Compatibility with Semen Sojae Praeparatum attenuates hepatotoxicity of Gardeniae Fructus by regulating the microbiota, promoting butyrate production and activating antioxidant response.

BACKGROUND: Herb-induced liver injury is a leading cause of drug-induced liver injury in China and its incidence is also increasing worldwide. Gardeniae Fructus (ZZ) has aroused wide concern for hepatotoxicity in recent decades. But when ZZ is administered in combination with Semen Sojae Praeparatum (DDC) to compose a herbal pair Zhizichi Decoction (ZZCD), lower hepatotoxicity is observed. The mechanism involved in the attenuated effect remains to be investigated. HYPOTHESIS/PURPOSE: Our previous studies showed that DDC benefited host metabolism by regulating the gut microbiota and it reduced the exposure of major toxic components of ZZ. The present study was aimed to investigate how DDC attenuated hepatotoxicity of ZZ from the perspective of gut microbiota. METHODS: Rats received ZZ and ZZCD treatment of different dosages and antibiotic treatment was applied to explore the involvement of gut microbiota. Biochemical assays and histopathological analysis were conducted to evaluate liver injury. Gut microbiota in caecal contents was profiled by 16S rRNA sequencing. Short-chain fatty acids (SCFAs) in caecal contents were measured by gas chromatography mass spectrometry (GCMS). To verify the protective effect of butyrate, it was administered with genipin, the major hepatotoxic metabolite of ZZ, to rats and HepG2 cells. Plasma lipopolysaccharide (LPS) level and colon tissue section were used to evaluate gut permeability. Expression level of Nuclear factor erythroid-derived 2-like 2 (Nrf2) was detected by immunohistochemistry in vitro and by western blot in vivo. RESULTS: Our study showed that ZZCD displayed lower hepatotoxicity than ZZ at the same dosage. ZZ induced gut dysbiosis, significantly reducing Lactobacillus and Enterococcus levels and increasing the Parasutterella level. In combination with DDC, these alterations were reversed and beneficial genus including Akkermansia and Prevotella were significantly increased. Besides, butyrate production was diminished by ZZ but was restored when in combination with DDC. Butyrate showed detoxification on genipin-induced liver injury by promoting colon integrity and promoting Nrf2 activation. Besides, it protected genipin-induced hepatocyte damage by promoting Nrf2 activation. CONCLUSION: DDC attenuates ZZ-induced liver injury by regulating the microbiota, promoting butyrate production and activating antioxidant response.

Acute and Subchronic Oral Toxicity Study of Gardenia Yellow E500 in Sprague-Dawley Rats.

OBJECTIVE: This study was conducted to evaluate the acute and subchronic toxicity of gardenia yellow, a natural colorant widely used in China and other Asian countries. An acute toxicity test was performed in S-D rats of both genders and the lethal dose (LD50) of per oral gardenia yellow was estimated to be more than 15.0 g/kg·bw. In the subchronic study, gardenia yellow was orally administered to rats by gavage at doses of 0, 0.50, 1.50 and 4.50 g/kg·bw/day for 90 days followed by a recovery period of 28 days. No appreciable toxic-related changes were observed in the 0.50 g/kg·bw/day group. When the animals received gardenia yellow at 1.50 g/kg·bw/day or more, body weight loss was observed, and pigments began to deposit in several vital organs, resulting in significant changes of several hematological and biochemical indicators related to the nutritional status of the body, liver and kidney function, more severe in the high dose group. In the recovery period, the alterations of the clinical symptoms and parameters were relieved a lot. Based on the results of the current study, the no observed adverse effect level (NOAEL) of gardenia yellow E500 in rats was set to be 0.50 g/kg·bw/day.

Synthesis of porous starch xerogels modified with mercaptosuccinic acid to remove hazardous gardenia yellow.

Mercaptosuccinic acid (MSA) molecules were inserted into potato starch, leading to the breaking of intrinsic H-bonds within macromolecular chains of starch and the formation of intermolecular H-bonds between MSA and starch, which could be verified by Fourier transform infrared spectroscopy (FT-TR). MSA modified porous starch xerogels (PSX/MSA) were obtained after freeze-drying the MSA modified starch, and they were characterized by field emission scanning electron microscopy (FESEM), exhibiting the intriguing porous structure due to the separation of starch chains by MSA molecules. The PSX/MSA were then used as the adsorbents to remove gardenia yellow (GY), a natural colorant with genotoxicity. Due to the porous structure of PSX and the introduced carboxyl groups from MSA, the adsorption capacity of the PSX/MSA was much higher than that of the starch xerogels alone (SX). The adsorption behaviors of GY by the PSX/MSA fitted both the Freundlich isotherm model and the pseudo-second-order kinetic model, and the efficient adsorption of GY suggested that the PSX/MSA might be potential adsorbents for the removal of dyes from contaminated aquatic systems.

Clinical Search for Undiagnosed Mesenteric Phlebosclerosis at Outpatient Departments Specializing in Herbal (Kampo) Medicine.

