Mitigating phospholipid peroxidation of macrophages in stress-induced tumor microenvironment by natural ALOX15/PEBP1 complex inhibitors.

BACKGROUND: The intricate interactions between chronic psychological stress and susceptibility to breast cancer have been recognized, yet the underlying mechanisms remain unexplored. Danzhi Xiaoyao Powder (DZXY), a traditional Chinese medicine (TCM) formula, has found clinical utility in the treatment of breast cancer. Macrophages, as the predominant immune cell population within the tumor microenvironment (TME), play a pivotal role in orchestrating tumor immunosurveillance. Emerging evidence suggests that lipid oxidation accumulation in TME macrophages, plays a critical role in breast cancer development and progression. However, a comprehensive understanding of the pharmacological mechanisms and active components of DZXY related to its clinical application in the treatment of stress-aggravated breast cancer remains elusive. PURPOSE: This study sought to explore the plausible regulatory mechanisms and identify the key active components of DZXY contributing to its therapeutic efficacy in the context of breast cancer. METHODS: Initially, we conducted an investigation into the relationship between the phagocytic capacity of macrophages damaged by psychological stress and phospholipid peroxidation using flow cytometry and LC-MS/MS-based phospholipomics. Subsequently, we evaluated the therapeutic efficacy of DZXY based on the results of the tumor size, tumor weight, the phospholipid peroxidation pathway and phagocytosis of macrophage. Additionally, the target-mediated characterization strategy based on binding of arachidonate 15-lipoxygenase (ALOX15) to phosphatidylethanolamine-binding protein-1 (PEBP1), including molecular docking analysis, microscale thermophoresis (MST) assay, co-immunoprecipitation analysis and activity verification, has been further implemented to reveal the key bio-active components in DZXY. Finally, we evaluated the therapeutic efficacy of isochlorogenic acid C (ICAC) based on the results of tumor size, tumor weight, the phospholipid peroxidation pathway, and macrophage phagocytosis in vivo. RESULTS: The present study demonstrated that phospholipid peroxides, as determined by LC-MS/MS-based phospholipidomics, triggered in macrophages, which in turn compromised their capacity to eliminate tumor cells through phagocytosis. Furthermore, we elucidate the mechanism behind stress-induced PEBP1 to form a complex with ALOX15, thereby mediating membrane phospholipid peroxidation in macrophages. DZXY, demonstrates potent anti-breast cancer therapeutic effects by disrupting the ALOX15/PEBP1 interaction and inhibiting phospholipid peroxidation, ultimately enhancing macrophages' phagocytic capability towards tumor cells. Notably, ICAC emerged as a promising active component in DZXY, which can inhibit the ALOX15/PEBP1 interaction, thereby mitigating phospholipid peroxidation in macrophages. CONCLUSION: Collectively, our findings elucidate stress increases the susceptibility of breast cancer by driving lipid peroxidation of macrophages and suggest the ALOX15/PEBP1 complex as a promising intervention target for DZXY.

An ethanolic extract of Arctium lappa L. leaves ameliorates experimental atherosclerosis by modulating lipid metabolism and inflammatory responses through PI3K/Akt and NF-κB singnaling pathways.

ETHNOPHARMACOLOGICAL RELEVANCE: Atherosclerosis (AS), a lipid-induced inflammatory condition of the arteries, is a primary contributor to atherosclerotic cardiovascular diseases including stroke. Arctium lappa L. leaf (ALL), an edible and medicinal herb in China, has been documented and commonly used for treating stroke since the ancient times. However, the elucidations on its anti-AS effects and molecular mechanism remain insufficient. AIM OF THE STUDY: To investigate the AS-ameliorating effects and the underlying mechanism of action of an ethanolic extract of leaves of Arctium lappa L. (ALLE). MATERIALS AND METHODS: ALLE was reflux extracted using with 70% ethanol. An HPLC method was established to monitor the quality of ALLE. High fat diet (HFD) and vitamin D3-induced experimental AS in rats were used to determine the in vivo effects; and oxidized low-density lipoprotein-induced RAW264.7 macrophage foam cells were used for in vitro assays. Simvatatin was used as positive control. Biochemical assays were implemented to ascertain the secretions of lipids and pro-inflammatory mediators. Haematoxylin-eosin (H&E) and Oil red O stains were employed to assess histopathological alterations and lipid accumulation conditions, respectively. CCK-8 assays were used to measure cytotoxicity. Immunoblotting assay was conducted to measure protein levels. RESULTS: ALLE treatment significantly ameliorated lipid deposition and histological abnormalities of aortas and livers in AS rats; improved the imbalances of serum lipids including total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C) and high-density lipoprotein cholesterol (HDL-C); notably attenuated serum concentrations of inflammation-associated cytokines/molecules including TNF-α, IL-6, IL-1ß, VCAM-1, ICAM-1and MMP-9. Mechanistic studies demonstrated that ALLE suppressed the phosphorylation/activation of PI3K, Akt and NF-κB in AS rat aortas and in cultured foam cells. Additionally, the PI3K agonist 740Y-P notably reversed the in vitro inhibitory effects of ALLE on lipid deposition, productions of TC, TNF-α and IL-6, and protein levels of molecules of PI3K/Akt and NF-κB singnaling pathways. CONCLUSIONS: ALLE ameliorates HFD- and vitamin D3-induced experimental AS by modulating lipid metabolism and inflammatory responses, and underlying mechanisms involves inhibition of the PI3K/Akt and NF-κB singnaling pathways. The findings of this study provide scientific justifications for the traditional application of ALL in managing atherosclerotic diseases.

