Screening for the extracorporeal coagulation activity quality markers (Q-markers) of dried and stir fried ginger

Ginger is a common condiment that is widely used as Chinese medicine in China and Southeast Asia. Dried ginger and stir fried ginger are two common processed products of ginger, with distinct clinical uses. The aim of this study was to identify the chemical components (quality markers, Q-mark) responsible for the differences in in vitro hemostatic activity between dried and stir fried ginger and to provide a basis for the selection between the two types of ginger in clinical application. In this study, methanolic extracts of dried and stir fried ginger were characterized using UPLC-Q/TOF-MS and then evaluated for in vitro coagulation activity. Spectral effect correlation analysis was used to identify quality markers, while molecular docking simulation was used to evaluate the binding energy between potential active compounds and target proteins. A total of 49 chemical constituents of the ginger extracts were identified using UPLC-Q/TOF-MS, 27 of which were significantly different between the two extracts. A fingerprint of 18 batches of dried and stir fried ginger established that zingiberone, 6-gingerol, 8-gingerol, 6-shogaol, 10-gingerol, 8-shogaol, and 10-shogaol were common constituents of the two extracts. Results of coagulation assays revealed that dried ginger had anti-coagulation effects, while stir fried ginger had hemostatic effect. Zingiberone, 6-shogaol and 10-shogaol were identified as the active components responsible for the hemostatic effect of Stir fried ginger through multivariate statistical analysis. In addition, molecular docking simulations suggested that these three components bound to Src proteins on platelets. Consequently, 6-gingerol, zingiberone, 6-shogaol and 10-shogaol were selected as quality markers to distinguish between dried and stir fried ginger. These results provide scientific evidence for the establishment of a quality evaluation system for the integrity and specificity of dried and stir fried ginger.     

Liquid chromatographic determination of 6-, 8-, 10-gingerol, and 6-shogaol in ginger (Zingiber officinale) as the raw herb and dried aqueous extract

The determination of 6-, 8-, 10-gingerol, and 6-shogaol in dried ginger (Zingiber officinale) and in the dried aqueous extract of ginger is reported. This is the first study to report a validated method for the determination of these 4 analytes. Several extraction solvents and methods were examined, and the optimum combination was determined. The samples were extracted at room temperature by sonication with methanol, and the extract was analyzed by liquid chromatography with photodiode array detection. A C18 column was used with a water-acetonitrile gradient mobile phase. Quantification was at 200 nm. The levels of 6-, 8-, 10-gingerol, and 6-shogaol in the raw herb were 9.3, 1.6, 2.3, and 2.3 mglg, respectively. The levels of gingerols found in the dried aqueous extract were between 5 and 16 times lower than those in the raw herb, but the level of 6-shogaol was higher. Analyte identity was confirmed by negative-ion electrospray ionization tandem mass spectrometry, in which 2 daughter ions were obtained for each analyte. The average recovery was 97% with a relative standard deviation of <8%. The limits of detection for 6-, 8-, 10-gingerol, and 6-shogaol in the raw herb were 0.22, 0.04, 0.09, and 0.07 mglg, respectively, and in the dried aqueous extract, 0.11, 0.02, 0.02, and 0.14 mg/g, respectively.     

Cytoprotection against Oxidative Stress by Methylnissolin-3-O-β-d-glucopyranoside from Astragalus membranaceus Mainly via the Activation of the Nrf2/HO-1 Pathway

Astragalus membranaceus is a famous herb found among medicinal and food plants in East and Southeastern Asia. The Nrf2-ARE assay-guided separation of an extract from Jing liqueur led to the identification of a nontoxic Nrf2 activator, methylnissolin-3-O-β-d-glucopyranoside (MNG, a component of A. membranaceus). Nrf2 activation by MNG has not been reported before. Using Western Blot, RT-qPCR and imaging, we investigated the cytoprotective effect of MNG against hydrogen peroxide-induced oxidative stress. MNG induced the expression of Nrf2, HO-1 and NQO1, accelerated the translocation of Nrf2 into nuclei, and enhanced the phosphorylation of AKT. The MNG-induced expression of Nrf2, HO-1, and NQO1 were abolished by Nrf2 siRNA, while the MNG-induced expression of Nrf2 and HO-1 was abated and the AKT phosphorylation was blocked by LY294002 (a PI3K inhibitor). MNG reduced intracellular ROS generation. However, the protection of MNG against the H₂O₂ insult was reversed by Nrf2 siRNA with decreased cell viability. The enhancement of Nrf2 and HO-1 by MNG upon H₂O₂ injury was reduced by LY294002. These data showed that MNG protected EA.hy926 cells against oxidative damage through the Nrf2/HO-1 and at least partially the PI3K/Akt pathways.     

