Preparation, characterization, and liver targeting evaluation of a novel sustained-release brucine self-assembled micelle mediated by glycyrrhetinic acid.

Background: Cancer is a serious threat to human life, health and social development. In recent years, nanomicelles, as an emerging drug carrier material, have gradually entered people's field of vision because of their advantages of improving bioavailability, maintaining drug levels, reducing systemic side effects and increasing drug accumulation at target sites. Methods: In this study, B-GPSG nano-micelles were prepared by film dispersion hydration method using brucine as model drug and glycyrrhetinic acid-polyethylene glycol-3-methylene glycol-dithiodipropionic acid-glycerol monostearate polymer as nano-carrier. The preparation process, characterization, drug release in vitro, pharmacokinetics and liver targeting were investigated. Results: The results showed that the range of particle size, polydispersion index and Zeta potential were 102.7 ± 1.09 nm, 0.201 ± 0.02 and -24.5 ± 0.19 mV respectively. The entrapment efficiency and drug loading were 83.79 ± 2.13% and 12.56 ± 0.09%, respectively. The drug release experiments in vitro and pharmacokinetic experiments showed that it had obvious sustained release effect. For pharmacokinetics study, it shows that both the B-GPSG solution group and the B-PSG solution group changed the metabolic kinetic parameters of brucine, but the B-GPSG solution group had a better effect. Compared with the B-PSG solution group, the drug was more prolonged in rats. The half-life in the body and the retention time in the body of B-GPSG are more helpful to improve the bioavailability of the drug and play a long-term effect. The tail vein injection results of mice indicate that B-GPSG can target and accumulate brucine in the liver without affecting other key organs. Cell uptake experiments and tissue distribution experiments in vivo show that glycyrrhetinic acid modified nano-micelles can increase the accumulation of brucine in hepatocytes, has a good liver targeting effect, and can be used as a new preparation for the treatment of liver cancer. Conclusion: The B-SPSG prepared in this experiment can provide a new treatment method and research idea for the treatment of liver cancer.

Exploring the Pivotal Immunomodulatory and Anti-Inflammatory Potentials of Glycyrrhizic and Glycyrrhetinic Acids.

Licorice extract is a Chinese herbal medication most often used as a demulcent or elixir. The extract usually consists of many components but the key ingredients are glycyrrhizic (GL) and glycyrrhetinic acid (GA). GL and GA function as potent antioxidants, anti-inflammatory, antiviral, antitumor agents, and immuneregulators. GL and GA have potent activities against hepatitis A, B, and C viruses, human immunodeficiency virus type 1, vesicular stomatitis virus, herpes simplex virus, influenza A, severe acute respiratory syndrome-related coronavirus, respiratory syncytial virus, vaccinia virus, and arboviruses. Also, GA was observed to be of therapeutic valve in human enterovirus 71, which was recognized as the utmost regular virus responsible for hand, foot, and mouth disease. The anti-inflammatory mechanism of GL and GA is realized via cytokines like interferon-γ, tumor necrotizing factor-α, interleukin- (IL-) 1ß, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12, and IL-17. They also modulate anti-inflammatory mechanisms like intercellular cell adhesion molecule 1 and P-selectin, enzymes like inducible nitric oxide synthase (iNOS), and transcription factors such as nuclear factor-kappa B, signal transducer and activator of transcription- (STAT-) 3, and STAT-6. Furthermore, DCs treated with GL were capable of influencing T-cell differentiation toward Th1 subset. Moreover, GA is capable of blocking prostaglandin-E2 synthesis via blockade of cyclooxygenase- (COX-) 2 resulting in concurrent augmentation nitric oxide production through the enhancement of iNOS2 mRNA secretion in Leishmania-infected macrophages. GA is capable of inhibiting toll-like receptors as well as high-mobility group box 1.

Synthesis and antitumor effects of novel 18ß-glycyrrhetinic acid derivatives featuring an exocyclic α,ß-unsaturated carbonyl moiety in ring A.