Tubson-2 decoction ameliorates rheumatoid arthritis complicated with osteoporosis in CIA rats involving isochlorogenic acid A regulating IL-17/MAPK pathway.

BACKGROUND: Osteoporosis (OP) is considered as one of the major comorbidities of rheumatoid arthritis (RA), and is responsible for fragility fracture. However, there is currently no effective treatment for RA complicated with OP. Tubson-2 decoction (TBD), a Mongolian medicine also known as Erwei Duzhong Decoction, has been shown to exert a preventive effect on post-menopausal osteoporosis (PMOP). The preventive effects of TBD on RA-induced OP, as well as the bioactive compound responsible and the underlying mechanisms, remain to be elucidated. OBJECTIVE: To explore the effects of TBD on RA-induced OP in vivo, and to elucidate the mechanism of isochlorogenic acid A (ICA), the effective component of TBD, in vitro. METHODS: To evaluate the anti-arthritic and anti-osteoporotic effects of TBD, we conducted H&E straining and safranine O/fast green, TEM, immunohistochemistry (IHC), bone histomorphometry, micro-CT imaging, and biomechanical testing in collagen induced arthritis (CIA) rats. The active ingredient in TBD was identified using network pharmacology and molecular docking. The identification was supported by in vivo IHC assay, and further confirmed using qRT-PCR, Western blot, and SEM analysis in TNF-α-treated MH7A cells and/or in LPS-exposed RAW264.7 cells. RESULTS: Oral administration of TBD attenuated the severity of arthritis and osteopenia as well as poor bone quality, in CIA rats. Additionally, TBD and the positive control, tripterygium glycosides (TG), exhibited similar effects in reducing inflammation in both the synovium and ankle joint. They also were both effective in improving bone loss, microarchitecture, and overall bone quality. TBD reduced the expression of MMP13, IL-17, and p-JNK protein in the synovium of CIA rats. ICA, which was screened, suppressed TNF-α or LPS-triggered inflammatory responses via down-regulating IL-17 signaling, involving in MMP13, IL-1ß, IL-23, and IL-17, and the MAPK pathway including p-ERK, p-JNK, and p-P38, both in MH7A cells and in RAW264.7 cells. Furthermore, ICA prevented osteoclasts from differentiating and bone resoprtion in a dose-dependent manner in vitro. CONCLUSION: This study provides the first evidence that TBD exerts intervening effects on RA-induced OP, possibly through the downregulation of the IL-17/MAPK signaling pathway by ICA. The findings of our study provides valuable insights for further research in this area.

Isochlorogenic acid A alleviates dextran sulfate sodium-induced ulcerative colitis in mice through STAT3/NF-кB pathway.

Isochlorogenic acid A (ICGA-A) is a dicaffeoylquinic acid widely found in various medicinal plants or vegetables, such as Lonicerae japonicae Flos and chicory, and multiple properties of ICGA-A have been reported. However, the therapeutic effect of ICGA-A on colitis is not clear, and thus were investigated in our present study, as well as the underlying mechanisms. Here we found that ICGA-A alleviated clinical symptoms of dextran sodium sulfate (DSS) induced colitis model mice, including disease activity index (DAI) and histological damage. In addition, DSS-induced inflammation was significantly attenuated in mice given ICGA-A supplementation. ICGA-A reduced the fraction of neutrophils in peripheral blood and the infiltration of neutrophils and macrophages in colon tissue, and reduced pro-inflammatory cytokine production and tight junctions in mouse models. Furthermore, ICGA-A down-regulated expression of STAT3 and up-regulated the protein level of IκBα. Our in vitro studies confirmed that ICGA-A inhibited the mRNA expression of pro-inflammatory cytokines. ICGA-A blocked the phosphorylation of STAT3, p65, and IκBα, suppressed the expression STAT3 and p65. In addition, the present study also demonstrated that ICGA-A had no obvious toxicity on normal cells and organs. Taken together, we conclude that ICGA-A mitigates experimental ulcerative colitis (UC) at least in part by inhibiting the STAT3/NF-кB signaling pathways. Hence, ICGA-A may be a promising and effective drug for treating UC.

