Distinction and quantification of Panax polysaccharide extracts via attenuated total reflectance-Fourier transform infrared spectroscopy with first-order derivative processing.

Derivative spectroscopy is used to separate the small absorption peaks superimposed on the main absorption band, which is widely adopted in modern spectral analysis to increase both the valid spectral information and the identification accuracy. In this study, a method based on attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR) with first-order derivative (FD) processing combined with chemometrics is proposed for rapid qualitative and quantitative analysis of Panax ginseng polysaccharides (PGP), Panax notoginseng polysaccharides (PNP), and Panax quinquefolius polysaccharides (PQP). First, ATR-FTIR with FD processing was used to establish the discriminant model combined with principal component analysis (PCA), partial least squares discriminant analysis (PLS-DA) and linear discriminant analysis (LDA). After that, two-dimensional ATR-FTIR based on single-characteristic temperature as external interference (2D-sATR-FTIR) was established using ATR-FTIR with FD processing. Then, ATR-FTIR with FD processing was combined with PLS to establish and optimize the quantitative regression model. Finally, the established discriminant model and 2D-sATR-FTIR successfully distinguished PGP, PNP and PQP, and the optimal PLS regression model had a good prediction ability for the Panax polysaccharide extracts content. This strategy provides an efficient, economical and nondestructive method for the distinction and quantification of PGP, PNP and PQP in a short detection time.

Oligo/polysaccharides from Cyathula officinalis and Achyranthes bidentata: a review of structures and bioactivities.

OBJECTIVES: Oligo-/polysaccharides from Cyathula officinalis Kuan (COPs) and Achyranthes bidentata Blume (ABPs) have attracted researchers' attention in the fields of healthy food supplements and traditional Chinese medicine (Niúxi) due to their multiple bioactivities combined with their nontoxic and highly biocompatible nature. The purpose of this paper was to provide a systematic and comprehensive overview of the extraction, purification, and structural analysis methods, chemical characteristics, biological activities, and structure bioactivity relationship. Furthermore, the possible development trends and perspectives for future research, and traditional uses of Niúxi are also summarized. METHODS: All the information was gathered from a library search and scientific databases. KEY FINDINGS: Although COPs and ABPs are derived from different plants, they have similar structural features in type, structure, and glycosidic linkage patterns and biological activities in vivo and in vitro. However, there are differences in monosaccharide compositions, which can be used as an identification mark. CONCLUSIONS: As traditional Chinese herbal medicine, C. officinalis and A. bidentata have similar pharmacological activities. The COPs and ABP possess wide pharmacological effects such as antitumor, antioxidant, anti-osteoporosis, and anti-inflammatory. Meanwhile, the biological activity and structure-activity relationship of purified COPs and ABPs are less studied, future research should focus on them.

Complete composition analysis of polysaccharides based on HPAEC-PAD coupled with quantitative analysis of multi-components by single marker.

INTRODUCTION: Monosaccharide compositions analysis (MCA) is indispensable for structural characterisations and structure-activity relationships of plant polysaccharides. OBJECTIVES: To develop a concise and direct MCA method, we established a quantitative analysis of the multi-monosaccharaides by single marker (QAMS) by high-performance anion-exchange chromatography with pulsed-amperometric detection (HPAEC-PAD) method. METHODOLOGY: A stable and reproducible HPAEC-PAD method for simultaneous determination of aldoses, ketoses and uronic acids (i.e., l-arabinose, d-xylose, d-ribose, l-rhamnose, d-fucose, d-mannose, d-glucose, d-galactose, d-fructose, d-glucuronic acid and d-galacturonic acid) was established by systematic optimisation of stationary phases, column temperatures and elution programmes. On this basis, the QAMS method was proposed through comprehensive investigations of relative correction factor (RCF) variations under different influencing factors, for example, sample concentrations, flow rates, and column temperatures. RESULTS: Using rhamnose as an internal reference standard, the contents of the other monosaccharide components in polysaccharides from Panax quinquefolium L. and Achyranthes bidentata Bl. samples were simultaneously determined by QAMS, and there was no significant difference between the results from the QAMS and external standard method (t test, P > 0.520). In addition, a MCA fingerprinting of 30 batches of P. quinquefolium polysaccharide was established by HPAEC-PAD, and six common peaks were assigned and determined. CONCLUSIONS: The established HPAEC-PAD-QAMS method was successfully applied to the MCA of polysaccharides from P. quinquefolium and A. bidentata after optimisation of hydrolysis conditions. HPAEC-PAD-QAMS was proposed and established for MCA of plant polysaccharides for the first time.

