Identification of an Alternative Glycyrrhizin Metabolite Causing Liquorice-Induced Pseudohyperaldosteronism and the Development of ELISA System to Detect the Predictive Biomarker.

Kan'Ichiro Ishiuchi,Tsubasa Kondo,Yuuna Tashita,Takao Namiki,Toshiaki Makino,Fangyi Lian,Miaki Mitamura,Tetsuhiro Yoshino,Masaru Mimura,Kenji Watanabe,Kiyoshi Minamizawa,Kakuyou Ogawa,Osamu Morinaga,Asuka Hirasawa,Yasuhito Maki,Keiko Ogawa-Ochiai

doi:10.3389/fphar.2021.688508

Abstract

Liquorice is usually used as crude drug in traditional Japanese Kampo medicine and traditional Chinese medicine. Liquorice-containing glycyrrhizin (GL) can cause pseudohyperaldosteronism as a side effect. Previously, we identified 18β-glycyrrhetyl-3-O-sulfate (3) as a GL metabolite in Eisai hyperbilirubinuria rats (EHBRs) with the dysfunction of multidrug resistance-related protein (Mrp2). We speculated that 3 was associated with the onset of liquorice-induced pseudohyperaldosteronism, because it was mainly detected in serum of patients with suspected to have this condition. However, it is predicted that other metabolites might exist in the urine of EHBRs orally treated with glycyrrhetinic acid (GA). We explored other metabolites in the urine of EHBRs, and investigated the pharmacokinetic profiles of the new metabolite in EHBRs and normal Sprague-Dawley rats. We further analyzed the serum concentrations of the new metabolite in the patients of pseudohyperaldosteronism. Finally, we developed the analyzing method of these metabolites as a preventive biomarker for the onset of pseudohyperaldosteronism using an enzyme-linked immunosorbent assay (ELISA). We isolated a new GL metabolite, 18β-glycyrrhetyl-3-O-sulfate-30-O-glucuronide (4). Compound 4 significantly inhibited rat type-2 11β-hydroxysteroid dehydrogenase (11β-HSD2) and was a substrate of both organic anion transporter (OAT) 1 and OAT3. Compound 4 was also detected in the serum of patients with suspected pseudohyperaldosteronism at an approximately 10-fold lower concentrations than 3, and these concentrations were positively correlated. Compound 4 showed a lower serum concentration and weaker inhibitory titer on 11β-HSD2 than 3. We developed an enzyme-linked immunosorbent assay system using an anti-18β-glycyrrhetyl-3-O-glucuronide (3MGA) monoclonal antibody to measure the serum concentration of 3 to facilitate the measurement of biomarkers to predict the onset of pseudohyperaldosteronism. Although we found 4 as the secondary candidate causative agent, 3 could be the main potent preventive biomarker of liquorice-induced pseudohyperaldosteronism. Compound 3 was detected in serum at a higher concentration than GA and 4, implying that 3 may be a pharmacologically active ingredient mediating not only the development of pseudohyperaldosteronism but anti-inflammatory effects in humans administered GL or other liquorice-containing preparations.

Highlights

Liquorice, the root or stolon of Glycyrrhiza uralensis or G. glabra, is a natural material commonly used as a sweetener for foods as well as a crude drug used in traditional Japanese Kampo medicine and traditional Chinese medicine (TCM)
Cortisol, which has a similar affinity to the mineralocorticoid receptor as aldosterone, is converted by 11β-HSD2 to cortisone, which has a low affinity for the receptor
We evaluated the association of the concentration of each GL metabolite in serum samples of patients with suspected pseudohyperaldosteronism with the laboratory data and various symptoms

Summary

Introduction

The root or stolon of Glycyrrhiza uralensis or G. glabra, is a natural material commonly used as a sweetener for foods as well as a crude drug used in traditional Japanese Kampo medicine and traditional Chinese medicine (TCM). Its component glycyrrhizin (5) (GL; Figure 1A) has various pharmacological actions such as anti-allergy, antiinflammation, anti-ulcer, anti-virus, anti-androgenic, and hepatorptective effects (Isbrucker and Burdock, 2006). Pseudohyperaldosteronism, a well-known side effect of liquorice, is characterized by hypertension, oedema, and hypokalaemia (Conn et al, 1968; Stewart et al, 1987). The onset of pseudohyperaldosteronism is associated with the inhibition of type-2 11β-hydroxysteroid dehydrogenase (11βHSD2) in renal distal tubular epithelial cells by some GL metabolites (Ploeger et al, 2001). Cortisol, which has a similar affinity to the mineralocorticoid receptor as aldosterone, is converted by 11β-HSD2 to cortisone, which has a low affinity for the receptor. When 11β-HSD2 is inhibited by GL metabolites, the local cortisol concentration increases, which stimulates the mineralocorticoid receptor

Methods

Results

Discussion

Conclusion