Quality markers of Zingiberis Rhizoma Carbonisata before and after processing / 中国中药杂志

Based on the previous research results of our group and literature research, the chemical components, mechanisms, pharmacodynamics, and pharmacokinetics of Zingiberis Rhizoma Carbonisata were summarized to determine the quality markers(Q-markers) of Zingiberis Rhizoma Carbonisata and Zingiberis Rhizoma. Our research group has clarified the differential components of Zingiberis Rhizoma Carbonisata and Zingiberis Rhizoma, the meridian-warming hemostatic effect of Zingiberis Rhizoma Carbonisata, the related targets and pathways of the effect, the endogenous biomarkers of Zingiberis Rhizoma Carbonisata, and the hemodynamic processes of Zingiberis Rhizoma Carbonisata and Zingiberis Rhizoma. Moreover, based on high-performance liquid chromatography-diode array detector-electrospray ionization mass spectrometry(HPLC-DAD-ESIMS), a method for determining the content of Q-mar-kers was established. In conclusion, the study finally determined that gingerone, 6-shogaol, and diacetyl-6-gingerol were the Q-mar-kers of Zingiberis Rhizoma Carbonisata decoction pieces, and 6-gingerol, 8-gingerol, and 10-gingerol were Q-markers of Zingiberis Rhizoma decoction pieces. The result is expected to provide a reference for the establishment of quality standards for Zingiberis Rhizoma Carbonisata decoction pieces and Zingiberis Rhizoma decoction pieces.

Influence of 6-shogaol potentiated on 5-fluorouracil treatment of liver cancer by promoting apoptosis and cell cycle arrest by regulating AKT/mTOR/MRP1 signalling / 中国天然药物

Currently, chemoresistance seriously attenuates the curative outcome of liver cancer. The purpose of our work was to investigate the influence of 6-shogaol on the inhibition of 5-fluorouracil (5-FU) in liver cancer. The cell viability of cancer cells was determined by MTT assay. Liver cancer cell apoptosis and the cell cycle were examined utilizing flow cytometry. Moreover, qRT-PCR and western blotting was used to analyse the mRNA and protein expression levels, respectively. Immunohistochemistry assays were used to examine multidrug resistance protein 1 (MRP1) expression in tumour tissues. In liver cancer cells, we found that 6-shogaol-5-FU combination treatment inhibited cell viability, facilitated G0/G1 cell cycle arrest, and accelerated apoptosis compared with 6-shogaol or 5-FU treatment alone. In cancer cells cotreated with 6-shogaol and 5-FU, AKT/mTOR pathway- and cell cycle-related protein expression levels were inhibited, and MRP1 expression was downregulated. AKT activation or MRP1 increase reversed the influence of combination treatment on liver cancer cell viability, apoptosis and cell cycle arrest. The inhibition of AKT activation to the anticancer effect of 6-shogaol-5-FU could be reversed by MRP1 silencing. Moreover, our results showed that 6-shogaol-5-FU combination treatment notably inhibited tumour growth in vivo. In summary, our data demonstrated that 6-shogaol contributed to the curative outcome of 5-FU in liver cancer by inhibiting the AKT/mTOR/MRP1 signalling pathway.

Anticancer effects of 6-shogaol via the AKT signaling pathway in oral squamous cell carcinoma

Abstract Objective Oral squamous cell carcinoma (OSCC) is one of the common type of cancer that leads to death; and is becoming a global concern. Due to the lack of efficient chemotherapeutic agents for patients with oral cancer, the prognosis remains poor. 6-shogaol, a bioactive compound of ginger, has a broad spectrum of bioactivities and has been widely used to relieve many diseases. However, its effects on human oral cancer have not yet been fully evaluated. In our study, we investigated the anticancer effects of 6-shogaol on the proliferation, migration, invasion, apoptosis, and underlying mechanisms within human OSCC cell lines. Methodology We investigated the effect of 6-shogaol on the growth of OSCC cells by cell viability and soft agar colony formation assay. Migration and invasion assays were conducted to confirm the effect 6-shogaol on OSCC cell metastasis. Apoptosis was detected by flow cytometry and the underlying mechanism on the antigrowth effect of 6-shogaol in OSCC cells was assessed using western blotting. Results In our results, 6-shogaol not only suppressed proliferation and anchorage-independent cell growth in OSCC cells, but also induced apoptosis by regulating the apoptosis-associated factors such as p53, Bax, Bcl-2, and cleaved caspase-3. Migration and invasion of OSCC cells were inhibited following the regulation of E-cadherin and N-cadherin by 6-shogaol. Additionally, 6-shogaol treatment significantly inhibited the PI3K/AKT signaling pathway. Conclusion Therefore, our results may provide critical evidence that 6-shogaol can be a potential new therapeutic candidate for oral cancer.