Protective Effects of 6-Shogaol,an Active Compound of Ginger,in a Murine Model of Cisplatin-Induced Acute Kidney Injury

Acute kidney injury (AKI) is a dose-limiting side effect of cisplatin therapy in cancer patients. However, effective therapies for cisplatin-induced AKI are not available. Oxidative stress, tubular cell death, and inflammation are known to be the major pathological processes of the disease. 6-Shogaol is a major component of ginger and exhibits anti-oxidative and anti-inflammatory effects. Accumulating evidence suggest that 6-shogaol may serve as a potential therapeutic agent for various inflammatory diseases. However, whether 6-shogaol exerts a protective effect on cisplatin-induced renal side effect has not yet been determined. The aim of this study was to evaluate the effect of 6-shogaol on cisplatin-induced AKI and to investigate its underlying mechanisms. An administration of 6-shogaol after cisplatin treatment ameliorated renal dysfunction and tubular injury, as shown by a reduction in serum levels of creatinine and blood urea nitrogen and an improvement in histological abnormalities. Mechanistically, 6-shogaol attenuated cisplatin-induced oxidative stress and modulated the renal expression of prooxidant and antioxidant enzymes. Apoptosis and necroptosis induced by cisplatin were also suppressed by 6-shogaol. Moreover, 6-shogaol inhibited cisplatin-induced cytokine production and immune cell infiltration. These results suggest that 6-shogaol exhibits therapeutic effects against cisplatin-induced AKI via the suppression of oxidative stress, tubular cell death, and inflammation.     

Coffee-Derived Phenolic Compounds Activate Nrf2 Antioxidant Pathway in I/R Injury In Vitro Model: A Nutritional Approach Preventing Age Related-Damages

Age-related injuries are often connected to alterations in redox homeostasis. The imbalance between free radical oxygen species and endogenous antioxidants defenses could be associated with a growing risk of transient ischemic attack and stroke. In this context, a daily supply of dietary antioxidants could counteract oxidative stress occurring during ischemia/reperfusion injury (I/R), preventing brain damage. Here we investigated the potential antioxidant properties of coffee-derived circulating metabolites and a coffee pulp phytoextract, testing their efficacy as ROS scavengers in an in vitro model of ischemia. Indeed, the coffee fruit is an important source of phenolic compounds, such as chlorogenic acids, present both in the brewed seed and in the discarded pulp. Therefore, rat brain endothelial cells, subjected to oxygen and glucose deprivation (OGD) and recovery (ogR) to mimic reperfusion, were pretreated or not with coffee by-products. The results indicate that, under OGD/ogR, the ROS accumulation was reduced by coffee by-product. Additionally, the coffee extract activated the Nrf2 antioxidant pathway via Erk and Akt kinases phosphorylation, as shown by increased Nrf2 and HO-1 protein levels. The data indicate that the daily intake of coffee by-products as a dietary food supplement represents a potential nutritional strategy to counteract aging.     

Gastrointestinal motility enhancing effect of ginger and its active constituents

The effect of ginger root (Zingiberis Rhizoma) on gastrointestinal motility was examined based on its ability to enhance charcoal meal transport in mice. Oral administrations of the acetone extract of ginger (which contains volatile oils and bitter substances) at 75 mg/kg, [6]-shogaol at 2.5 mg/kg, or a [6]-, [8]- or [10]-gingerol at 5 mg/kg enhanced the transport of a charcoal meal. The effects of these substances were similar to or slightly weaker than those of metoclopramide and donperidone.     

A Novel Synthetic Precursor of Styryl Sulfone Neuroprotective Agents Inhibits Neuroinflammatory Responses and Oxidative Stress Damage through the P38 Signaling Pathway in the Cell and Animal Model of Parkinson’s Disease

A novel class of styryl sulfones were designed and synthesized as CAPE derivatives by our work team, which showed a multi-target neuroprotective effect, including antioxidative and anti-neuroinflammatory properties. However, the underlying mechanisms remain unclear. In the present study, the anti-Parkinson’s disease (PD) activity of 10 novel styryl sulfone compounds was screened by the cell viability test and the NO inhibition test in vitro. It was found that 4d exhibited the highest activity against PD among them. In a MPTP-induced mouse model of PD, the biological activity of 4d was validated through suppressing dopamine neurotoxicity, microglial activation, and astrocytes activation. With compound 4d, we conducted the mechanistic studies about anti-inflammatory responses through inhibition of p38 phosphorylation to protect dopaminergic neurons, and antioxidant effects through promoting nuclear factor erythroid 2-related factor 2 (Nrf2). The results revealed that 4d could significantly inhibit 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/1-methyl-4-phenylpyridinium (MPTP/MPP⁺)-induced p38 mitogen-activated protein kinase (MAPK) activation in both in vitro and in vivo PD models, thus inhibiting the NF-κB-mediated neuroinflammation-related apoptosis pathway. Simultaneously, it could promote Nrf2 nuclear transfer, and upregulate the expression of antioxidant phase II detoxification enzymes HO-1 and GCLC, and then reduce oxidative damage.     

