Extraction, characterization and intestinal anti-inflammatory and anti-oxidative activities of polysaccharide from stems and leaves of Chuanminshen violaceum M. L. Sheh & R. H. Shan.

ETHNOPHARMACOLOGICAL RELEVANCE: Chuanminshen violaceum M. L. Sheh & R. H. Shan (CV) is used as a medicine with roots, which have the effects of benefiting the lungs, harmonizing the stomach, resolving phlegm and detoxifying. Polysaccharide is one of its main active components and has various pharmacological activities, but the structural characterization and pharmacological activities of polysaccharide from the stems and leaves parts of CV are still unclear. AIM OF THE STUDY: The aim of this study was to investigate the optimal extraction conditions for ultrasound-assisted extraction of polysaccharide from CV stems and leaves, and to carry out preliminary structural analyses, anti-inflammatory and antioxidant effects of the obtained polysaccharide and to elucidate the underlying mechanisms. MATERIALS AND METHODS: The ultrasonic-assisted extraction of CV stems and leaves polysaccharides was carried out, and the response surface methodology (RSM) was used to optimize the extraction process to obtain CV polysaccharides (CVP) under the optimal conditions. Subsequently, we isolated and purified CVP to obtain the homogeneous polysaccharide CVP-AP-I, and evaluated the composition, molecular weight, and structural features of CVP-AP-I using a variety of technical methods. Finally, we tested the pharmacological activity of CVP-AP-Ⅰ in an LPS-induced model of oxidative stress and inflammation in intestinal porcine epithelial cells (IPEC-J2) and explored its possible mechanism of action. RESULTS: The crude polysaccharide was obtained under optimal extraction conditions and subsequently isolated and purified to obtain CVP-AP-Ⅰ (35.34 kDa), and the structural characterization indicated that CVP-AP-Ⅰ was mainly composed of galactose, galactose, rhamnose and glucose, which was a typical pectic polysaccharide. In addition, CVP-AP-Ⅰ attenuates LPS-induced inflammation and oxidative stress by inhibiting the expression of pro-inflammatory factor genes and proteins and up-regulating the expression of antioxidant enzyme-related genes and proteins in IPEC-J2, by a mechanism related to the activation of the Nrf2/Keap1 signaling pathway. CONCLUSION: The results of this study suggest that the polysaccharide isolated from CV stems and leaves was a pectic polysaccharide with similar pharmacological activities as CV roots, exhibiting strong anti-inflammatory and antioxidant activities, suggesting that CV stems and leaves could possess the same traditional efficacy as CV roots, which is expected to be used in the treatment of intestinal diseases.

Ultrasonic-assisted extraction of polysaccharide from Paeoniae Radix alba: Extraction optimization, structural characterization and antioxidant mechanism in vitro.

Paeoniae Radix alba is used in Traditional Chinese Medicine for the treatment of gastrointestinal disorders, immunomodulatory, cancer, and other diseases. In the current study, the yield of Paeoniae Radix alba polysaccharide (PRP) was significantly increased with optimal ultrasound-assisted extraction compared to hot water extraction. Further, an acidic polysaccharide (PRP-AP) was isolated from PRP after chromatographic separation and was characterized as a typical pectic polysaccharide with side chains of arabinogalactans types I and II. Moreover, it showed antioxidant effects on LPS-induced damage on IPEC-J2 cells determined by qRT-PCR and ELISA, including decreasing the pro-inflammatory factors' expressions and increasing the antioxidant enzymes activities, which was shown to be related to the Nrf2/Keap1 pathway modulated by PRP-AP. The metabolites change (such as itaconate, cholesterol sulfate, etc.) detected by untargeted metabolomic analysis in cells was also shown to be modulated by PRP-AP, and these metabolites were further utilized and protected cells damaged by LPS. These results revealed the cellular active mechanism of the macromolecular PRP-AP on protecting cells, and supported the hypothesis that PRP-AP has strong benefits as an alternative dietary supplement for the prevention of intestinal oxidative stress by modulating cellular metabolism.

Polysaccharide from aerial part of Chuanminshen violaceum alleviates oxidative stress and inflammatory response in aging mice through modulating intestinal microbiota.

Aging is a biological process of progressive deterioration of physiological functions, which poses a serious threat to individual health and a heavy burden on public health systems. As population aging continues, research into anti-aging drugs that prolong life and improve health is of particular importance. In this study, the polysaccharide from stems and leaves of Chuanminshen violaceum was obtained with water extraction and alcohol precipitation, and then separated and purified with DEAE anion exchange chromatography and gel filtration to obtain CVP-AP-I. We gavaged natural aging mice with CVP-AP-I and performed serum biochemical analysis, histological staining, quantitative real-time PCR (qRT-PCR) and ELISA kit assays to analyze inflammation and oxidative stress-related gene and protein expression in tissues, and 16SrRNA to analyze intestinal flora. We found that CVP-AP-I significantly improved oxidative stress and inflammatory responses of the intestine and liver, restored the intestinal immune barrier, and balanced the dysbiosis of intestinal flora. In addition, we revealed the potential mechanism behind CVP-AP-I to improve intestinal and liver function by regulating intestinal flora balance and repairing the intestinal immune barrier to regulate the intestinal-liver axis. Our results indicated that C. violaceum polysaccharides possessed favorable antioxidant, anti-inflammatory and potentially anti-aging effects in vivo.