Validation Of Reversed-Phase High-Performance Liquid Chromatography Method For Simultaneous Determination Of 6-, 8-, 10-Gingerols And 6-Shogaol From Ginger Extracts

Objective: Development and validation of a High-Performance Liquid Chromatography (HPLC) method for the simultaneous estimation of 6-, 8-, 10-Gingerols and 6-Shogaol in ginger extract using authentic standards. Methods: The chromatographic separation was achieved by using a C18 column and a mobile phase composed of acetonitrile, ortho-phospohoric acid in water and methanol. The proposed method was validated in terms of the analytical parameters such as specificity, accuracy, precision, linearity, range, the limit of detection (LOD) and limit of quantification (LOQ) according to ICH guidelines. Results: Linear calibration curves were obtained over concentration ranges of 10-250 µg/ml for 6-, 8-, 10-gingerols and 6-shogaol with determination coefficients more than 0.99 for each analyte. Intra and inter-day precisions of the method were found to be below 2% for each analyte, with relative standard deviation (% RSD) values in the range of 0.47 to 1.55% for 6-gingerol, 0.44 to 1.51% for 8-gingerol, 0.24 to 1.90% for 10-gingerol and 0.25 to 1.67% for 6-shogaol. The percentage recovery of gingerols and shogaol was obtained with an average of 99.53%, 99.97%, 100.13% and 100.53% respectively, which was well within acceptance range. Conclusion: Simple, accurate, precise and rapid HPLC method was developed for the simultaneous analysis of 6-, 8-, 10-gingerols and 6-shogaol and validated in accordance with ICH guidelines. The developed method was found to be suitable for the standardization of herbal extracts and polyherbal formulations for the content of 6-, 8-, 10-gingerols and 6-shogaol.

Study on the Action Mechanism of 6-Shogaol in the Treatment of Ulcerative Colitis from the Signaling Pathway of Notch and TLR4/ NF-κB / 中国药学杂志

OBJECTIVE: To explore the effect and mechanism of 6-gingerenol on ulcerative colitis based on TLR4/NF-κB signaling pathway related to immune inflammation and Notch signaling pathway related to mucosal repair. METHODS: Forty Kunming mice were randomly divided into the control group (n=10), the model group (n=10), the sulfasalazine group (n=10) and the 6-gingerenol group (n=10). Ulcerative colitis model mice were induced by oral administration of 2% dextran sulfate sodium(DSS) except those in the control group. The mice in the sulfasalazine group and the 6-gingerenol group were given sulfasalazine and 6-gingerenol (ig 100 mg•kg-1) respectively, and mice in the other groups were given normal saline. The serum and colon were isolated after 20-days treatment. The expressions of Hes-1, Math-1, MUC2 and olfm4 proteins in colonic tissues were detected by immunofluorescence. The expression levels of notch-1, Hes-1, Math-1, TLR4, NF-κBp65 mRNA and proteins in colonic tissues were detected by RT-PCR and Western blot. The levels of TNF-α and IL-1β in serum were detected by ELISA. RESULTS: Compared with the model group, the expressions of Notch-1, Hes-1, TLR4 and NF-κB p65 proteins and the relative expressions of mRNA in colon epithelial tissue of mice in sulfasalazine group and 6-gingerenol group were significantly decreased, while the relative expression of mRNA in Math-1 protein was significantly increased, and the contents of TNF-α and IL-1β in serum were significantly decreased. The expression of olfm4 protein was significantly decreased and the expression of MUC2 protein was significantly increased in 6-gingerenol group, but there was no significant change in olfm4 and MUC2 proteins expressions in sulfasalazine group. Compared with sulfasalazine group, the expressions of Notch-1 and Hes-1 proteins and the relative expression of mRNA in colon epithelial tissue in the 6-gingerenol group were significantly decreased, the relative expression of mRNA and protein of Math-1 were significantly increased, the expression of olfm4 protein was significantly decreased, and the expression of MUC2 protein was significantly increased, but the expressions of TLR4 and NF-κBp65 proteins and mRNA, and the TNF-α and IL-1β in serum were not significantly changed in 6-gingerenol group. CONCLUSION: 6-Gingerenol could treat ulcerative colitis though inhibiting intestinal immune inflammation and promoting the repair of damaged colonic mucosa. The mechanism is related to its inhibition on activation of Notch and TLR4/NF-κB signaling pathway.