A series of novel 18ß-glycyrrhetinic acid (GA) derivatives featuring an exocyclic α,ß-unsaturated carbonyl moiety in ring A were synthesized and evaluated for their antitumor activities. Compounds 5c and 5l showed stronger cytotoxicity than other compounds and reported GA analogue CDODA-Me (methyl 2-cyano-3,11-dioxo-18ß-olean-1,12-dien-30-oate). 5c and 5l induced apoptosis in cancer cells accompanying with c-Flip reduction and Noxa induction, associated with decreased HDAC3 expression and increased acetylation of H3. 5l displayed better stability properties than 5c and CDODA-Me in microsomes and plasma, 5l also showed favorable pharmacokinetic profiles and inhibited tumor growth in mice. Compound 5l represents a new type of GA derivatives with improved antitumor activity.

A 18ß-glycyrrhetinic acid conjugate with Vorinostat degrades HDAC3 and HDAC6 with improved antitumor effects.

Semisynthetic 18ß-glycyrrhetinic acid (GA) analogues bearing 1-en-2-cyano-3-oxo substitution on ring A have enhanced antitumor effects with reduced levels of HDAC3 and HDAC6 proteins. Aiming to inhibit both HDAC protein and activity, we developed a hybrid molecule by tethering active GA analogue methyl 2-cyano-3,11-dioxo-18ß-olean-1,12-dien-30-oate (CDODA-Me) and Vorinostat (SAHA). We tested the proper hybrid approaches of GA with hydroxamic acid and turned out that GA conjugated with SAHA by a piperazine linker was the best. The conjugate (15) of CDODA-Me and SAHA linked through a piperazine group was a potent cytotoxic agent against cancer cells with apoptosis induction. Compound 15 was more effective than the simple combination of CDODA-Me and SAHA to induce apoptosis. Mechanistic studies revealed that 15 was less effective than SAHA to inhibit HDAC activity, but was more effective than CDODA-Me to decrease the levels of HDAC3 and HDAC6 proteins with upregulated levels of acetylated H3 and acetylated α-tubulin. Compound 15 represents a new HDAC3 and HDAC6 inhibitor by reducing protein levels.

Tumor-Associated Fibroblast-Targeted Regulation and Deep Tumor Delivery of Chemotherapeutic Drugs with a Multifunctional Size-Switchable Nanoparticle.

Tumor-associated fibroblasts (TAFs), which form a predominant stromal cellular component of the tumor microenvironment, hinder the delivery of nanomedicine to deep tumor cells and lead to poor prognosis of tumors. However, depletion of TAFs by therapeutic agents results in the secretion of damage response program (DRP) molecules to weaken the efficacy of tumor treatment. This paper reports a multifunctional size-switchable nanoparticle (denoted DGL (dendrigraft poly-l-lysine) (DGL)/GEM@PP/GA) for TAF-targeted regulation and deep tumor penetration. After accumulation at the tumor site, in response to overexpressed matrix metalloproteinase-2 (MMP-2) in the tumor microenvironment, gemcitabine (GEM)-conjugated small nanoparticles (DGL/GEM) are released from DGL/GEM@PP/GA, leaving 18ß-glycyrrhetinic acid (GA)-loaded large nanoparticles (PP/GA). The released DGL/GEM can penetrate to the deep region of the tumor as well as intracellularly release GEM to kill tumor cells. However, residual GA-loaded nanoparticles with lower tumor penetration ability could accumulate around tumor vessels and be preferentially absorbed by TAFs to regulate the secretion of Wnt 16, which is an important DRP molecule. By taking actions on both tumor cells and TAFs, DGL/GEM@PP/GA displayed significant and long-term antitumor effect in stroma-rich pancreatic cancer and breast cancer models.

Comparative pharmacokinetics and metabolites study of seven major bioactive components of Shaoyao-Gancao decoction in normal and polycystic ovary syndrome rats by ultra high pressure liquid chromatography with tandem mass spectrometry.