Angelica sinensis aboveground part polysaccharide and its metabolite 5-MT ameliorate colitis via modulating gut microbiota and TLR4/MyD88/NF-κB pathway.

The roots of Angelica sinensis have been used in Traditional Chinese Medicine for thousands of years. However, tons of aerial parts of this herb (aboveground part) are commonly discarded during the process of root preparations. A polysaccharide (ASP-Ag-AP) in the aboveground parts of A. sinensis was isolated and preliminarily characterized as typical plant pectin. ASP-Ag-AP exhibited noticeable protective effects against dextran sodium sulfate (DSS)-induced colitis, including reduction of colonic inflammation, modulation of barrier function, and alteration of gut microbiota and serum metabolite profile. Anti-inflammatory effects of ASP-Ag-AP were observed by inhibiting TLR4/MyD88/NF-κB signaling pathway in vitro and in vivo. Additionally, the level of serum metabolite 5-methyl-dl-tryptophan (5-MT) was reduced by DSS and restored by ASP-Ag-AP, which also negatively correlated with Bacteroides, Alistipes, Staphylococcus and pro-inflammatory factors. The protection from inflammatory stress on intestinal porcine enterocytes cells (IPEC-J2) of 5-MT was observed through the inhibition of TLR4/MyD88/NF-κB pathway. Besides, 5-MT also exhibited robust anti-inflammatory effect in colitis mice with improving colitis symptoms, barrier function and gut microbiota, which was the same as presented by ASP-Ag-AP. Therefore, ASP-Ag-AP could be a promising agent for colitis prevention and 5-MT could be the signal metabolite of ASP-Ag-AP on defending against intestinal inflammatory stress.

Salvia miltiorrhiza polysaccharide and its related metabolite 5-methoxyindole-3-carboxaldehyde ameliorate experimental colitis by regulating Nrf2/Keap1 signaling pathway.

The roots of Salvia miltiorrhiza have been used in Traditional Chinese Medicine for thousands of years. However, tons of aerial parts of this plant are usually discarded in the production of roots preparation. To make better use of these plant resources, the polysaccharide isolated from the aerial part of S. miltiorrhiza was investigated for its potential protection against intestinal diseases. A pectic polysaccharide (SMAP-1) was isolated and characterized being composed of homogalacturonan as the main chain and rhamnogalacturonan type I as ramified region, with side chains including arabinans and possible arabinogalactan type I and II. SMAP-1 exhibited robust protective effects against dextran sodium sulfate (DSS)-induced colitis and restored colitis symptoms, colonic inflammation, and barrier functions. Anti-oxidative effects were also observed by up-regulating Nrf2/Keap1 signaling pathway. Additionally, the level of serum 5-methoxyindole-3-carboxaldehyde (5-MC) was restored by SMAP-1 identified in metabolomic analysis, being correlated with the aforementioned effects. Protection against oxidative stress on intestinal porcine enterocyte cells (IPEC-J2) by 5-MC was observed through the activation of Nrf2/Keap1 system, as also shown by SMAP-1. In conclusion, SMAP-1 could be a promising candidate for colitis prevention, and 5-MC could be the signal metabolite of SMAP-1 in protecting against oxidative stress in the intestine.

Structural features and antioxidant activities of polysaccharides from different parts of Codonopsis pilosula var. modesta (Nannf.) L. T. Shen.

In this study, three acidic polysaccharides from different plant parts of Codonopsis pilosula var. Modesta (Nannf.) L. T. Shen were obtained by ion exchange chromatography and gel filtration chromatography, and the yields of these three polysaccharides were different. According to the preliminary experimental results, the antioxidant activities of the polysaccharides from rhizomes and fibrous roots (CLFP-1) were poor, and was thus not studied further. Due to this the structural features of polysaccharides from roots (CLRP-1) and aerial parts (CLSP-1) were the object for this study and were structurally characterized, and their antioxidant activities were evaluated. As revealed by the results, the molecular weight of CLRP-1and CLSP-1 were 15.9 kDa and 26.4 kDa, respectively. The monosaccharide composition of CLRP-1 was Ara, Rha, Fuc, Xyl, Man, Gal, GlcA, GalA in a ratio of 3.8: 8.4: 1.0: 0.8: 2.4: 7.4: 7.5: 2.0: 66.7, and Ara, Rha, Gal, GalA in a ratio of 5.8: 8.9: 8.0: 77.0 in for CLSP-1. The results of structural elucidation indicated that both CLRP-1 and CLSP-1 were pectic polysaccharides, mainly composed of 1, 4-linked galacturonic acid with long homogalacturonan regions. Arabinogalactan type I and arabinogalactan type II were presented as side chains. The antioxidant assay in IPEC-J2 cells showed that both CLRP-1 and CLSP-1 promoted cell viability and antioxidant activity, which significantly increase the level of total antioxidant capacity and the activity of superoxide dismutase, catalase, and decrease the content of malondialdehyde. Moreover, CLRP-1 and CLSP-1 also showed powerful antioxidant abilities in Caenorhabditis elegans and might regulate the nuclear localization of DAF-16 transcription factor, induced antioxidant enzymes activities, and further reduced reactive oxygen species and malondialdehyde contents to increase the antioxidant ability of Caenorhabditis elegans. Thus, these finding suggest that CLRP-1 and CLSP-1 could be used as potential antioxidants.