HPLC-ELSD fingerprint of Zhenwu Decoction and simultaneous determination of 13 components / 中草药

Objective: To establish HPLC-ELSD fingerprint of Zhenwu Decoction(ZWD), screen out the signature components of ZWD through chemical pattern recognition, so as to establish the content determination method of ZWD based on this index. Methods: The fingerprint of 16 batches of ZWD was established by HPLC-ELSD method. The similarity evaluation system of traditional Chinese medicine chromatographic fingerprint (2012 Version) was used for similarity evaluation to determine the common peaks and its attribution. Cluster analysis (CA), principal component analysis (PCA) and orthogonal partial least squares discriminant analysis (OPLS-DA) were used to select the index components of ZWD. Results: The fingerprint of ZWD was established, 38 common peaks were confirmed, and the similarity was > 0.95. The results of CA, PCA and OPLS-DA were consistent and the samples were divided into three categories. Benzoylmesaconine, benzoylaconitine, benzoylhypacoitine, polyporenic acid C, pachymic acid, atractylenolide II, atractylenolide III, oxypaeoniflorin, albiflorin, paeoniflorin and benzoylpaeoniflorin were identified as the 11 index components with significant difference contribution in different batches of ZWD samples. 6-Gingerol and 6-shogaol were the main active components of ginger, so the above 13 components were taken as the index components of ZWD. The chromatographic peak separation degree and linear relationship were good. The average recovery rate was 96.46%-99.80%, RSD ≤ 3.15%. The mass fraction range of benzoylmesaconine, benzoylaconitine, benzoylhypacoitine, polyporenic acid C, pachymic acid, atractylenolide II, atractylenolide III, oxypaeoniflorin, albiflorin, paeoniflorin, benzoylpaeoniflorin, 6-gingerol, 6-shogaol in 16 batches were 283.93-576.86, 25.05-147.39, 62.96-303.37, 31.24-131.27, 9.76-44.04, 32.15-83.55, 76.55-333.13, 17.48-146.61, 456.58-1554.14, 3 322.48-5 590.01, 158.21-556.50, 525.85-582.92 and 68.52-74.73 mg/g, respectively. Conclusion: The fingerprint combined with PCA, CA and OPLS-DA can comprehensively evaluate the quality of ZWD. This method is stable and reliable, providing reference for the quality evaluation.

Effect of Coptidis Rhizoma-Zingiberis Rhizoma Extract on Colonic Epithelium TLR4/NF-κB Signaling Pathway in Mice with Ulcerative Colitis / 中国实验方剂学杂志