Difference analysis of different parts of chicory based on HPLC fingerprint and multi-component content determination.

Objective: To establish HPLC fingerprints of different parts of chicory stems, leaves, roots, flowers and seeds, and compare the similarities and differences of chemical components in different parts, so as to provide a scientific basis for the comprehensive utilization of chicory. Methods: To establish the HPLC fingerprint of chicory, the chromatographic column was chosen with Agilent ZORBAX Eclipse XDB-C18, the mobile phase was methanol (A) - 0.2% formic acid (B), the flow rate was 1 mL/min, the column temperature was 30 °C, and the detection wavelength was 254 nm. The Similarity Evaluation System of Chromatographic Fingerprint of Traditional Chinese Medicine (2012 Edition) was used to evaluate the similarity of different parts of decoction pieces, and the determination method of multi-component content was established based on fingerprint identification chromatographic peaks, and the determination results were analyzed. Results: The HPLC fingerprinting method of chicory was established. Sixteen chromatographic peaks were identified and 10 of them were identified as: caftaric acid (1), esculin (2), chlorogenic acid (3), esculetin (4), caffeic acid (5), cichoric acid (8), hyperoside (11), rutin (12), isochlorogenic acid C (14) and luteolin (16). The similarity of different parts was 0.084-0.701. At the same time, the total content of detected chemical components was ranked as flower > leaf > stem > root > seed. Roots did not contain caftaric acid, rutin, and luteolin, flowers did not contain luteolin, and seeds did not contain caftaric acid, cichoric acid, and luteolin. The content of cichoric acid in leaves was the most, and esculin in flowers was the most. Conclusion: The results of HPLC fingerprint and multi-component content determination revealed the similarity and difference of different parts of chicory from chemical composition, indicating that there were certain differences in different parts of chicory. The established HPLC fingerprinting method can provide a reference for quality control and evaluation of different parts of the chicory.

[Evaluation and optimization of content determination method of Chrysanthemi Flos in Chinese Pharmacopoeia].

This study discovered that the resolution of 3,5-O-dicaffeoylquinic acid(isochlorogenic acid A) in the content determination method of Chrysanthemi Flos in Chinese Pharmacopoeia(ChP)(2020 edition) was poor, which affected accurate quantification. We tested the method in ChP with chromatographic columns of seven brands to clarify the problems in the existing method, optimized the chromatographic conditions by adjusting the mobile phase composition and elution ratio and replacing the chromatographic column packing, and carried out the reproducibility assay for the new method. The two methods were compared for the content determination results of Chrysanthemi Flos prepared from six different varieties. As evaluated by the resolution based on different chromatographic columns of seven brands, the existing method failed to separate isochlorogenic acid A and isochlorogenic acid D well. The peaks of the two components were not completely separated on three chromatographic columns, and isochlorogenic acid A and isochlorogenic acid D generated a co-effluent peak in the other four columns. Isochlorogenic acid A and isochlorogenic acid D could be completely separated under the optimized chromatographic conditions. The difference in the peak areas of isochlorogenic acid A+isochlorogenic acid D obtained by the optimized method and the method in ChP was not significant, with deviation less than 3.0%, which further proved that the result measured by the method in ChP was the co-effluent of isochlorogenic acid A and isochlorogenic acid D. The optimized method can ensure the accurate quantification of isochlorogenic acid A. The existing content determination method of Chrysanthemi Flos has the problem of poor resolution. It is recommended to revise the chromatographic conditions for the content determination method of Chrysanthemi Flos to improve the resolution of isochlorogenic acid A and ensure its accurate quantification.

Comparative effectiveness of phlegm-heat clearing Chinese medicine injections for AECOPD: A systematic review and network meta-analysis.