A simple and sensitive liquid chromatography with tandem mass spectrometry method was developed for simultaneous quantification of paeoniflorin, albiflorin, oxypaeoniflorin, liquiritin, liquiritigenin, glycyrrhetinic acid, and glycyrrhizin in rat plasma after oral administration of Shaoyao-Gancao decoction, which is traditionally used in the treatment of polycystic ovary syndrome. The plasma samples were pretreated with methanol as precipitant. The method exhibited good linearity (correlation coefficient (R2 ) > 0.99) with lower quantification limits of 0.595-4.69 ng/mL for all analytes. Intra- and interbatch precision, accuracy, recovery, and stability of the method were all within accepted criteria. The results showed that the pharmacokinetic behaviors of the seven compounds were altered in the pathological status of polycystic ovary syndrome. Furthermore, a total of 36 metabolites were structurally identified based on their accurate masses and fragment ions. The major metabolic pathway involves phase I metabolic reactions (such as hydroxylation), phase II metabolic reactions (such as sulfation and glucuronidation conjugation) as well as the combined multiple-step metabolism. This study is the first report on the pharmacokinetic and metabolic information of Shaoyao-Gancao decoction in both normal and model rats, which would provide scientific evidences for the bioactive chemical basis of herbal medicines and also promote the clinical application of Shaoyao-Gancao decoction for treating polycystic ovary syndrome.

Inhibiting hepatic gluconeogenesis by chitosan lactate nanoparticles containing CRTC2 siRNA targeted by poly(ethylene glycol)-glycyrrhetinic acid.

Diabetes mellitus is a chronic metabolic disorder characterized by insulin deficiency and impaired glucose metabolism. Overexpression of cAMP response element binding protein (CREB)-regulated transcriptional coactivator 2 (CRTC2) plays an important role in high gluconeogenesis in patients with diabetes type II. Using RNA interference technology for silencing CRTC2 gene expression could be helpful in controlling the level of blood glucose and gluconeogenesis. In this study, we designed a siRNA delivery platform comprising blended chitosan lactate (CT) and polyethylene glycol (PEG) conjugated with glycyrrhetinic acid (GA) for controlling gluconeogenesis. The nanoparticles showed spherical and smooth surface with ~ 189-nm size and + 5.1 zeta potential. Targeted nanoparticles were efficiently stable in serum and different levels of heparin media over 48 h. The gene knockdown efficiency of nanoparticles was comparable to Lipofectamine®, while they had no significant in vitro and in vivo toxicity. The in vivo therapeutic efficacy of targeted nanoparticles was also confirmed by reduced amount of fasting blood sugar in diabetic rat models. Furthermore, the nanoparticles were mostly accumulated in the liver after 2 h indicating the significant targeting ability of the prepared nanoparticles. Therefore, CT/PEG-GA nanoparticles can be considered as a potential candidate for targeted delivery of siRNA into hepatocytes in order to regulate gluconeogenesis in diabetes.

Glycyrrhetinic acid-conjugated polymeric prodrug micelles co-delivered with doxorubicin as combination therapy treatment for liver cancer.

Significant synergy of doxorubicin (DOX) and glycyrrhizic acid (GA) in inhibiting the proliferation of cancer cells was demonstrated in the human hepatocellular carcinoma cell line, HepG2. A novel polymeric prodrug micellar carrier based on polyethylene glycol-derivatized GA (PEG-Fmoc-GA), was developed for co-delivery of DOX as a combined anti-cancer treatment. The PEG-Fmoc-GA polymeric prodrug micelles achieved a more effective synergistic action on cell proliferation inhibition and apoptosis induction, when co-delivered with DOX, which can be attributed to the dual effect of the cleaved GA and loaded DOX. PEG-Fmoc-GA conjugated micelles significantly facilitated the intracellular uptake of DOX by HepG2 cells, when compared to a DOX solution alone. In addition, DOX encapsulated in PEG-Fmoc-GA micelles displayed longer blood circulation time, larger drug concentration area under the curve, decreased volume distribution and clearance than DOX solution. Biodistribution studies showed that DOX/PEG-Fmoc-GA micelles were preferentially accumulated at the tumor site. Importantly, DOX/PEG-Fmoc-GA micelles demonstrated a more pronounced therapeutic efficacy in vivo compared with DOX alone with respect to both tumor growth inhibition and overall survival in a HepG2 xenograft model. Thus, PEG-Fmoc-GA polymeric prodrug micelles represent a promising dual-function co-delivery system to achieve anti-cancer synergistic activity of DOX and GA.