6,7,4′-Trihydroxyflavanone Mitigates Methamphetamine-Induced Neurotoxicity in SH-SY5y Cells via Nrf2/heme Oxyganase-1 and PI3K/Akt/mTOR Signaling Pathways

Methamphetamine (METH) is a synthetic psychostimulant drug that has detrimental effects on the health of its users. Although it has been investigated as a cause of neurodegenerative disease due to its neurotoxicity, whether small molecules derived from natural products attenuate these side effects remains elusive. 6,7,4′-trihydroxyflavanone (THF) is a flavanone family that possesses various pharmacological activities, including anti-rheumatic, anti-ischemic, anti-inflammatory, anti-osteoclastogenic, and protective effects against METH-induced deactivation of T cells. However, little is known about whether THF protects neuronal cells from METH-induced neurotoxicity. Here, we investigated the protective effects of THF on neurotoxicity induced by METH exposure by enhancing the Nrf2/HO-1 and PI3K/Akt/mTOR signaling pathways in SH-SY5y cells. Treatment with THF did not lead to cytotoxicity, but attenuated METH-induced neurotoxicity by modulating the expression of apoptosis-related proteins, METH-induced oxidative stress, and PI3K/Akt/mTOR phosphorylation in METH-exposed SH-SY5y cells. Moreover, we found THF induced Nrf2 nuclear translocation and HO-1 expression. An inhibitor assay confirmed that the induction of HO-1 by THF attenuates METH-induced neurotoxicity. Therefore, we suggest that THF preserves neuronal cells from METH-induced neurotoxicity by upregulating HO-1 expression through the Nrf2 and PI3K/Akt/mTOR signaling pathways. Thus, THF has therapeutic potential for use in the treatment of METH-addicts suffering from neurodegenerative diseases.     

Identification of a Sesquiterpene Lactone from Arctium lappa Leaves with Antioxidant Activity in Primary Human Muscle Cells

Many pathologies affecting muscles (muscular dystrophies, sarcopenia, cachexia, renal insufficiency, obesity, diabetes type 2, etc.) are now clearly linked to mechanisms involving oxidative stress. In this context, there is a growing interest in exploring plants to find new natural antioxidants to prevent the appearance and the development of these muscle disorders. In this study, we investigated the antioxidant properties of Arctium lappa leaves in a model of primary human muscle cells exposed to H₂O₂ oxidative stress. We identified using bioassay-guided purification, onopordopicrin, a sesquiterpene lactone as the main molecule responsible for the antioxidant activity of A. lappa leaf extract. According to our findings, onopordopicrin inhibited the H₂O₂-mediated loss of muscle cell viability, by limiting the production of free radicals and abolishing DNA cellular damages. Moreover, we showed that onopordopicrin promoted the expression of the nuclear factor-erythroid-2-related factor 2 (Nrf2) downstream target protein heme oxygenase-1 (HO-1) in muscle cells. By using siRNA, we demonstrated that the inhibition of the expression of Nrf2 reduced the protective effect of onopordopicrin, indicating that the activation of the Nrf2/HO-1 signaling pathway mediates the antioxidant effect of onopordopicrin in primary human muscle cells. Therefore, our results suggest that onopordopicrin may be a potential therapeutic molecule to fight against oxidative stress in pathological specific muscle disorders.     

Synergistic Anti-Inflammatory Activity of Ginger and Turmeric Extracts in Inhibiting Lipopolysaccharide and Interferon-γ-Induced Proinflammatory Mediators

This study aims to investigate the combined anti-inflammatory activity of ginger and turmeric extracts. By comparing the activities of individual and combined extracts in lipopolysaccharide and interferon-γ-induced murine RAW 264.7 cells, we demonstrated that ginger-turmeric combination was optimal at a specific ratio (5:2, w/w) in inhibiting nitric oxide, tumour necrosis factor and interleukin 6 with synergistic interaction (combination index < 1). The synergistic inhibitory effect on TNF was confirmed in human monocyte THP-1 cells. Ginger-turmeric combination (5:2, w/w) also upregulated nuclear factor erythroid 2–related factor 2 activity and heme oxygenase-1 protein expression. Additionally, 6-shogaol, 8-shogaol, 10-shogaol and curcumin were the leading compounds in reducing major proinflammatory mediators and cytokines, and a simplified compound combination of 6-s, 10-s and curcumin showed the greatest potency in reducing LPS-induced NO production. Our study provides scientific evidence in support of the combined use of ginger and turmeric to alleviate inflammatory processes.     

