Optimization of parameters for stir-frying of Kansui Radix with vinegar based on conversion of toxic components / 中国中药杂志

This study aims to optimize the parameters for stir-frying of Kansui Radix with vinegar based on the conversion of representative toxic diterpenes, which is expected to serve as a reference for the standardized production of Kansui Radix stir-fried with vinegar. To be specific, the toxic components [3-O-(2'E,4'Z-decadienoyl)-20-O-acetylingenol(3-O-EZ), kansuiphorin C(KPC)] in Kansui Radix and the products(ingenol, 20-deoxyingenol) after the stir-frying with vinegar were selected. The toxicity to intestine and water-draining activity were evaluated with NCM460(normal human colon mucosal epithelial cell line) and HT-29(a human colorectal adenocarcinoma cell line). An HPLC method was then developed to assess the conversion of toxic components. On this basis, temperature, time, and amount of vinegar for the processing of Kansui Radix were optimized with the Box-Behnken design and the content of ingenol and 20-deoxyingenol as evaluation index. The results showed that after the stir-frying of Kansui Radix with vinegar, 3-O-EZ and KPC were first converted to monoester 3-O-(2'E,4'Z-decadienoyl)ingenol(3-EZ) and 5-O-benzoyl-20-deoxyingenol(5-O-Ben) and finally to almost non-toxic ingenol and 20-deoxyingenol, respectively. Meanwhile, the water-draining activity was retained. Six compounds had a good linear relationship with the peak area in the corresponding concentration ranges(R~2≥0.999 8), and the average recovery fell in the range of 98.20%-102.3%(RSD≤2.4%). The content of representative diterpenes and intermediate products was 14.78%-24.67% lower in the Kansui Radix stir-fried with vinegar than in the Kansui Radix, while the content of the conversed products was 14.37%-71.37% higher. Among the process parameters, temperature had significant influence on the total content of products, followed by time. The optimal parameters were 210 ℃, 15 min, and 30% vinegar. The relative error between the experimental results and the predicted values was 1.68%, indicating that the process was stable and reproducible. The strategy of screening optimal parameters for stir-frying of Kansui Radix with vinegar based on the transformation of toxic components can help improve the production stability, reduce the toxicity, and ensure the efficacy of Kansui Radix stir-fried with vinegar, which can serve as a reference for the process optimization of similar toxic Chinese medicinals.

Effect of Kansui Radix stir-fried with vinegar on fecal metabolomics in Walker-256 rats with ascites / 中国中药杂志

Utilizing metabolomics technology, this study explored the change of fecal endogenous metabolites in Walker-256 rats with malignant ascites after the administration with Kansui Radix(KR) stir-fried with vinegar(VKR), sought the potential biomarkers in feces which were related to the treatment of malignant ascites by VKR and revealed the biological mechanism of water-expelling effect of VKR. Ultra-fast liquid chromatography-quadrupole-time-of-flight mass spectrometry(UFLC-Q-TOF-MS) was employed to detect the feces of rats in all groups. Principle component analysis(PCA) and partial least squares discriminant analysis(PLS-DA) were conducted to achieve pattern recognition. Combining t-test and variable importance in the projection(VIP) enabled the screening of potential biomarkers for the malignant ascites. Metabolic pathway analysis was accomplished with MetaboAnalyst. Correlation analysis was finally conducted integrating the sequencing data of gut microbiota to elucidate the mechanism underlying the water-expelling effect of VKR. The results showed that both KR and VKR could restore the abnormal metabolism of model rats to some extent, with VKR being inferior to KR in the regulation. Eleven potential biomarkers were identified to be correlated with the malignant ascites and five metabolic pathways were then enriched. Four kinds of gut microbiota were significantly related to the potential biomarkers. The water-expelling effect of VKR may be associated with the regulation of phenylalanine metabolism, biosynthesis of phenylalanine, tyrosine and tryptophan, tryptophan metabolism, glycerophospholipid metabolism, and glycosylphosphatidylinositol(GPI)-anchor biosynthesis. This study can provide a scientific basis for comprehensive understandings of the interaction between gut microbiota and host which has relation to the water-expelling effect of VKR and guide the reasonable clinical application of VKR.