Liver-Targeted Co-delivery of Entecavir and Glycyrrhetinic Acid Based on Albumin Nanoparticle To Enhance the Accumulation of Entecavir.

Hepatitis B, one of the most common contagious viral hepatitis with high infection rate, is challenging to treat. Although the treatment for hepatitis B has been improved over the years, many therapeutic drugs still have either severe adverse effects or insufficient effectiveness via systemic administration. In this study, we confirmed that glycyrrhetinic acid can enhance the accumulation of entecavir in HepaRG cell and liver. Then we constructed a novel albumin nanoparticle co-loading entecavir and glycyrrhetinic acid (ETV-GA-AN) to improve liver accumulation of entecavir and investigated its ability to deliver both drugs to liver. In vitro cellular uptake study and in vivo tissue distribution experiment showed that these negatively charged ETV-GA-AN (112 ± 2 nm in diameter) can increase the accumulation of entecavir in hepatic HepaRG cells and improve entecavir distribution in liver. We also revealed the mechanism that glycyrrhetinic acid enhances intracellular accumulation of entecavir by inhibiting the activity of specific efflux transporters. Our delivery system is the first liver-targeted albumin nanoparticle that utilizes the site-specific co-delivery strategy to delivery entecavir and glycyrrhetinic acid. As it combines high efficiency and low toxicity, it possess great potential for treating hepatitis B.

Acid-sensitive polymeric vector targeting to hepatocarcinoma cells via glycyrrhetinic acid receptor-mediated endocytosis.

Liver cancer is one of the top death causing cancers, traditional treatments have not settled for the requirement of patients. In this work, a smart acid-responsive micelle based on glycyrrhetinic acid modified chitosan-polyethyleneimine-4-Hydrazinobenzoic acid-doxorubicin (GA-CS-PEI-HBA-DOX) was synthesized for targeted delivery of DOX to liver cancer. A dual pH-sensitive and receptor-mediated strategy has been exploited to enhance the delivery efficiency. The micelle possesses positive charges under pH 6.8 and can be turned into negative charges above pH 7.0, which help to be accumulated in tumor tissues (pH 6.0-7.0). In the intracellular environment (pH 4.5-6.5) of tumor cells, the pH-sensitive hydrazone bonds between DOX and GA-CS-PEI-HBA would break and release as much as 90.3% of the encapsulated payloads in 48 h. In addition, GA was modified to improve the targeting abilities. The micelles exhibited high lethality to HepG2 cells while showed much lower cytotoxicity to HUVEC cells. With high drug-loading capacity and the targeted release ability, the GA-CS-PEI-HBA-DOX micelle might be employed as a promising candidate for targeted cancer treatment.

An intelligent re-shieldable targeting system for enhanced tumor accumulation.

Programmed ligand targeting strategy promotes the blood circulation stability of nanoparticles by shielding the ligand. However, the irreversible shielding causes the deshielded nanoparticles to be easily recognized and cleared by the reticuloendothelial system (RES), impeding their further retention in the tumor. Here, we for the first time prove the superiority of the intelligent re-shieldable targeting system that is based on the pH-responsive self-assembly/disassembly of gold nanoparticles. The system can enhance the stability of gold nanoparticles in the blood circulation (2.6-fold at 24h), reduce uptake by the RES (35% lower) and improve tumor accumulation (41% higher by analysis of gold content in tumor) effectively compared with the conventional irreversible system. Furthermore, preliminary study indicates that the system could be applied as computed tomography contrast agent in tumor imaging. The in vivo validity of the intelligent re-shieldable targeting system provides inspiration for the design of nanomaterials for cancer diagnosis and treatment.

A Glycyrrhetinic Acid-Modified Curcumin Supramolecular Hydrogel for liver tumor targeting therapy.