Objective::To observe the effect of berberine and 6-shogaol, main components of Coptiae Rhizoma and Zingiberis Rhizoma, on the inflammatory signaling pathway of Toll-like receptors 4 (TLR4)/nuclear factor kappa B (NF-κB) in colonic epithelial cells of mice with ulcerative colitis. Method::Fifty Kunming mice were randomly divided into normal group, model group, berberine group (100 mg·kg-1), 6-shogaol group (100 mg·kg-1), and 6-shogaol combined with berberine group (200 mg·kg-1), with 10 mice in each group. A mouse model of ulcerative colitis was established through oral administration with 2% dextroan sulfate for two weeks. Each group was given corresponding drugs by gavage, while normal group and model group were given equal amount of normal saline. Serum and colon tissue samples were taken 20 days after administration. Enzyme-linked immunosorbent method was used to detect serum interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α) expressions, and real-time fluorescence quantitative polymerase chain reaction(Real-time PCR) and Western blot method were used to detect TLR4, NF-κB p65 mRNA and protein expressions in colon epithelial tissue. Result::Compared with the normal group, relative expressions of TLR4 and NF-κB p65 mRNA and protein were increased in the model group (P<0.01), and the contents of serum IL-1β and TNF-α were increased (P<0.01). Compared with the model group, relative expressions of TLR4 and NF-κB p65 mRNA and protein were significantly decreased in 6-shogaol group, berberine group and 6-shogaol combined with berberine group (P<0.01), and the contents of serum IL-1β and TNF-α were significantly decreased (P<0.01). Among the three groups, 6-shogaol combined with berberine group had the strongest effect (P<0.01). Conclusion::Both 6-shogaol and berberine can inhibit colonic inflammation, reduce inflammatory damage and treat ulcerative colitis. The combined application of 6-shogaol and berberine has a significant synergism effect. The mechanism is related to the excessive activation of TLR4/NF-κB pathway and the regulation of non-controllable intestinal inflammation.

Effect of berberine combined with 6-shogaol on notch signaling pathway in colon epithelial cells of ulcerative colitis mice / 中草药

Objective: To investigate the effects of berberine and 6-shogaol on Notch signaling pathway in colonic epithelial cells of mice with ulcerative colitis. Methods: A total of 50 Kunming mice were randomly selected as normal group, and the remaining 40 mice were treated with po 2% DSS to induce ulcerative colitis. The rats were randomly divided into model group, berberine group, 6-shogaol group, and berberine + 6-shogaol group. The corresponding drugs were given by ig administration. Colon tissues were taken after 20 d of administration. The protein expression of Hes-1, Math-1, MUC2, and olfm4 in colon epithelium was detected by immunofluorescence method, and the gene and protein expression of Notch-1, Hes-1, Math-1 in colon epithelium were detected by qRT-PCR and Western blotting. Results: Compared with the model group, the protein and mRNA expression of Notch-1, Hes-1, and olfm4 in colon epithelium of mice in 6-shogaol group, berberine group, and berberine + 6-shogaol group were significantly decreased, while the expression of Math-1 and MUC2 were significantly increased. Among three groups, berberine + 6-shogaol group showed the best effect. Conclusion: Both 6-shogaol and berberine can effectively repair damaged colonic mucosa and treat ulcerative colitis. The combined use of berberine and 6-shogaol is more effective than the single application. Its mechanism is related to inhibiting the over-activation of Notch pathway and regulating the proliferation and differentiation of colonic epithelium.

6-Shogaol reduces progression of experimental endometriosis in vivo and in vitro via regulation of VGEF and inhibition of COX-2 and PGE2-mediated inflammatory responses

Endometriosis (EM) is one of the most common gynaecological disorder affecting women in their reproductive age. Mechanisms involved in the pathogenesis of EM remains poorly understood, however inflammatory responses have been reported to be significantly involved. The efficacy of 6-shogaol on proliferation of endometriotic lesions and inflammatory pathways in experimentally-induced EM model was explored in this study. EM was stimulated in Sprague-Dawley rats by implantation of autologous endometrium onto the peritoneum abdominal wall. Separate groups were treated with 6-shogaol (50, 100 or 150 mg/kg b.wt/day) via oral gavage for one month period. Gestrinone (GTN) group received GTN (0.5 mg/kg/day) as positive control. Five weeks after implantation, the spherical volume of ecto-uterine tissues was determined. Treatment with 6-shogaol significantly reduced the implant size. Histological analysis reported atrophy and regression of the lesions. 6-shogaol administration effectively down-regulated NF-κB signaling, VEGF and VEGFR-2 (Flk-1) expression in the endometriotic lesions. Excess production of IL-1β and IL-6 (pro-inflammatory cytokines), PGE2 and nitric oxide (NO) were reduced. Overall, the results of the study reveal the efficacy of 6-shogaol against endometriosis via effectively suppressing proliferation of the lesions and modulating angiogenesis and COX-2/NF-κB-mediated inflammatory cascades.