Chemical ingredient groups B and C in Kansui Radix stir-fried with vinegar affect diversity of gut microbiota in rat model of malignant ascites induced by Walker-256 cells / 中国中药杂志

This study aims to explore the effects of chemical ingredient groups B and C in Kansui Radix stir-fried with vinegar on the diversity of gut microbiota in the rat model of malignant ascites, identify the key differential microbial taxa, and reveal the biological mechanism of water-expelling effect of the two chemical ingredient groups. The rat model of malignant ascites induced by Walker-256 cells was established, and phenolphthalein was used as the positive drug. The rats were orally administrated with corresponding agents for consecutive 7 days. On day 6, fresh feces samples were collected from the rats, and 16 S rDNA high-throughput sequencing and GC-MS were employed to determine the composition of gut microbiota and the content of short-chain fatty acids, respectively. On day 7, serum and intestinal tissue samples were collected for the determination of related indicators. Compared with the control group, the model group showed decreased feces volume and urine volume(P<0.01), increased volume of ascites and levels of Na~+, K~+, and Cl~- in urine(P<0.01), down-regulated mRNA and protein levels of intestinal AQP8(P<0.01), lowered abundance of beneficial Lactobacillus(P<0.01) while risen abundance of potential pathogenic Lachnospiraceae and Anaeroplasma(P<0.01), and reduced content of short-chain fatty acids(P<0.01). Compared with the model group, administration with chemical ingredient groups B and C alleviated all the above indicators(P<0.01). In conclusion, chemical ingredient groups B and C in Kansui Radix stir-fried with vinegar could alleviate the disordered gut microbiota in rats with malignant ascites to expel water through increasing the abundance of beneficial Lactobacillus and reducing the abundance of harmful Lachnospiraceae and Anaeroplasma. This study can provide a reference for the reasonable clinical application of Kansui Radix stir-fried with vinegar.

Preliminary investigation on processing mechanism of Kansui Radix by liquorice based on thermal analysis technology / 中国中药杂志

In the thermal analysis, the pyrolysis characteristics of crude Kansui Radix, alcohol extract of Kansui Radix, petroleum ether extract, chloroform extract, ethyl acetate extract, n-butanol extract, and licorice vinegar were analyzed with simulated air (N₂-O₂ 4:1) as the carrier gas, at a temperature increase rate of 10 °C·min⁻¹ and a volume flow rate of 60 mL·min⁻¹, respectively. The results showed that due to the different polarity of the extraction solvent, the type and quantity of the chemical components contained in each polar part were different, and with the increase in the amount of solid powder of licorice, the peak of the maximum heat loss rate occurred in advance. For petroleum ether, chloroform, and ethyl acetate fractions, (157.40±1.06), 3.50, (25.83±1.66) °C in advance respectively, but the weight loss rate of the chloroform fraction was increased by (2.62±5.19) °C, while decreased by (33.90±1.72), (19.28±1.11) °C for the petroleum ether and ethyl acetate fractions. So we can conclude that with the addition of licorice, the pyrolysis rate of the petroleum ether and chloroform fractions in the toxic part of Kansui Radix was increased; the temperature point at the peak of the maximum weight loss rate was decreased, and the ethyl acetate fraction (effective part) showed a decrease in temperature rising process, but its overall ratio of weight loss and weight loss rate were relatively small, retaining the effect of medicinal ingredients. This proved the mechanism of licorice system Kansui Radix on attenuating toxicity after processing and the scientificity and rationality of licorice system Kansui Radix. At the same time, as the proportion of glycyrrhizin was increased, the peak of the maximum heat loss rate of petroleum ether, chloroform and ethyl acetate fractions occurred in advance; the peak temperature was decreased, with easy pyrolysis. Among them, the thermogravimetric rate of the mixture of petroleum ether and chloroform fractions (10:1) was relatively large, with a low peak temperature, while ethyl acetate fraction showed opposite results. This conclusion has certain guiding significance for the ratio of gansui to licorice.