Curcumin (Cur), a phenolic anti-oxidant compound obtained from Curcuma longa plant, possesses a variety of therapeutic properties. However, it is suffered from its low water solubility and low bioavailability property, which seriously restricts its clinical application. In this study, we developed a glycyrrhetinic acid (GA) modified curcumin supramolecular pro-gelator (GA-Cur) and a control compound Nap-Cur by replacing GA with the naphthylacetic acid (Nap). Both compounds showed good water solubility and could form supramolecular gels by disulfide bond reduction triggered by glutathione (GSH) in vitro. Both formed gels could sustainedly release Cur in buffer solutions. We also investigated the cytotoxicity of pro-gelators to HepG2 cells by a MTT assay and determined the cellular uptake behaviours of them by fluorescence microscopy and LC-MS. Due to the over expression of GA receptor in liver cancer cells, our pro-gelator of GA-Cur showed an enhanced cellular uptake and better inhibition capacity to liver tumor cells than Nap-Cur. Therefore, the GA-Cur could significantly inhibit HepG2 cell growth. Our study provides a novel nanomaterial for liver tumor chemotherapy.

Effect of 18ß-glycyrrhetinic acid and hydroxypropyl γcyclodextrin complex on indomethacin-induced small intestinal injury in mice.

Non-steroidal anti-inflammatory drugs (NSAIDs)-induced small intestinal injury is a serious clinical event with recent advances of diagnostic technologies, but a successful therapeutic method to treat such injuries is still lacking. Licorice, a traditional herbal medicine, and its derivatives have been widely used for the treatment of a variety of diseases due to their extensive biological actions. However, it is unknown whether these derivatives have an effect on NSAIDs-induced small intestinal damage. Previously, the anti-inflammatory effects of three compounds extracted from the licorice root, glycyrrhizin, 18ß-glycyrrhetinic acid, and dipotassium glycyrrhizinate, were compared in vitro cell culture. The most prominent inhibitory effect on the tumor necrosis factor-α (TNF-α) production was observed with the administration of 18ß-glycyrrhetinic acid as an active metabolite of glycyrrhizin. In this study, a complex compound of 18ß-glycyrrhetinic acid and hydroxypropyl γcyclodextrin was examined to improve the oral bioavailability. After administration of this complex to indomethacin treated mice, a significantly high plasma concentration of 18ß-glycyrrhetinic acid was detected using the tandem mass spectrometry coupled with the HPLC. Furthermore, the complex form of 18ß-glycyrrhetinic acid and hydroxypropyl γcyclodextrin reduced mRNA expressions of TNF-α, interleukin (IL)-1ß, and IL-6, which was histologically confirmed in the improvement of indomethacin-induced small intestinal damage. These results suggest that the complex of 18ß-glycyrrhetinic acid and hydroxypropyl γcyclodextrin has the potential therapeutic value for preventing the adverse effects of indomethacin-induced small intestinal injury.

Synthesis and plasma pharmacokinetics in CD-1 mice of a 18ß-glycyrrhetinic acid derivative displaying anti-cancer activity.

OBJECTIVES: The plasma pharmacokinetic profile in CD-1 mice of a novel 18ß-glycyrrhetinic acid (GA) derivative, which displays in vitro anti-cancer activity, was assessed. METHODS: This study involved an original one-step synthesis of N-(2-{3-[3,5-bis(trifluoromethyl)phenyl]ureido}ethyl)-glycyrrhetinamide, (2) a compound that displays marked anti-proteasome and anti-kinase activity. The bioselectivity profile of 2 on human normal NHDF fibroblasts vs human U373 glioblastoma cells was assessed. Maximal tolerated dose (MTD) profiling of 2 was carried out in CD1 mice, and its serum pharmacokinetics were profiled using an acute intravenous administration of 40 mg/kg body weight. KEY FINDINGS: Compound 2 displayed IC(50) in vitro growth inhibitory concentrations of 29 and 8 µm on NHDF fibroblasts and U373 glioblastoma cells, respectively, thus a bioselectivity index of ∼4. The intravenous pharmacokinetic parameters revealed that 2 was rapidly distributed (t(1/2dist) of ∼3 min) but slowly eliminated (t(1/2elim) = ∼77 min). CONCLUSIONS: This study describes an original and reliable nanoemulsion of a GA derivative with both anti-proteasome and anti-kinase properties and that should be further tested in vivo using various human xenograft or murine syngeneic tumour models with both single and chronic intravenous administration.

