to 27-hydroxycholesterol (27-OHC). Since 27-OHC is a natural ligand for LXR, we hypothesised that CYP27A1 deficiency may up-regulate the activity of HSD11B1. In a patient with cerebrotendinous xanthomatosis carrying a loss-of-function mutation in CYP27A1, the plasma concentrations of 27-OHC were dramatically reduced (3.8 vs 90-140 ng/ml in healthy controls) and the urinary ratios of (THFC5a-THF)/THE and F/E were increased, demonstrating enhanced HSD11B1 and diminished HSD11B2 activities. Similarly, in Cyp27a1 knockout (KO) mice, the plasma concentrations of 27-OHC were undetectable (!1 vs 25-120 ng/ml in Cyp27a1 WT mice). The urinary ratio of (THBC5a-THB)/THA was fourfold and that of B/A was twofold higher in KO mice than in their WT littermates. The (THBC5a-THB)/THA ratio was also significantly increased in the plasma, liver and kidney of KO mice. In the liver of these mice, the increase in the concentrations of active glucocorticoids was due to increased liver weight as a consequence of Cyp27a1 deficiency. In vitro, 27-OHC acts as an inhibitor of the activity of HSD11B1. Our studies suggest that the expression of CYP27A1 modulates the concentrations of active glucocorticoids in both humans and mice and in vitro. Journal of EndocrinologyResearch I VÖ GELI and others CYP27A1 and glucocorticoid metabolism in vivo

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“…have been described in the literature but exhibit insufficient separation and quite high limits of detection (18,21,22 ). Derivatization, for example as picolinyl esters (23 ) or Girard P hydrazones (19 ), in LC-based methods enhanced the performance and decreased the detection limit but appeared more laborious than GCbased methods.…”

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confidence: 99%

Gas Chromatography–Tandem Mass Spectrometry Method for the Simultaneous Determination of Oxysterols, Plant Sterols, and Cholesterol Precursors

2012

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Determination of 24S- and 27-hydroxycholesterol in plasma by high-performance liquid chromatography-mass spectrometry

Cited by 80 publications

References 23 publications

Highly sensitive quantification of key regulatory oxysterols in biological samples by LC-ESI-MS/MS

Highly sensitive quantification of key regulatory oxysterols in biological samples by LC-ESI-MS/MS

Evidence for a role of sterol 27-hydroxylase in glucocorticoid metabolism in vivo

Gas Chromatography–Tandem Mass Spectrometry Method for the Simultaneous Determination of Oxysterols, Plant Sterols, and Cholesterol Precursors

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