Drying processing method for Zingiberis Rhizoma based on multiple bioactive constituents / 中草药

Objective To compare the effects of different drying methods on six bioactive constituents of Zingiberis Rhizoma (ZR), and explore the dynamic changes of bioactive constituents and water content during the drying process. Methods The multiple components in ZR were simultaneously measured by HPLC, and 6-gingerol, 8-gingerol, 10-gingerol, 6-shogaol, α-curcumene, (E)- β-farnesene were used as indexes to evaluate ZR obtained from different drying methods. The Weibull function was used to simulate the dynamic change of water content, which was combined with the dynamic changes of components during the drying process of ZR to explore the principle of drying process. Results A total of 12 kinds of drying methods had a certain effect on the multiple components of ZR, and the components presented the fluctuation change in the drying process. The coefficient of correlation of Weibull functional simulation of ZR drying process was greater than 0.990. Conclusion ZR obtained by drying at 60 ℃ was better. Water content range of 6%-15% was suitable for processing ZR. 6-gingerol, 8-gingerol, 10-gingerol, and 6-shogaol were significantly negatively correlated with the moisture content of ZR. The Weibull distribution model could well simulate the fluctuation change of water content in the drying process, and it was of great significance for the prediction and quality control of ZR during drying process, which could also provide a technical basis for the use of modern drying technology to dry ZR at the same time.

Research Progress in Zingiber officinale / 中国药学杂志

Zingiber officinale has a long application history in China, it is used as medicine, also condiment, food and drinks. The chemical constituents of Zingiber officinale include volatile oil, gingerol, diaryl-heptnaoids and so on. Scientific research showed that Zingiber officinale is widely used in anti-nausea, resisting gastric ulcer, anti-bacterial, anti-phlogistic, analgesia, antioxidant, resisting motion sickness, anti-tumor, hypoglycemic, antilipidemic, and improving cardiocerebral vascular system and so on. Zingiber officinale is recommended as a healthy food by doctor of traditional Chinese medicine in all times. In summary, it's worth to be researched and developed in detail.

Characteristics and thermodynamics of the interaction of 6-shogaol with human serum albumin as studied by isothermal titration calorimetry

ABSTRACT The interaction between 6-shogaol, a pharmacologically active ginger constituent, and human serum albumin (HSA), the main in vivo drug transporter, was investigated using isothermal titration calorimetry (ITC). The value of the binding constant, Ka (5.02 ± 1.37 × 104 M−1) obtained for the 6-shogaol-HSA system suggested intermediate affinity. Analysis of the ITC data revealed feasibility of the binding reaction due to favorable enthalpy and entropy changes. The values of the thermodynamic parameters suggested involvement of van der Waals forces, hydrogen bonds and hydrophobic interactions in the 6-shogaol-HSA complex formation.

Correlation research between in vivo tissue distribution in rats and channel tropism of active constituents in dried ginger / 中草药

Objective: To observe the in vivo tissue distribution of 6-gingerols, 6-shogaol, and 8-gingerols from dried ginger (the rhizome of Zingiber officinale) in rats and to discuss the channel tropism of them. Methods: The method of "symptoms-efficacy-pharmacokinetics" was used and the ginger solution was ig given to the rats which were deficiency-cold in spleen and stomach; Then the blood, heart, liver, spleen, lung, kidney, brain, stomach, large intestine, and small intestine were immediately taken out after the rats were ig given the medince in 10, 20, 40, 60, 80, 120, and 360 min, respectively; Finally, HPLC was used to detect the concentration of 6-gingerols, 6-shogaol, and 8-gingerols from dried ginger in different tissues of rats in each group of deficiency-cold in spleen and stomach and normal by calculating the pharmacokinetic parameters. Results: We found that in the group of deficiency-cold in spleen and stomach, the concentration of the three active components was the highest in stomach, small intestine, liver, and lung, and in the group of normal, the three components were mostly distributed in the stomach, kidney, small intestine, large intestine, and lung. What's more, in the digestive organs, such as stomach and small intestine, the concentration was obviously higher in the group of deficiency-cold in spleen and stomach than that in the group of normal. Conclusion: The main ingredients of dried ginger mostly distribute in spleen, stomach, lung, and liver. This view conform to the traditional channel trpism of dried ginger in traditional Chinese medicine theory.