ETHNOPHARMACOLOGICAL RELEVANCE: Qingkailing (QKL), Reduning (RDN), Xiyanping (XYP), Tanreqing (TRQ) and Yuxingcao (YXC) injections are all phlegm-heat clearing Chinese medicine (CM) injections composed of the extract from traditional CM materials. Evidence from clinical studies and animal experiments indicates that the above CM injections are effective supplementary therapy for acute exacerbation chronic obstructive pulmonary disease (AECOPD), and clinicians are faced with a difficult choice on the optimal phlegm-heat clearing CM injection for AECOPD. AIM OF THE STUDY: This systematic review and Bayesian network meta-analysis aimed to evaluate the comparative effectiveness of five commonly used phlegm-heat clearing CM injections for COPD. MATERIALS AND METHODS: A pairwise and network meta-analyses were performed to assess the effectiveness of QKL, RDN, TRQ, XYP and YXC on AECOPD. Randomized controlled trials (RCTs) were identified by searching English and Chinese databases. The primary outcome was lung function (forced expiration volume [FEV1] and forced vital capacity [FVC]), blood gas analysis index was secondary outcome measure. Winbugs and Stata 15.0 software were used for data analysis. RESULTS: A total of 57 RCTs were included. The pairwise analyses showed that each of the injections combined with routine treatment were superior to routine treatment alone [FEV1: QKL, MD 0.20, 95% CI (0.06, 0.35); RDN, MD 0.24, 95% CI (0.08, 0.40); TRQ, MD 0.24, 95% CI (0.19, 0.29); XYP, MD 0.26, 95% CI (0.20, 0.32); YXC MD 0.73, 95% CI (0.06, 1.41)]. The network meta-analysis provided the following rank of lung function improvement: FEV1: YXC > TRQ > XYP > RDN > QKL; FVC: YXC > TRQ > QKL > RDN > XYP. RDN and YXC ranked highest in blood gas analysis index. RDN was the highest ranked injection for effectiveness, followed by QKL, TRQ, XYP, then YXC. Most of the injections appeared safe, with severe adverse events rarely reported. CONCLUSION: This study suggests that YXC and TRQ are the most effective therapies in treating AECOPD patients. RDN and YXC are more effective in the alleviation of clinical symptoms. Given that the safety of YXC is controversial, TRQ and RDN may be preferable as phlegm-heat clearing CM injections in the adjuvant treatment of AECOPD.

Weed Suppression and Molecular Mechanisms of Isochlorogenic Acid A Isolated from Artemisia argyi Extract via an Activity-Guided Method.

Allelopathy is considered an environmentally friendly and resource-conserving approach to weed control because allelochemicals degrade easily and cause less pollution than traditional chemical herbicides. In this study, the allelopathic active constituents of Artemisia argyi were elucidated by activity-guided isolation and ultraperformance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS). First, a crude extract prepared in water was fractionated using macroporous resin D101 to obtain three fractions (Fr.A-C). Combined with the allelopathic activity assay on Setaria viridis and Portulaca oleracea, Fr.C was determined to be the most active fraction. We identified 14 compounds in the active fraction (Fr.C) using UPLC-QTOF-MS, including 13 phenolic compounds. Accordingly, phenolic components have been suggested as the main allelochemicals in A. argyi. Thereafter, Fr.C was further isolated by octadecylsilyl (ODS) chromatography to obtain eight subfractions (Fr.C-1-Fr.C-8). Finally, isochlorogenic acid A (ICGAA) was purified from Fr.C-3 by semipreparative liquid chromatography, which was detected in the growth environment of A. argyi. Furthermore, we evaluated the allelopathic effects of ICGAA on six weeds from different families and genera for the first time. The results showed that ICGAA is a novel allelochemical with broad herbicidal activity. In addition, we analyzed the inhibitory effect and molecular mechanism of ICGAA on the growth of S. viridis seedlings. Optical microscopy and transmission electron microscopy (TEM) revealed the degradation of membrane structures and organelles after ICGAA treatment. Transcriptome and real-time polymerase chain reaction (RT-qPCR) analysis showed that ICGAA inhibited the growth of weeds mainly by inhibiting the diterpenoid biosynthesis pathway (especially gibberellins, GAs). The decrease of gibberellin (GA) contents after ICGAA treatment also confirmed these results. In brief, this study provides new material sources and theoretical support for developing biological herbicides for agroecosystems.

Evaluation and optimization of content determination method of Chrysanthemi Flos in Chinese Pharmacopoeia / 中国中药杂志

