Dataset Information

Targeted metabolic profiling of urinary steroids with a focus on analytical accuracy and sample stability.

ABSTRACT:

Introduction

Preoperative diagnostic workup of adrenal tumors is based on imaging and hormone analyses, but charged with uncertainties. Steroid profiling by liquid chromatography tandem mass spectrometry (LC-MS/MS) in 24-h urine has shown potential to discriminate benign and malignant adrenal tumors. Our aim was to develop and validate a specific and accurate LC-MS/MS method for the quantification of deconjugated urinary marker steroids, to evaluate their pre-analytical stability and to apply the method to clinical samples of patients with adrenal tumors.

Methods

A method for the quantification of 11 deconjugated steroids (5-pregnenetriol, dehydroepiandrosterone, cortisone, cortisol, α-cortolone, tetrahydro-11-deoxycortisol, etiocholanolone, pregnenolone, pregnanetriol, pregnanediol, and 5-pregnenediol) in human urine was developed and validated based on international guidelines. Steroids were enzymatically deconjugated and extracted by solid phase extraction before LC-MS/MS quantification in positive electrospray ionization mode.

Results

Excellent linearity with R² > 0.99 and intra- and inter-day precisions of < 10.1 % were found. Relative matrix effects were between 96.4 % and 101.6 % and relative recovery was between 98.2 % and 115.0 %. Sufficient pre-freeze stability for all steroids in urine was found at 20-25 °C for seven days and at 4-6 °C for up to 28 days. Samples were stable during long-term storage at -20 °C and -80 °C for 6 months.

Conclusions

A sensitive and robust LC-MS/MS method for the quantification of 11 urinary steroids was developed and validated according to international guidelines. Pre-analytical matrix stability was evaluated and the suitability of the method for the analysis of clinical samples and prospective validation studies was shown.

SUBMITTER: Vogg N

PROVIDER: S-EPMC9334310 | biostudies-literature | 2022 Aug

REPOSITORIES: biostudies-literature

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Publications

Targeted metabolic profiling of urinary steroids with a focus on analytical accuracy and sample stability.

Vogg Nora N Müller Tobias T Floren Andreas A Dandekar Thomas T Scherf-Clavel Oliver O Fassnacht Martin M Kroiss Matthias M Kurlbaum Max M

Journal of mass spectrometry and advances in the clinical lab 20220725

<h4>Introduction</h4>Preoperative diagnostic workup of adrenal tumors is based on imaging and hormone analyses, but charged with uncertainties. Steroid profiling by liquid chromatography tandem mass spectrometry (LC-MS/MS) in 24-h urine has shown potential to discriminate benign and malignant adrenal tumors. Our aim was to develop and validate a specific and accurate LC-MS/MS method for the quantification of deconjugated urinary marker steroids, to evaluate their pre-analytical stability and to ...[more]

PMID: 35910411

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Project description:BackgroundTo evaluate the metabolic changes in urinary steroids in pre- and post-menopausal women and men with papillary thyroid carcinoma (PTC).MethodsQuantitative steroid profiling combined with gas chromatography-mass spectrometry was used to measure the urinary concentrations of 84 steroids in both pre- (n = 21, age: 36.95 ± 7.19 yr) and post-menopausal female (n = 19, age: 52.79 ± 7.66 yr), and male (n = 16, age: 41.88 ± 8.48 yr) patients with PTC. After comparing the quantitative data of the patients with their corresponding controls (pre-menopause women: n = 24, age: 33.21 ± 10.48 yr, post-menopause women: n = 16, age: 49.67 ± 8.94 yr, male: n = 20, age: 42.75 ± 4.22 yr), the levels of steroids in the patients were normalized to the mean concentration of the controls to exclude gender and menopausal variations.ResultsMany urinary steroids were up-regulated in all PTC patients compared to the controls. Among them, the levels of three active androgens, androstenedione, androstenediol and 16α-hydroxy DHEA, were significantly higher in the pre-menopausal women and men with PTC. The corticoid levels were increased slightly in the PTC men, while progestins were not altered in the post-menopausal PTC women. Estrogens were up-regulated in all PTC patients but 2-hydroxyestrone and 2-hydroxy-17β-estradiol were remarkably changed in both pre-menopausal women and men with PTC. For both menopausal and gender differences, the 2-hydroxylation, 4-hydroxylation, 2-methoxylation, and 4-methoxylation of estrogens and 16α-hydroxylation of DHEA were differentiated between pre- and post-menopausal PTC women (P < 0.001). In particular, the metabolic ratio of 2-hydroxyestrone to 2-hydroxy-17β-estradiol, which could reveal the enzyme activity of 17β-hydroxysteroid dehydrogenase, showed gender differences in PTC patients (P < 1 × 10-7).ConclusionsThese results are expected be helpful for better understanding the pathogenic differences in PTC according to gender and menopausal conditions.

Project description:Synthetic cathinones constitute the second largest groups of new psychoactive substances (NPS), which are especially popular among adolescents and young adults. Due to their potential toxicity, the recreational use of these NPS constitute a serious worldwide public health problem. However, their fast appearance in the market renders the continuous updating of NPS information highly challenging for forensic authorities. The unavailability of pharmacokinetic data for emerging NPS is critical for forensic and clinical verifications. With the ultimate goal of having a proactive approach towards the NPS issue, high resolution mass spectrometry was used in the current work to assess preliminary pharmacokinetic data for 8 selected cathinones: 4 reported substances (4-CIC, 3-CMC, 4-CMC and 4-MEAP) and 4 previously unreported ones (3-CIC, 4-MDMB, 4-MNEB and 4-MDMP) for which the emergence on the NSP market is expected to be eminent, were also included in this study. Based on the calculation of pharmacokinetic parameters, half-life and intrinsic clearance, 4-CMC and 4-MDMB are low and high clearance compounds, respectively, and all the remaining cathinones included in this study are intermediate clearance compounds. This fact anticipates the key role of metabolites as suitable biomarkers to extend detection windows beyond those provided by the parent cathinones. Reduction of the keto group and hydroxylation on the alkyl chains were the common metabolic pathways identified for all cathinones. However, the relative importance of these metabolic transformations is dependent on the cathinone substituents. The glucuronic acid conjugation to metabolites stemming for keto group reduction constituted the sole Phase II transformation identified. To our knowledge, this study constitutes the first metabolite profiling of the already reported synthetic cathinones 4-CIC, 3-CMC and 4-CMC. Noteworthy is the fact that 3-CMC accounts for almost a quarter of the quantity of powders seized during 2020. The analytical methods developed, and the metabolites characterized, are now available to be included in routine screening methods to attest the consumption of the 8 cathinones studied.

Dataset Information

Targeted metabolic profiling of urinary steroids with a focus on analytical accuracy and sample stability.

Introduction

Methods

Results

Conclusions

Publications

Targeted metabolic profiling of urinary steroids with a focus on analytical accuracy and sample stability.

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