Gas chromatographic–mass spectrometric analysis of urinary glycyrrhetinic acid: an aid in diagnosing liquorice abuse

Abstract

Abstract. Kerstens MN, Guillaume CPF, Wolthers BG, Dullaart RPF (University Hospital Groningen, Groningen, The Netherlands). Gas chromatographic±mass spectrometric analysis of urinary glycyrrhetinic acid: an aid in diagnosing liquorice abuse. J Intern Med 1999; 246: 539±547.Objectives. Liquorice abuse can lead to severe clinical complications, caused by its active compound 18b-glycyrrhetinic acid (18bGA). 18bGA inhibits dehydrogenase activity of 11b-hydroxysteroid dehydrogenase (11bHSD). This enzyme catalyses the interconversion between cortisol and cortisone and normally protects the mineralocorticoid receptor from being activated by cortisol. Diagnosing liquorice abuse can be notoriously difficult. The aim of our study was to develop an accurate and clinically applicable 18bGA urinary assay. Design. We developed a urinary 18bGA assay based on gas chromatography and mass spectrometry (GCMS) with sufficient sensitivity to detect 18bGA at low concentrations. The assay was validated in four volunteers consuming different amounts of liquorice. We applied its use in two patients with hypokalaemic hypertension and suppressed plasma renin activity and serum aldosterone, who were suspected of liquorice abuse. Results. The detection limit for 18bGA of the GC assay was 10 mg L 21 , which was lowered to 3 mg L 21 by subsequent application of MS. In all volunteers, urinary 18bGA was detected during liquorice intake. Urinary 18bGA remained detectable until 5 days after stopping continued liquorice intake and until at least 51 h after ingestion of a single large amount. Urinary 18bGA was demonstrated in both patients, establishing a diagnosis of liquorice abuse. One patient showed changes in urinary cortisol metabolites, consistent with 11bHSD inhibition. Changes in cortisol metabolites were less pronounced in the other patient. Conclusion. Liquorice abuse can result in hypokalaemic hypertension with prolonged suppression of plasma renin activity and aldosterone concentration. This is caused by 18bGA-mediated inhibition of 11bHSD, resulting in activation of the renal mineralocorticoid receptor by cortisol. Urinary 18bGA measurement by GCMS is a useful aid in establishing liquorice abuse.

Glycyrrhetinic acid attenuates vascular smooth muscle vasodilatory function in healthy humans

“…Increased MR activation in our subjects is supported by the reduction in plasma potassium and suppression of plasma aldosterone levels at the end of the GA treatment period. These results are consistent with previous reports of liquorice administration [20,21]. In our cohort, there was no change in BP attributable to GA treatment.…”

Biomarkers of food intake for cocoa and liquorice (products): a systematic review

“…of men) Age range (years) BMI range b (kg/m 2 ) Analytical method Approach Sample type and time Candidate biomarkers of food intake HMDB ID* Primary ref. Liquorice (products) Solid Katzen Kinder by Katjes (0.05% GL) 200 g 135 Acute single-dose study 4 (1) 26–29 18.6–28.6 LC-MS/MS (LC-ESI-MS) Targeted Blood (0, 0:05, 1, 2, 3, 5 and 7 h) Urine (regularly) Blood, 18-GA (highest after 6 h; 150–434 ng/ml) Urine − HMDB0011628 [ 53 ] Solid liquorice (0.23% GL) 50, 100 and 200 g daily for 5 days 500 g within 4 h 68, 135 and 271 676 Parallel intervention study (5 days) Acute single-dose study 3 (0) 1 (1) 19–23 22 U U GC-MS Targeted Urine (all up to 5 days) Urine, 18-GA [ 52 ] Isolated compounds Glycyrrhizin (GL) 600 mg in 2 dl water (excretion study) 353 Acute single-dose study 6 (5) 24–40 U HPLC-UV Targeted Urine (all up to 4 days) 18-GA 3-MGA (Cmax 0.49–2.69 μg/ml) HMDB0037827 [99] Glycyrrhetinic acid (GA) 500 mg 509 c Acute single-dose study 10 (10) 24–38 U LC analyser Targeted Serum (0, 2, 4, 7, 10 and 24 h) Urine (0, 2, 4, 7, 10 and 24 h) Serum, 18-GA (max after 2–4 h 13.4 μmol/l) Urine − [ 54 ] Liquorice extract (LE) vs. glycyrrhizin 21 g LE vs. 1600 mg GL 941 …”

“…Krahenbuhl et al [ 51 ] observed a clear dose-response relationship for 18-GA in plasma; however, the concentration GA used in this study was comparable to an intake of liquorice ranging from 1.7 to 5.2 kg; therefore, no relevant intake levels of the targeted food were examined in this study. In the study of Kerstens et al [ 52 ], 50–200 g of liquorice were consumed, showing a dose-response relationship of 18-GA in urine (question 2). 18-GA was found to be traceable in urine after the intake of 50 g of solid liquorice, and around 0.04% of the total intake could be traced back after 51 h in total [ 52 ].…”

Determination of reference intervals for urinary steroid profiling using a newly validated GC-MS/MS method

“…For example, congenital adrenal hyperplasia (CAH), in particular the milder variants, might be diagnosed for the first time in adulthood [6,7] by noticing moderately elevated P3, PTL and PDL excretions and/or elevated diagnostic ratios for these enzyme deficiencies (Supplementary Table 4). In addition, USP can be helpful to evaluate the efficacy of glucocorticoid treatment in adult patients with CAH or for the diagnosis of licorice induced hypertension, which is reflected by an impaired hydroxysteroiddehydrogenase activity [8]. Patients with ACC often demonstrate several disorders in steroid biosynthesis.…”

“…USP has a broad range of applications. For example, it can be used for the diagnosis and follow-up of disorders resulting from steroid biosynthetic enzyme deficiencies, licorice-induced hypertension, hirsutism and other related diseases [6][7][8]. Furthermore, USP might be helpful in monitoring patients with an adrenocortical carcinoma (ACC) and could be useful in discriminating between malignant and benign adrenal tumors [9][10][11][12].…”