Ginger and Its Pungent Constituents Non-Competitively Inhibit Serotonin Currents on Visceral Afferent Neurons

Nausea and emesis are a major side effect and obstacle for chemotherapy in cancer patients. Employ of antiemetic drugs help to suppress chemotherapy-induced emesis in some patients but not all patients. Ginger, an herbal medicine, has been traditionally used to treat various kinds of diseases including gastrointestinal symptoms. Ginger is effective in alleviating nausea and emesis, particularly, for cytotoxic chemotherapy drug-induced emesis. Ginger-mediated antiemetic effect has been attributed to its pungent constituents-mediated inhibition of serotonin (5-HT) receptor activity but its cellular mechanism of action is still unclear. Emetogenic chemotherapy drugs increase 5-HT concentration and activate visceral vagal afferent nerve activity. Thus, 5-HT mediated vagal afferent activation is essential to provoke emesis during chemotherapy. In this experiment, water extract of ginger and its three major pungent constituent's effect on 5-HT-evoked responses were tested on acutely dispersed visceral afferent neurons with patch-clamp methods. The ginger extract has similar effects to antiemetic drug ondansetron by blocking 5-HT-evoked responses. Pungent constituents of the ginger, [6]-shogaol, [6]-gingerol, and zingerone inhibited 5-HT responses in a dose dependent manner. The order of inhibitory potency for these compounds were [6]-shogaol>[6]-gingerol>zingerone. Unlike well-known competitive 5-HT3 receptor antagonist ondansetron, all tested ginger constituents acted as non-competitive antagonist. Our results imply that ginger and its pungent constituents exert antiemetic effects by blocking 5-HT-induced emetic signal transmission in vagal afferent neurons.

Evaluation of ginger (Zingiber officinale Roscoe) bioactive compounds in increasing the ratio of T-cell surface molecules of CD3+CD4+:CD3+CD8+ in-vitro

The potential ability of ginger bioactive compounds in increasing the ratio of T-cell surface molecules of CD3+CD4+:CD3+CD8+ was investigated using dual tagging FITC and PE of monoclonal antibody anti-human with its fluorescence measured by flow cytometer. Oleoresin was extracted using sinkhole distillation technique. Its components namely, gingerol in fraction-1, shogaol in fraction 2 and zingeron in fraction-3 were separated by column vacuum chromatography method. The doses of oleoresin, gingerol, shogaol, and zingeron tested were 50, 100,150, 200, and 250 μg/ml. Lymphocytes (2x106 cell/ml) from human peripheral blood were isolated using ficoll density gradient technique, and cultured in the presence of the compounds in RPMI-1640 medium and phytohemaglutinin (PHA) mitogen for 96 h under normal conditions. Percentages of T-cell surface molecules (CD4+ and CD8+) were determined using dual-tagging FITC and PE fluorescents labeled on monoclonal antibody anti human. The fluorescence-labeled bands on the T-cell surface molecules were counted using flow cytometer. The experiment revealed that oleoresin and its three fractions increased the percentage of CD3+CD4+. The compound in fraction 3 of oleoresin at 200 μg/ml increased by the highest percentage of CD3+CD4+ of 9%, but slightly decreased the percentage of CD3+CD8+. These ginger bioactive compounds increased the ratio of CD3+CD4:CD3+CD8+ T-cells with the highest increment of 30% from effects of 200 μg/ml fraction 3 of oleoresin. This in vitro finding revealed that ginger bioactive compounds potentially increased cellular and humoral immune response. Further clinical studies are needed to confirm the benefits of these ginger bioactive compounds as a potential functional food for testing on HIV infected patients.