This study discovered that the resolution of 3,5-O-dicaffeoylquinic acid(isochlorogenic acid A) in the content determination method of Chrysanthemi Flos in Chinese Pharmacopoeia(ChP)(2020 edition) was poor, which affected accurate quantification. We tested the method in ChP with chromatographic columns of seven brands to clarify the problems in the existing method, optimized the chromatographic conditions by adjusting the mobile phase composition and elution ratio and replacing the chromatographic column packing, and carried out the reproducibility assay for the new method. The two methods were compared for the content determination results of Chrysanthemi Flos prepared from six different varieties. As evaluated by the resolution based on different chromatographic columns of seven brands, the existing method failed to separate isochlorogenic acid A and isochlorogenic acid D well. The peaks of the two components were not completely separated on three chromatographic columns, and isochlorogenic acid A and isochlorogenic acid D generated a co-effluent peak in the other four columns. Isochlorogenic acid A and isochlorogenic acid D could be completely separated under the optimized chromatographic conditions. The difference in the peak areas of isochlorogenic acid A+isochlorogenic acid D obtained by the optimized method and the method in ChP was not significant, with deviation less than 3.0%, which further proved that the result measured by the method in ChP was the co-effluent of isochlorogenic acid A and isochlorogenic acid D. The optimized method can ensure the accurate quantification of isochlorogenic acid A. The existing content determination method of Chrysanthemi Flos has the problem of poor resolution. It is recommended to revise the chromatographic conditions for the content determination method of Chrysanthemi Flos to improve the resolution of isochlorogenic acid A and ensure its accurate quantification.

Anti-inflammatory property and functional substances of Lonicerae Japonicae Caulis.

ETHNOPHARMACOLOGICAL RELEVANCE: Lonicerae Japonicae Caulis, the dried stem and branch of Lonicera japonica Thunb., is a Chinese Materia Medica known as Ren Dong Teng in Chinese with long use history in the traditional Chinese medicine (TCM) prescriptions. Lonicerae Japonicae Caulis possesses heat-clearing and detoxifying functions according to the TCM theory. In recent years, a large amount of experimental and clinical studies proved good anti-inflammatory effects of some heat-clearing and detoxifying herbs. The present study aims to reveal the anti-inflammatory property and functional substances of Lonicerae Japonicae Caulis. MATERIALS AND METHODS: For anti-inflammatory activity test, LPS-induced RAW 264.7 macrophages, DSS-induced SPF male C57BL/6J mice model, and LPS-induced SPF male ICR mice model were used in vitro and in vivo, respectively. The behavioral changes, organ damage, and the expression of inflammatory factors such as TNT-α and IL-6 mRNA expression were measured for activity evaluation. Lonicerae Japonicae Caulis samples were prepared by solvent extraction and subsequent column chromatography. The main components were identified and determined using UPLC-UV analysis as well as NMR interpretation after purification. To testify the contribution of main components for the anti-inflammatory activity, different samples were also prepared by compound-knockout strategy. RESULTS: Ethanol extract of Lonicerae Japonicae Caulis could attenuate sickness symptoms in mice such as diarrhea, less activity, and depression. It could also alleviate multiple organ damage, and significantly inhibit the expression of pro-inflammatory factors such as TNF-α, IL-1ß, IL-6 and IFN-γ in mice. Furthermore, the isochlorogenic acid-rich and biflavonoid-rich fractions and isochlorogenic acids A and C, and ochnaflavone could significantly down-regulate the mRNA expression of TNF-α and IL-6 in LPS-induced RAW 264.7 macrophages. CONCLUSIONS: Lonicerae Japonicae Caulis possesses anti-inflammatory property. Its isochlorogenic acid-rich and biflavonoid-rich fractions do the major contribution. And their main components, isochlorogenic acids A and C, and ochnaflavone, take main responsibility for the anti-inflammatory property.

Isochlorogenic acid (ICGA): natural medicine with potentials in pharmaceutical developments.

Natural products have attracted a great deal of attention as significant resources in traditional Chinese medicine (TCM) and in chemical medicine, as well as in cosmetic ingredients, nutraceuticals and food products. Isochlorogenic acid (ICGA), which has medicinal value, has been discovered in various plants. As a widespread natural medicine, ICGA should be the subject of further research and development. However, there have been no systematic analyses of ICGA. According to our investigation, ICGA was initially isolated from green coffee extracts by Barnes et al. in 1950. To date, it has been discovered in a variety of tea, vegetables, medicinal diet and TCM materials. ICGA is used as a chemical marker for the quality control of these TCM materials. The metabolic process of ICGA has been studied in detail, conforming to be linear dynamics. Thus, the clear pharmacokinetics of ICGA offers a solid foundation for its research and development. ICGA has multiple biological and pharmacological effects, and studies have mainly focused on its antioxidant, anti-inflammatory, antimicrobial, hypoglycemic, neuroprotective, and cardiovascular protective effects, and hepatoprotective properties. The mechanisms underlying these effects are summarized in this review to provide scientific support and inspiration for the future research and development of ICGA and ICGA-rich natural products.

Three common caffeoylquinic acids as potential hypoglycemic nutraceuticals: Evaluation of α-glucosidase inhibitory activity and glucose consumption in HepG2 cells.