Compatibility Rule of Herb-pair in Rhubarb Based on Anti-inflammation Function / 医药导报

Objective To study compatibility rule of herb-pair in rhubarb after compatibilities with fructus aurantii Immaturus,Chinese goldthread rhizome,moutan cortex,peach seed and kansui radix stir-baked with vinegar, and to observe its anti-inflammation and acute toxicity. Methods Mice were gavaged by 15,30 g?kg-1 rhubarb and rhubarb, rhubarb frutus aurantii immaturus rhizoma coptidis, radix et rhizoma rhei peony, rhubarb, semen persicae and rhubarb extracts from euphorbia kansui, respectively, in the morning and evening once, for 7 d.The effects of five different compatibility of Rhubarb on acute inflammation were observed in the mouse paw swelling induced by carrageenan. The classical method were used to determine acute toxicity of rhubarb and the contents of five different compatibility of rhubarb. Results Compared with the control group, the contents of five different compatibility of rhubarb with high and low dosage groups(30,15 g?kg-1 ) could inhibit the paw edema in mice,reduce NO and MDA production and enhanced activity of SOD in mice inflammatory tissue. The LD50 was not determined. Calculated by crude drug content, the MLD of rhubarb compatibilities with fructus aurantii immaturus, Chinese goldthread rhizome,moutan cortex,peach seed and kansui radix stir-baked with vinegar were 145.33,142.30,117.53,103.45, 113.09,182.36 g?kg-1 respectively, which were respectively equal to 581,569,470,418,452 ,729 times of people, s daily dried medicinal herb dosage. Conclusion The five different herb-pair have anti-inflammation effects after compatibility of Rhubarb, and it got best effects when rhubarb compatibility with kansui radix stir-baked, the next were rhubarb compatibility with Chinese goldthread rhizome and moutan cortex. It was basic security and low toxicity of herb-pair in rhubarb after compatibilities.

Dosage-toxicity-efficacy relationship of Kansui Radix in malignant pleural effusion models based on Walker 256 analysis / 中国中药杂志

To investigate the dosage-toxicity-efficacy relationship of Kansui Radix, explore its regularity of the toxicity and efficacy change, and provide scientific basis for its clinical rational application, the malignant pleural effusion models were used to observe the effect of Kansui Radix with larger dose range (0.045-1.620 g•kg•d⁻¹ for Kansui Radix) on biochemical indexes and hydrothorax volume in experimental animals. Factor analysis method was also used to comprehensively assess the dosage-toxicity-efficacy relationship of Kansui Radix. The results showed that the rats in model group had larger hydrothorax volume, and ALT, AST, LDH, HBDH, IFN-γ, IL-2 and TNF-α levels were significantly increased (P<0.05), while TP and ALB levels were decreased (P<0.05) as compared with the blank group. After drug administration, various treatment groups decreased hydrothorax volume, IFN-γ, IL-2, TNF-α and increased TP and ALB levels as compared with model group, indicating certain therapeutic effect; and increased ALT, AST, LDH and HBDH levels, indicating certain liver and cardiac toxicity. In the factor analysis, two common factors were extracted from nine indexes, explaining 89.1% of the information. The analysis results suggested that there was no obvious toxicity in case of independent use of Kansui Radix within the dosage range set in pharmacopeia, while it would produce liver toxicity and cardiac toxicity upon 3 times of the dosage set in pharmacopeia, and the toxicity was increased with the increase of dose. At the same time, Kansui Radix can decrease the hydrothorax volume in malignant pleural effusion models and improve relevant physical indexes in a dose-dependent manner. Comprehensive analysis results of its toxic effect characteristics indicated that the upper-limit dose of Kansui Radix in pharmacopeia shall be regarded as the relatively optimal therapeutic dose.

Comparative study on expelling water retention with drastic purgative of crude and vinegar stir-baked Kansui Radix in cancerous ascites rats / 中草药

Objective: To compare the function of expelling water retention with drastic purgative of crude and vinegar stir-baked Kansui Radix in cancerous ascites model rats. Methods: Furosemide was taken as positive control drug, and the cancerous ascites model rats were respectively orally administered with powder of crude and vinegar stir-baked Kansui Radix and their alcohol and water extract for 7 d. The amounts of urine and ascites, the levels of urinary sodium, potassium, chlorideion, and pH value, and the contents of PRA, Ang II, ALD, and ADH in serum were investigated. Results: Compared with model group, the amount of urine of each medication administration group significantly increased (P < 0.05, 0.01), the amount of ascites, the levels of urinary sodium, potassium, chlorideion, and pH value (P < 0.01), and the contents of PRA, Ang II, ALD, and ADH in serum all showed a significant decrease (P < 0.05, 0.01). Among them, the groups which were administered with powder of crude and vinegar stir-baked Kansui Radix were the most significant, and there was no significant difference between the two groups. Conclusion: The powders of crude and vinegar stir-baked Kansui Radix have a remarkable effect on expelling water retention with drastic purgative, and they could improve the symptom of cancerous ascites model rats.