HPLC assay and pharmacokinetics and tissue distribution study of glycyrrhetinic acid liposomes modified with galactosylated lipid.

The aim of this study was to evaluate the pharmacokinetics and tissue distribution of the glycyrrhetinic acid (GA) liposome modified with galactosylated lipid (NOH-GA-LP), compared with GA conventional liposome (GA-LP) and GA solution in mice. The pharmacokinetics and biodistribution of liposomal and solution formulation of GA in mice were studied after intravenous administration. Plasma and tissues were treated using liquid-liquid extraction and determined using reversed-phase high-performance liquid chromatography. Results showed that the mean residence times of NOH-GA-LP (2.99-fold) and GA-LP (2.94-fold) were higher than that of the GA solution in plasma. NOH-GA-LP produced a drug concentration in the liver that was markedly higher than that in other tissues and was approximately 2.0- and 4.8-fold of that of GA-LP and GA solution, respectively. In conclusion, the NOH-GA-LP prepared in this study is a promising sustained-release and drug-targeting system for antitumor drugs.

Final report on the safety assessment of Glycyrrhetinic Acid, Potassium Glycyrrhetinate, Disodium Succinoyl Glycyrrhetinate, Glyceryl Glycyrrhetinate, Glycyrrhetinyl Stearate, Stearyl Glycyrrhetinate, Glycyrrhizic Acid, Ammonium Glycyrrhizate, Dipotassium Glycyrrhizate, Disodium Glycyrrhizate, Trisodium Glycyrrhizate, Methyl Glycyrrhizate, and Potassium Glycyrrhizinate.