超高效液相色谱-串联质谱法快速同时测定生姜中10种营养成分

建立了超高效液相色谱-串联质谱（UHPLC-MS/MS）快速同时测定生姜中姜辣素类和姜黄素类营养成分的分析方法，具体包括6-姜酚、8-姜酚、10-姜酚、6-姜烯酚、8-姜烯酚、10-姜烯酚、四氢姜黄素、姜黄素、去甲氧基姜黄素、双去甲氧基姜黄素10种目标物。采用ZORBAX RRHD Eclipse Plus C18色谱柱（100 mm×2.1 mm，1.8 μm）分离，以0.1%（v/v）甲酸水溶液和0.1%（v/v）甲酸甲醇溶液为流动相进行梯度洗脱，采用电喷雾电离（ESI）源、正离子和多反应监测（MRM）模式对目标物进行定性确证和定量分析。10种营养成分的线性相关系数（r）均≥ 0.9995，方法的定量限为0.10~7.71 μg/L，样品基质在3个水平下的平均加标回收率为82.8%~115.3%，相对标准偏差（RSD）为0.58%~11.49%。分析结果显示，生姜中10种营养成分均有检出，其中6-姜酚的含量最高且集中分布于373.35~702.48 mg/kg。该法简便快速，准确可靠，适用于生姜中姜辣素类和姜黄素类营养成分的分析，可为生姜质量鉴定和控制提供技术手段。     

Preparation of the monomers of gingerols and 6-shogaol by flash high speed counter-current chromatography

The flash high speed counter-current chromatographic (FHSCCC) separation of gingerols and 6-shogaol was performed on a HSCCC instrument equipped with a 1200-ml column (5 mm tubing i.d.) at a flow rate of 25 ml/min. The performance met the FHSCCC feature that the flow rate of mobile phase (ml) is equal to or greater than the square of the diameter of the column tubing (mm). The separation employed the upper phase of stationary phase of the n-hexane-ethyl acetate-methanol-water (3:2:2:3, v/v) as the stationary phase. A stepwise elution was performed by eluting with the lower phase of n-hexane-ethyl acetate-methanol-water (3:2:2:3, v/v) for first 90 min and the lower phase of the n-hexane-ethyl acetate-methanol-water (3:2:6:5, v/v) for the second 90 min. In each separation 5 g of the ethyl acetate extract of rhizomes of ginger was loaded, yielding 1.96 g of 6-gingerol (98.3%), 0.33 g of 8-gingerol (97.8%), 0.64 g of 6-shogaol (98.8%) and 0.57 g of 10-gingerol (98.2%). The separation can be expected to scale up to industrial separation.     

Protective Effect of Djulis (Chenopodium formosanum) Extract against UV- and AGEs-Induced Skin Aging via Alleviating Oxidative Stress and Collagen Degradation

Skin aging is a complex process involving photoaging and glycation stress, which share some fundamental pathways and have common mediators. They can cause skin damage and collagen degradation by inducing oxidative stress and the accumulation of reactive oxygen species (ROS). Chenopodium formosanum (CF), also known as Djulis, is a traditional cereal in Taiwan. This study investigated the protection mechanisms of CF extract against ultraviolet (UV) radiation and advanced glycation end products (AGEs)-induced stress. The results indicated that CF extract had strong antioxidant and free radical scavenging effects. It could reduce UV-induced intracellular ROS generation and initiate the antioxidant defense system by activating the nuclear factor erythroid 2-related factor 2 (Nrf2)/heme oxygenase-1 (HO-1) signaling pathway in human skin fibroblasts. CF extract modulated mitogen-activated protein kinase (MAPK) and transformed growth factor-beta (TGF-β) signaling pathways to alleviate oxidative stress-induced skin aging. Moreover, the results revealed that CF extract not only promoted collagen synthesis but also improved aging-induced collagen degradation. CF extract attenuated AGEs-induced ROS production and the upregulation of receptor for AGEs (RAGE). The overall results suggest that CF extract provides an effective anti-aging strategy by preventing skin damage from oxidative stress and collagen loss with potent antioxidant, anti-photoaging, and antiglycation activities.