The demand for plant-derived antidiabetic nutraceuticals is increasing. In this study, the effects of three common caffeoylquinic acids (CQAs) (chlorogenic acid, isochlorogenic acid A, and cynarin) on α-glucosidase activity and glucose consumption in HepG2 cells were systematically compared. Their α-glucosidase inhibitory activities followed the order of isochlorogenic acid A > chlorogenic acid > cynarin. The fluorescence analysis indicated that they exerted the inhibitory role by forming the complex with α-glucosidase at the molar ratio of 1:1. Isochlorogenic acid A possessed the highest binding capacity, followed by chlorogenic acid and cynarin. The effect of caffeoyl group distribution on the α-glucosidase inhibitory activity was clarified by the molecular docking results. In the HepG2 cells, isochlorogenic acid A also showed the best glucose consumption with negligible cytotoxicity, which might be related to its reactive oxygen species scavenging capacity in cells. Our results confirm its potential application as the antidiabetic nutraceutical. PRACTICAL APPLICATIONS: The hypoglycemic activities of three common CQAs (chlorogenic acid, isochlorogenic acid A, and cynarin) were systemically evaluated in this study. Isochlorogenic acid A exhibited the strongest α-glucosidase inhibitory activity and highest glucose consumption in HepG2 cells with low cytotoxicity. The results suggest that isochlorogenic acid A can be used as the potential hypoglycemic nutraceutical in functional foods.

Identification of Transient Receptor Potential Vanilloid 3 Antagonists from Achillea alpina L. and Separation by Liquid-Liquid-Refining Extraction and High-Speed Counter-Current Chromatography.

Bioassay-guided fractionation of the ethanol extract of whole herbs of Achillea alpina led to the isolation of isochlorogenic acids A and B as transient receptor potential vanilloid 3 (TRPV3) channel antagonists by using a calcium fluorescent assay. The structures were identified by spectroscopic analysis and the inhibitory activities of isochlorogenic acids A and B were confirmed by whole-cell patch clamp recordings of human embryonic kidney 293 (HEK293) cells expressing human TRPV3. Molecular docking results revealed that these two compounds reside in the same active pocket of human TRPV3 channel protein with lower binding energy than the agonist 2-aminoethoxydiphenyl borate (2-APB). High-speed counter-current chromatography (HSCCC) coupled with a liquid-liquid extraction approach was successfully established for the separation of isochlorogenic acids A and B from the whole herbs of A. alpina. Ethyl acetate and n-hexane-ethyl acetate-water (3:3:4 and 1:5:4, v/v/v) were selected as liquid-liquid extraction solvent systems to remove high- and low-polarity impurities in the mixture. Sixty g of ethanol extract was refined by solvent partition to yield 1.7 g of the enriched fraction, of which 480 mg in turn obtained 52.5 mg of isochlorogenic acid B (purity 98.3%) and 37.6 mg isochlorogenic acid A (purity 96.2%) after HSCCC with n-hexane-ethyl acetate-water containing 1% acetic acid (1:4:8, v/v/v).

Isochlorogenic Acid A Attenuates the Progression of Liver Fibrosis Through Regulating HMGB1/TLR4/NF-κB Signaling Pathway.

Liver fibrosis, a chronic damage process related to further progression of hepatic cirrhosis, has yet no truly effective treatment. Isochlorogenic acid A (ICQA), isolated from a traditional Chinese herbal medicine named Laggera alata (DC.) Sch.Bip. ex Oliv. (Asteraceae), is proved to exhibit anti-inflammatory, hepatoprotective and antiviral properties. However, the actions of ICQA on liver fibrosis are poorly understood. The purpose of this study was to evaluate the actions of ICQA on liver fibrosis and clarify the underlying mechanism. It was found that ICQA had significant protective actions on liver injury, inflammation as we as fibrosis in rats. Meanwhile, ICQA prevented hepatic stellate cells (HSC) activation, indicated by its inhibitory effect on the overexpression of α-smooth muscle actin (α-SMA). In addition, the reduced fibrosis was found to be associated with the decreased protein expression of high-mobility group box 1 (HMGB1) as well as toll like receptor (TLR) 4. Simultaneously, ICQA can suppress the cytoplasmic translocation of HMGB1 in rat liver. Further investigations indicated that ICQA treatment dramatically attenuated the nuclear translocation of the nuclear factor-kB (NF-κB) p65 and suppressed the hepatic expression of p-IκBα in rats with liver fibrosis. Taken together, our study indicated that ICQA could protect against CCl4-induced liver fibrosis probably through suppressing the HMGB1/TLR4/NF-κB signaling pathways.