Glycyrrhetinic Acid and its salts and esters and Glycyrrhizic Acid and its salts and esters are cosmetic ingredients that function as flavoring agents or skin-conditioning agents - miscellaneous or both. These chemicals may be isolated from licorice plants. Glycyrrhetinc Acid is described as at least 98% pure, with 0.6% 24-OH-Glycyrrhetinic Acid, not more than 20 mu g/g of heavy metals and not more than 2 mu g/g of arsenic. Ammonium Glycyrrhizate has been found to be at least 98% pure and Dipotassium Glycyrrhizate has been found to be at least 95% pure. Glycyrrhetinic Acid is used in cosmetics at concentrations of up to 2%; Stearyl Glycyrrhetinate, up to 1%; Glycyrrhizic Acid, up to 0.1%; Ammonium Glycyrrhizate, up to 5%; Dipotassium Glycyrrhizate, up to 1%; and Potassium Glycyrretinate, up to 1%. Although Glycyrrhizic Acid is poorly absorbed by the intestinal tract, it may be hydrolyzed to Glycyrrhetinic Acid by a beta -glucuronidase produced by intestinal bacteria. Glycyrrhetinic Acid and Glycyrrhizic Acid bind to rat and human albumin, but do not absorb well into tissues. Glycyrrhetinic Acid and Glycyrrhizic Acid and metabolites are mostly excreted in the bile, with very little excreted in urine. Dipotassium Glycyrrhizate was undetectable in the receptor chamber when tested for transepidermal permeation through pig skin. Glycyrrhizic Acid increased the dermal penetration of diclofenac sodium in rat skin. Dipotassium Glycyrrhizate increased the intestinal absorption of calcitonin in rats. In humans, Glycyrrhetinic Acid potentiated the effects of hydrocortisone in the skin. Moderate chronic or high acute exposure to Glycyrrhizic Acid, Ammonium Glycyrrhizate, and their metabolites have been demonstrated to cause transient systemic alterations, including increased potassium excretion, sodium and water retention, body weight gain, alkalosis, suppression of the renin-angiotensis-aldosterone system, hypertension, and muscular paralysis; possibly through inhibition of 11beta -hydroxysteroid dehydrogenase-2 (11beta -OHSD2) in the kidney. Glycyrrhetinic Acid and its derivatives block gap junction intracellular communication in a dose-dependent manner in animal and human cells, including epithelial cells, fibroblasts, osteoblasts, hepatocytes, and astrocytes; at high concentrations, it is cytotoxic. Glycyrrhetinic Acid and Glycyrrhizic Acid protect liver tissue from carbon tetrachloride. Glycyrrhizic Acid has been used to treat chronic hepatitis, inhibiting the penetration of the hepatitis A virus into hepatocytes. Glycyrrhetinic Acid and Glycyrrhizic Acid have anti-inflammatory effects in rats and mice. The acute intraperitoneal LD(50) for Glycyrrhetinic Acid in mice was 308 mg/kg and the oral LD(50) was > 610 mg/kg. The oral LD(50) in rats was reported to be 610 mg/kg. Higher LD(50) values were generally reported for salts. Little short-term, subchronic, or chronic toxicity was seen in rats given ammonium, dipotassium, or disodium salts of Glycyrrhizic Acid. Glycyrrhetinic Acid was not irritating to shaved rabbit skin, but was considered slightly irritating in an in vitro test. Glycyrrhetinic Acid inhibited the mutagenic activity of benzo[a]pyrene and inhibited tumor initiation and promotion by other agents in mice. Glycyrrhizic Acid inhibited tumor initiation by another agent, but did not prevent tumor promotion in mice. Glycyrrhizic Acid delayed mortality in mice injected with Erlich ascites tumor cells, but did not reduce the mortality rate. Ammonium Glycyrrhizate was not genotoxic in in vivo and in vitro cytogenetics assays, the dominant lethal assay, an Ames assay, and heritable translocation tests, except for possible increase in dominant lethal mutations in rats given 2000 mg/kg day(-1) in their diet. Disodium Glycyrrhizate was not carcinogenic in mice in a drinking water study at exposure levels up to 12.2 mg/kg day(-1) for 96 weeks. Glycyrrhizate salts produced no reproductive or developmental toxicity in rats, mice, golden hamsters, or Dutch-belted rabbits, except for a dose-dependent increase (at 238.8 and 679.9 mg/kg day(-1)) in sternebral variants in a study using rats. Sedation, hypnosis, hypothermia, and respiratory depression were seen in mice given 1250 mg/kg Glycyrrhetinic Acid intraperitoneally. Rats fed a powdered diet containing up to 4% Ammonium Glycyrrhizate had no treatment related effects in motor function tests, but active avoidance was facilitated at 4%, unaffected at 3%, and depressed at 2%. In a study of 39 healthy volunteers, a no effect level of 2 mg/kg/day was determined for Glycyrrhizic Acid given orally for 8 weeks. Clinical tests in seven normal individuals given oral Ammonium Glycyrrhizate at 6 g/day for 3 days revealed reduced renal and thermal sweat excretion of Na+ and K+, but carbohydrate and protein metabolism were not affected. Glycyrrhetinic Acid at concentrations up to 6% was not a skin irritant or a sensitizer in clinical tests. Neither Glycyrrhizic Acid, Ammonium Glycyrrhizate, nor Dipotassium Glycyrrhizate at 5% were phototoxic agents or photosensitizers. Birth weight and maternal blood pressure were unrelated to the level of consumption of Glycyrrhizic Acid in 1049 Finnish women with infants, but babies whose mother consumed > 500 mg/wk were more likely to be born before 38 weeks. The Cosmetic Ingredient Review (CIR) Expert Panel noted that the ingredients in this safety assessment are not plant extracts, powders, or juices, but rather are specific chemical species that may be isolated from the licorice plant. Because these chemicals may be isolated from plant sources, however, steps should be taken to assure that pesticide and toxic metal residues are below acceptable levels. The Panel advised the industry that total polychlorobiphenyl (PCB)/pesticide contamination should be limited to not more than 40 ppm, with not more than 10 ppm for any specific residue, and that toxic metal levels must not contain more than 3 mg/kg of arsenic (as As), not more than 0.002% heavy metals, and not more than 1 mg/kg of lead (as Pb). Although the Panel noted that Glycyrrhizic Acid is cytotoxic at high doses and ingestion can have physiological effects, there is little acute, short-term, subchronic, or chronic toxicity and it is expected that these ingredients would be poorly absorbed through the skin. These ingredients are not considered to be irritants, sensitizers, phototoxic agents, or photosensitizers at the current maximum concentration of use. Accordingly, the CIR Expert Panel concluded that these ingredients are safe in the current practices of use and concentration. The Panel recognizes that certain ingredients in this group are reportedly used in a given product category, but the concentration of use is not available. For other ingredients in this group, information regarding use concentration for specific product categories is provided, but the number of such products is not known. In still other cases, an ingredient is not in current use, but may be used in the future. Although there are gaps in knowledge about product use, the overall information available on the types of products in which these ingredients are used and at what concentration indicate a pattern of use. Within this overall pattern of use, the Expert Panel considers all ingredients in this group to be safe.