Comparative effects of different extracts of Kansui Radix stir-baked with vinegar on function of expelling water retention with drastic purgative in cancerous ascites model rats / 中草药

Objective: To study the effects of different extracts of Kansui Radix stir-baked with vinegar (KRV) on the function of expelling water retention with drastic purgative in cancerous ascites model rats. Methods: The cancerous ascites model rats were respectively ig administered with KRV powder, ethanol extract, aqueous extract, and ehanol and aqueous extract of KRV (340 mg/kg) for 7 d. The amounts of urine and ascites, the levels of urinary sodium, potassium, chloride ion, and pH value, and the contents of PRA, Ang II, and ALD in serum were investigated. UPLC-QTOF MS technology was used to explore the components differences in various extracts of KRV. Results: Compared with the control group, the amount of urine in model group was significantly reduced (P<0.05), the ascites generated, and the urinary sodium, potassium, chloride ion, pH value, and the contents of PRA, Ang II, and ALD in serum were all significantly increased (P<0.01). Compared with the model groups, the treatment groups showed decreasing trend in ascites; The amounts of urine in positive groups, powder groups, ethanol and aqueous extract groups showed a significant increase (P<0.05); The level of urinary sodium of water extraction groups showed significant decrease (P<0.05); The levels of urinary sodium, potassium, chloride ion, pH value, and the contents of PRA, Ang II, and ALD in serum of positive groups, powder groups, ethanol extract groups, and ethanol and aqueous extract groups all showed a significant decrease (P<0.05, 0.01). Diterpenes were inspected in the alcohol extract and alcohol and aqueous extract, fewer in the aqueous extract. Conclusion: Powder groups and ethanol and aqueous extract groups of KRV have remarkable effect on expelling water retention with drastic purgative, and there is no significant difference between the two groups, which could provide the basis for clinical medication of KRV that is made into the pill and powder. Diterpenes in KRV may be the active components on the function of expelling water retention with drastic purgative.

Attenuation by stir baking with vinegar on effect of ethyl acetate fraction from Kansui Radix on mice gastrointestinal permeability / 中草药

Objective: To study the mechanism of attenuation by stir baking with vinegar on effect of ethyl acetate fraction of Kansui Radix on mice gastrointestinal permeability. Methods: Mice were randomly divided into blank control, Kansui Radix, and Kansui Radix stir-baked with vinegar groups. The gastric and intestinal tissues were removed from mice after ig administration for 7 d and were observed by the transmission electron microscope (TEM). The expression of E-cadherin and VCAM-l was examined by immunohistochemical techniques. Results: Using TEM, the arrangement and karyon morphology of gastric glandular epithelium cells were irregular with the uneven electronic of cytoplasm and lots of lysosomes in Kansui Radix group; The morphology of the gastric epithelial cells was irregular with some abnormal mitochondrions in the cytoplasm in Kansui Radix stir-baked with vinegar group; The intestinal mucosa cells were covered with short microvilli with well-connected intercellular junctions, but the tight junctions on the near-surface were short in Kansui Radix group; The arrangement and morphology of intestinal epithelial cells were fairly uniform with few and short microvilli and intact mitochondrion in Kansui Radix stir-baked with vinegar group. Using immunohistochemical techniques, compared with the control group, Kansui Radix could decrease the expression of E-cadherin and increase the expression of VCAM-l (P < 0.05, 0.01). While compared with Kansui Radix group, Kansui Radix stir-baked with vinegar could obviously increase the expression of E-cadherin (P < 0.05, 0.01) and decrease the expression of VCAM-l (P < 0.05, 0.01). Conclusion: Stir-baking with vinegar can attenuate the Kansui Radix-induced gastrointestinal inflammation. The mechanism may be that it can improve the morphology of mice gastrointestinal mucosal cells, reduce the membrane permeability, and decrease the stimulating effect of Kansui Radix.