Isochlorogenic acid (ICGA): natural medicine with potentials in pharmaceutical developments / 中国天然药物

Natural products have attracted a great deal of attention as significant resources in traditional Chinese medicine (TCM) and in chemical medicine, as well as in cosmetic ingredients, nutraceuticals and food products. Isochlorogenic acid (ICGA), which has medicinal value, has been discovered in various plants. As a widespread natural medicine, ICGA should be the subject of further research and development. However, there have been no systematic analyses of ICGA. According to our investigation, ICGA was initially isolated from green coffee extracts by Barnes et al. in 1950. To date, it has been discovered in a variety of tea, vegetables, medicinal diet and TCM materials. ICGA is used as a chemical marker for the quality control of these TCM materials. The metabolic process of ICGA has been studied in detail, conforming to be linear dynamics. Thus, the clear pharmacokinetics of ICGA offers a solid foundation for its research and development. ICGA has multiple biological and pharmacological effects, and studies have mainly focused on its antioxidant, anti-inflammatory, antimicrobial, hypoglycemic, neuroprotective, and cardiovascular protective effects, and hepatoprotective properties. The mechanisms underlying these effects are summarized in this review to provide scientific support and inspiration for the future research and development of ICGA and ICGA-rich natural products.

Anti-HIV activity of southern African plants: Current developments, phytochemistry and future research.

ETHNOPHARMACOLOGICAL RELEVANCE: The African continent is home to a large number of higher plant species used over centuries for many applications, which include treating and managing diseases such as HIV. Due to the overwhelming prevalence and incidence rates of HIV, especially in sub-Saharan Africa, it is necessary to develop new and affordable treatments. AIM OF THE STUDY: The article provides an extensive overview of the status on investigation of plants from the southern African region with ethnobotanical use for treating HIV or HIV-related symptoms, or the management of HIV. The review also provide an account of the in vitro assays, anti-viral activity and phytochemistry of these plants. MATERIALS AND METHODS: Peer-reviewed articles investigating plants with ethnobotanical information for the treatment or management of HIV or HIV-related symptoms from the southern African region were acquired from Science Direct, PubMed central and Google Scholar. The selection criteria was that (1) plants should have a record of traditional/popular use for infectious or viral diseases, HIV treatment or symptoms similar to HIV infection, (2) if not traditionally/popularly used, plants should be closely related to plants with popular use and HIV activity identified by means of in vitro assays, (3) plants should have been identified scientifically, (4) should be native to southern African region and (5) anti-HIV activity should be within acceptable ranges. RESULTS: Many plants in Africa and specifically the southern African region have been used for the treatment of HIV or HIV related symptoms and have been investigated suing various in vitro techniques. In vitro assays using HIV enzymes such as reverse transcriptase (RT), integrase (IN) and protease (PR), proteins or cell-based assays have been employed to validate the use of these plants with occasional indication of the selectivity index (SI) or therapeutic index (TI), with only one study, that progressed to in vivo testing. The compounds identified from plants from southern Africa is similar to compounds identified from other regions of the world, and the compounds have been divided into three groups namely (1) flavonoids and flavonoid glycosides, (2) terpenoids and terpenoid glycosides and (3) phenolic acids and their conjugated forms. CONCLUSIONS: An investigation of the plants from southern Africa with ethnobotanical use for the treatment of HIV, management of HIV or HIV-related symptoms, therefore provide a very good analysis of the major assays employed and the anti-viral compounds and compound groups identified. The similarity in identified anti-viral compounds worldwide should support the progression from in vitro studies to in vivo testing in development of affordable and effective anti-HIV agents for countries with high infection and mortality rates due to HIV/AIDS.

Characterization of isochlorogenic acid A metabolites in rats using high-performance liquid chromatography/quadrupole time-of-flight mass spectrometry.

Isochlorogenic acid A is widely present in fruits, vegetables and herbal medicines, and is characterized by anti-inflammatory, hepatoprotective and antiviral properties. However, little is known about its metabolic fate and pharmacokinetic properties. This study is thus designed to investigate the metabolic fate of isochlorogenic acid A. An analytical method based on high-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (HPLC/Q-TOF MS) was established to characterize the metabolites of isochlorogenic acid A in the plasma, urine and feces of rats. A total of 32 metabolites were identified. The metabolic pathways mainly include hydrolyzation, dehydroxylation, hydrogenation and conjugation with methyl, glucuronic acid, glycine, sulfate, glutathione and cysteine. Moreover, the pharmacokinetic profiles of all the circulating metabolites were investigated. M11 resulting from hydrolyzation, dehydroxylation and hydrogenation was the dominant circulating metabolite after the intragastric administration of isochlorogenic acid A. The results obtained will be useful for further study of elucidating potential bioactive metabolites which can provide better explanation of the pharmacological and/or toxicological effects of this compound.