OBJECTIVE: Mesenteric phlebosclerosis (MP) is a disease characterized by calcification of the mesenteric vein, which causes chronic mesenteric ischemia. Recently, the long-term intake of gardenia fruit ('Sanshishi' in Japanese) has been attracting attention as a possible cause. Usually, only advanced, severe MP cases get reported. However, we suspected that some latent cases of this disease may exist. We performed this study in order to determine the prediagnostic cases at our outpatient departments of herbal (Kampo) medicine, with particular attention paid to the initial changes, such as any slight color change of the colon, as shown in colonoscopy. METHODS: We recommend colonoscopy and computed tomography (CT) scans for patients with a long-term history of taking herbal medicines containing gardenia fruit. Clinical examinations were performed upon receiving patients' consent from December 2013 to November 2014. RESULTS: Of the 103 patients who took gardenia fruit long-term, 29 agreed to be checked for MP. 14 patients underwent colonoscopy. Four patients were confirmed to have MP due to the presence of fibrotic deposition of the colonic membrane on histological inspection. Twenty-one patients underwent abdominal CT screening. Characteristic calcification of the mesenteric vein was observed on CT scans in 2 patients. All 4 MP patients took Kampo formulas containing gardenia fruit for more than 6.8 years. The other patients did not develop MP, despite long-term gardenia fruit intake. CONCLUSION: We detected the latent and undiagnosed MP cases. All diagnoses were made while paying careful attention to any slight changes in colonoscopy and CT scans.

[Comparative study on hepatic toxicity of gardeniae fructus and Huanglian Jiedu decoction].

OBJECTIVE: To observe the effect of ingredients in Huanglian Jiedu decoction (HLJDT) combined with Gardeniae Fructus on the hepatic toxicity of Gardeniae Fructus and its mechanism. METHOD: Rats were given Gardeniae Fructus and HLJDT decoction at the dose of 10 times of clinical dosage for 3 days. Their ALT AST, ALP, TBA were detected, and their liver weight index was calculated. SOD activity, MDA content, GSH-PX activity, TNF-alpha content in hepatic tissues were determined. The cell apoptosis in liver tissue was determined by TUNEL, and the expressions of apoptosis related proteins Bax, Bcl-2 were measured by immunohistochemical method. RESULT: Compared with the normal control group, the Gardeniae Fructus group showed significant increase in the liver weight index, ALT, AST, TBA and ALP, notable decrease in SOD, SOD/MDA and GSH-PX, and remarkable rise in MDA, TNF-a concentration, accumulated optical density, apoptosis index, Bax and Bax/Bcl-2. Compare with that in the Gardeniae Fructus group, the liver index, ALT, AST, TBA, ALP reduced obviously; SOD, SOD/MDA and GSH-PX markedly increased; MDA and TNF-alpha significantly reduced; the accumulated optical density and apoptosis index significantly reduced; and Bax/Bcl-2 was much lower in HLJDT group. CONCLUSION: The hepatic toxicity caused by Gardeniae Fructus may be related to inflammation, oxidative stress-induced hepatocyte necrosis and apoptosis. Other ingredients in HLJDT, apart from Gardeniae Fructus, can decrease the hepatic toxicity caused by Gardeniae Fructus by increasing the enzyme activity eliminating radicals and inhibiting hepatocyte injury caused by inflammatory reaction against Gardeniae Fructus.

A 13-week oral dose subchronic toxicity study of gardenia yellow containing geniposide in rats.

Gardenia yellow powders A, B and C, containing geniposide at 0.284%, 0.938% and 2.783%, respectively, were administered orally to male and female SD rats as 3% feed admixtures for 13-weeks to evaluate any potential toxicity. Mean geniposide intake values were 5.72, 18.9 and 56.3mg/kg/day in groups receiving these feed admixtures, respectively. All animals survived the duration of the study. The following findings were evident in the gardenia yellow C group: chromatouria, slightly increased plasma total bilirubin, blackish brown discoloration of the kidneys and liver, brown pigments in the proximal tubular epithelium of the kidneys. Slightly increased plasma total bilirubin was considered to be due to interference of metabolite of geniposide with the system of measurement and not to be a toxic effect since there were no related changes in histopathology of the liver or in any blood chemistry parameters. Other findings were limited to pigmentations or discolorations attributable to metabolites of geniposide. No treatment-related effects were evident on body weight, food consumption, ophthalmology, hematology or organ weights in any group. Therefore, it was concluded that 3-month ingestion of the gardenia yellow powder containing geniposide at 2.783% (approximately 60 mg/kg/day as geniposide intake) does not cause any severe toxic effects.

[Research and development of Fructus Gardeniae].

Survey on research and development of Fructus Gardeniae in the recent 10 years. Gardenia yellow has been used for food colorent, medicine, feedingstuff and cosmetic. Garnedia blue has been used for developing another pigment with red and yellow. Fructus Gardeniae has been used in digestive system for cholecyst constracting and gall-stone eliminating, for declining peroxide on SAP mouse and increasing immune ability, for protecting liver against cancel, anti-acetylcholinic restraining on stomach enginery, in cardiovascular system it has been used for centrally anti-hypertension, preventing atheroma and thrombus, also Fructus Gardeniae has been used for anti-inflammation, treating parenchyma injure etc. Geniposide used for increasing production in agriculture has wider perspect.

Investigation of the antimutagenic effects of selected South African medicinal plant extracts.

Dichloromethane extracts from different parts of Rhamnus prinoides, Ornithogalum longibracteatum, Gardenia volkensii, Spirostachys africana, Diospyros whyteana, Syzigium cordatum and Prunus africana were investigated for mutagenic and antimutagenic effects in Salmonella/microsome and micronucleus tests. None of the extracts tested in the Ames test were found to induce mutations or to modify the effect of the mutagen 4-nitroquinoline-oxide (4NQO). In the micronucleus test, extracts from twigs/bark of R. prinoides, twigs of D. whyteana, P. africana and S. cordatum significantly lowered the effect of the mutagen mitomycin C (MMC). Extracts from twigs/bark of G. volkensii and S. africana were genotoxic in the micronucleus test, while extracts of O. longibracteatum leaves potentiated the genotoxicity of MMC. This preliminary investigation shows that plant extracts used in traditional medicine may have particular effects with regard to mutagenicity and antimutagenicity indicating careful use in some instances and the need to isolate their active principles for further research.