The comparison of preliminary structure and intestinal anti-inflammatory and anti-oxidative activities of polysaccharides from different root parts of Angelica sinensis (Oliv.) Diels.

ETHNOPHARMACOLOGICAL RELEVANCE: The root of Angelica sinensis, has been commonly used in gynecology for centuries, and is normally applied divided into different parts in various clinical applications. At present, the majority of existing studies focus on the volatile oil and ferulic acid extracted from different parts of A. sinensis, but there is a dearth of scientific information on its water-soluble polysaccharides. AIM OF THE STUDY: The structures of polysaccharides from plants, have been reported contributing to multiple pharmacological activities such as anti-oxidative, anti-inflammatory, anti-tumor and liver protection. Therefore, the focus of this study was on its anti-oxidative and anti-inflammatory activities in vitro, which would be based on the various polysaccharides with distinct structures obtained from different parts of the A. sinensis root. MATERIALS AND METHODS: Four parts of A. sinensis root were separated according to the Chinese Pharmacopoeia: head, body, tail and whole body. Crude polysaccharides were obtained by water extraction and ethanol precipitation method, and were further fractionated by DEAE Sepharose chromatographic column and gel filtration. The comparison of ASPs from different root parts were performed, including chemical compositions determined by colorimetric analysis, monosaccharide compositions measured by high performance liquid chromatography (HPLC), glycosidic linkage units determined by methylation and gas chromatography-mass spectrometry (GC-MS), organic functional groups determined by FT-IR, molecular weight (Mw) demarcated by gel permeation chromatography, and the viscosities and solubilities were measured according to method published in the previous report with minor modification. In vitro biological activities of APSs were compared on lipopolysaccharide (LPS)-induced inflammatory and oxidative stress models on IPEC-J2 cells. RESULTS: Four purified polysaccharides, ASP-H-AP, ASP-B-AP, ASP-T-AP and ASP-Hb-AP from the root of A. sinensis, were obtained, and consisted of various contents of protein and the polyphenol. They were possibly pectic polysaccharides with a long homogalacturonan region as the main backbone and ramified with rhamnogalacturonan I region, but they were differed by subregions and the relative contents of glycosidic units. The Mw of four pectic polysaccharides were ranged from 67.9-267.7 kDa. The infrared spectrum also showed that the four polysaccharide fractions contained the characteristic peaks of polysaccharides. Their distinct primary structure could lead to a variety of biological activities. In vitro biological assays suggested that four polysaccharide fractions can protect IPEC-J2 cells against the LPS-induced inflammation by down-regulating inflammation factors and related genes on IPEC-J2 cells. These polysaccharides also could alleviate oxidative stress on IPEC-J2 cells by up-regulating the gene and protein expressions of antioxidant enzymes. It was concluded that ASP-H-AP possessed better anti-inflammatory and anti-oxidative effects, while those of ASP-T-AP was relatively poor among the four polysaccharide fractions. CONCLUSION: All results indicated that the structure of pectic polysaccharides from different root parts of A. sinensis differed, which lead to their distinct anti-inflammatory and anti-oxidative activities. This may also be one of the factors why different parts of A. sinensis showed various pharmacological activities and applied independently in traditional use. In addition, it would be valuable for further studies on structure-activity relationship of polysaccharides obtained by different root parts of A. sinensis.

Restorative Effects of Inulin From Codonopsis pilosula on Intestinal Mucosal Immunity, Anti-Inflammatory Activity and Gut Microbiota of Immunosuppressed Mice.

An inulin (CPPF), isolated from a traditional Chinese herbal medicine Codonopsis pilosula, was characterized and demonstrated with potential prebiotic activity in vitro before. Based on its non-digested feature, the intestinal mucosa and microbiota modulatory effects in vivo on immunosuppressed mice were investigated after oral administration of 200, 100 and 50 mg/kg of CPPF for 7 days. It was demonstrated that the secretions of sIgA and mucin 2 (Muc2) in ileum were improved by CPPF, and the anti-inflammatory activities in different intestine parts were revealed. The intestine before colon could be the target active position of CPPF. As a potential prebiotic substance, a gut microbiota restorative effect was also presented by mainly modulating the relative abundance of Eubacteriales, including Oscillibacter, unidentified Ruminococcus and Lachnospiraceae after high-throughput pyrosequencing of V4 region of 16S rRNA analysis. All these results indicated that this main bioactive ingredient inulin from C. pilosula was a medicinal prebiotic with enhancing mucosal immune, anti-inflammatory and microbiota modulatory activities.