Isochlorogenic acid A affects P450 and UGT enzymes in vitro and in vivo.

Isochlorogenic acid A (ICQA), which has anti-inflammatory, hepatoprotective, and antiviral properties, is commonly presented in fruits, vegetables, coffee, plant-based food products, and herbal medicines. These herbal medicines are usually used in combination with other medicines in the clinic. However, little is known about the regulatory effects of ICQA on drug-metabolizing enzymes and the herb-drug interactions. In the present study, we evaluated the inhibitory potentials of ICQA on CYP1A2, CYP2C9, CYP2C19, CYP3A4, CYP2D6, and CYP2E1 in vitro based on a cocktail approach. The P450 and UGT activities in mice treated with ICQA for a prolonged period were also determined. Our results demonstrated that ICQA exhibited a weak inhibitory effect on CYP2C9 in human liver microsomes with IC50 being 57.25 µmol·L-1 and Ki being 26.77 µmol·L-1. In addition, ICQA inhibited UGT1A6 activity by 25%, in the mice treated with ICQA (i.p.) at 30 mg·kg-1 for 14 d, compared with the control group. Moreover, ICQA showed no mechanism-based inhibition on CYP2C9 or UGT1A6. In conclusion, our results further confirm a safe use of ICQA in clinical practice.

Immuno-enhancement effects of Yifei Tongluo Granules on cyclophosphamide-induced immunosuppression in Balb/c mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Traditional Chinese medicine Yifei Tongluo Granules has been employed clinically with the combination of chemotherapy agents to treat patients with multidrug-resistant tuberculosis. However, the mechanisms underlying the therapeutic potential have not been well elucidated. The present study was employed to verify immunomodulatory effect and to investigate the underlying mechanisms which have not been explored. MATERIALS AND METHODS: The study samples of total extracts (FB-E) and polysaccharides (FB-P) were prepared by the extraction of the Yifei Tongluo Granules using appropriate techniques. A simple immunodeficient mice model was established by challenging Balb/c mice with cyclophosphamide in order to avoid the handling of tuberculosis viruses. The in vivo study was thus designed to systematically elucidate the immuno-enhancement effects of Yifei Tongluo Granules extracts in immunosuppressed mice induced by cyclophosphamide. Balb/c mice were orally ingested once daily with the low and high doses of two different extracts for ten consecutive days, respectively, accompanied by intraperitoneal injection of cyclophosphamide (60mg/kg) on days 1-3 and 10. RESULTS: Compared with the model group, the treatment of immunodeficient mice with the low and high doses of the extracts FB-E or FB-P enhanced spleen and thymus indices, T- and B-cell proliferation as well as increased the activities of splenic natural killer, lymphokine activated killer, cytotoxic T lymphocyte cells and peritoneal macrophage phagocytosis. In addition, the FB-E or FB-P treatment balanced the ratio of Th1/Th2 and up-regulated the CD4+/CD8+ ratio in the serum. CONCLUSIONS: These results demonstrate, for the first time, that the treatment of the cyclophosphamide-challenged mice with the Yifei Tongluo Granules extracts resulted in accelerated recovery of immunosuppression, sugguesting that the immunomodulation might be the mechanism for the observed clinical benefits of Yifei Tongluo Granules. Our findings provide preliminary mechanistic study evidences for clinical application of Yifei Tongluo Granules in patients with immunodeficient diseases such as tuberculosis.

Isochlorogenic acid A affects P450 and UGT enzymes in vitro and in vivo / 中国天然药物

Isochlorogenic acid A (ICQA), which has anti-inflammatory, hepatoprotective, and antiviral properties, is commonly presented in fruits, vegetables, coffee, plant-based food products, and herbal medicines. These herbal medicines are usually used in combination with other medicines in the clinic. However, little is known about the regulatory effects of ICQA on drug-metabolizing enzymes and the herb-drug interactions. In the present study, we evaluated the inhibitory potentials of ICQA on CYP1A2, CYP2C9, CYP2C19, CYP3A4, CYP2D6, and CYP2E1 in vitro based on a cocktail approach. The P450 and UGT activities in mice treated with ICQA for a prolonged period were also determined. Our results demonstrated that ICQA exhibited a weak inhibitory effect on CYP2C9 in human liver microsomes with IC being 57.25 μmol·L and Ki being 26.77 μmol·L. In addition, ICQA inhibited UGT1A6 activity by 25%, in the mice treated with ICQA (i.p.) at 30 mg·kg for 14 d, compared with the control group. Moreover, ICQA showed no mechanism-based inhibition on CYP2C9 or UGT1A6. In conclusion, our results further confirm a safe use of ICQA in clinical practice.