A population physiologically based pharmacokinetic/pharmacodynamic model for the inhibition of 11-beta-hydroxysteroid dehydrogenase activity by glycyrrhetic acid.

Glycyrrhizic acid is widely applied as a sweetener in food products and chewing tobacco. Habitual consumption of this compound may lead to hypertension and electrolyte disturbances due to inhibition of 11-beta-hydroxysteroid dehydrogenase by the metabolite glycyrrhetic acid. The effect of 130 mg glycyrrhetic acid/day for 5 days on 11-beta-hydroxysteroid dehydrogenase activity was studied by measuring the cortisol-cortisone ratio in 24-h urine. A twofold increase in this ratio was observed. It took 4 days for the elevated urinary cortisol-cortisone ratio to return to the baseline ratio after cessation of the treatment. The pharmacokinetics of glycyrrhetic acid were studied after the first and last dose. Using data from a previously performed single-dose study and present multiple-dose treatment, a physiologically based pharmacokinetic model for glycyrrhetic acid was developed. The variability of the pharmacokinetics of glycyrrhetic acid in the population studied could be explained for a considerable part by interindividual differences in gastrointestinal transit of glycyrrhetic acid metabolites. The relationship between glycyrrhetic acid exposure and changes in urinary cortisol-cortisone ratio was described by a pharmacodynamic model, using nonlinear mixed-effect modeling. Literature data on the inhibitory effect of glycyrrhetic acid on 11-beta-hydroxysteroid dehydrogenase activity under various exposure scenarios could be adequately described by the model. Due to the relationship between the pharmacokinetics of glycyrrhetic acid and its inhibitory effect on 11-beta-hydroxysteroid dehydrogenase activity, reflected by a change in the urinary cortisol-cortisone ratio, this ratio might serve as a noninvasive marker to identify individuals at risk for glycyrrhizic acid over-consumption.

Physiologically based pharmacokinetic modeling of glycyrrhizic acid, a compound subject to presystemic metabolism and enterohepatic cycling.

Glycyrrhizic acid is currently of clinical interest for treatment of chronic hepatitis. It is also applied as a sweetener in food products and chewing tobacco. In some highly exposed subgroups of the population, serious side effects such as hypertension and electrolyte disturbances have been reported. In order to analyze the health risks of exposure to this compound, the kinetics of glycyrrhizic acid and its active metabolites were evaluated quantitatively. Glycyrrhizic acid and its metabolites are subject to complex kinetic processes, including enterohepatic cycling and presystemic metabolism. In humans, detailed information on these processes is often difficult to obtain. Therefore, a model was developed that describes the systemic and gastrointestinal tract kinetics of glycyrrhizic acid and its active metabolite glycyrrhetic acid in rats. Due to the physiologically based structure of the model, data from earlier in vitro and in vivo studies on absorption, enterohepatic cycling, and presystemic metabolism could be incorporated directly. The model demonstrates that glycyrrhizic acid and metabolites are transported efficiently from plasma to the bile, possibly by the hepatic transfer protein 3-alpha-hydroxysteroid dehydrogenase. Bacterial hydrolysis of the biliary excreted metabolites following reuptake of glycyrrhetic acid causes the observed delay in the terminal plasma clearance of glycyrrhetic acid. These mechanistic findings, derived from analysis of experimental data through physiologically based pharmacokinetic modeling, can eventually be used for a quantitative health risk assessment of human exposure to glycyrrhizic acid containing products.