Revealing the "Yin-Jing" mystery veil of Platycodon grandiflorum by potentiating therapeutic effects and lung-oriented guidance property.

ETHNIC PHARMACOLOGICAL RELEVANCE: "Yin-Jing" medicine (YJM) has been widely used by both ancient and modern Chinese medicine practitioners during long-term clinical practice. However, it remains unclear how to best guide other medicines to the targeted organs in a traditional Chinese medicine (TCM) prescription. Here, in an attempt to explain the scientific connotation of the YJM property (YJMP) attributed to a basic TCM theory, Platycodon grandiflorum (PG) was chosen as a case study to reveal the mystery of YJMP theory. AIM OF THE STUDY: The main purpose of this study is to employ modern chemical and molecular biology methods to confirm the "Yin-Jing" effect of PG, and further clarify its material basis and related possible mechanism. MATERIALS AND METHODS: The ammonia-induced lung injury rat model was utilized to determine the optimal dosage of traditional prescription Hui Yan Zhu Yu decoction (HYZYD) using Wright Giemsa staining, HE staining, Masson staining, and TUNEL analysis. With the same way, PG was confirmed to have potentiating therapeutic effect (PTE) by comparison with HYZYD and [HYZYD-PG]. TMT proteomics was used to reveal the "Yin-Jing" mechanism of action. Western blot assay (WB) was employed for verification of differentially expressed proteins. Additionally, four non-crossing fragmentations (Fr. A-D) were characterized by RPLC/SEC-ELSD and HILIC-ESI--Q-OT-IT-MS techniques. The PTE and guidance property assays were utilized to evaluate "Yin-Jing" functions by a compatible combination of hydroxysafflor yellow A (HYA) using qPCR, FCM, WB, HPLC, high content cell imaging (HCI) and high-resolution live-cell imaging (HRLCI) techniques. RESULTS: The HYZYD-M (medium dose group) significantly improved the lung injury level in a pneumonia model of rats. PG enhanced the therapeutic effect of HYZYD ascribed to Yin-Jing PTE functions. TMT proteomics revealed a category of differentially expressed proteins ascribed to Golgi-ER between HYZYD and [HYZYD-PG]. Fr. C (i.e., saponins) and Fr. D (i.e., lipids) were determined as therapeutic fragmentations via the LPS-induced A549 cell injury model; however, Fr. B (fructooligosaccharides and small Mw fructans) had no therapeutic effect. Further compatibility PTE assays confirmed Fr. B significantly improved efficiency by a combination of HYA. The guidance assays showed Fr. B could significantly increase the uptake and distribution of HYA into lung cells and tissues. HCI assays showed that Fr. B increased uptake of HYA accompanied by significant activation of Golgi-ER. Unlike Fr. B, HRLCI showed that Fr. A, C and D were not only unobvious activations of Golgi-ER but also insignificant facilitation of colocalizations between HYA and Golgi-ER. CONCLUSIONS: Fr. B is believed to be a key YJMP material basis of PG attributed to Yin-Jing PTE with characteristic of lung-oriented guidance property, whereas another abound Fr. C was determined to have synergistic effects rather than Yin-Jing material basis.

TMT-Based Proteomics Reveal the Mechanism of Action of Amygdalin against Rheumatoid Arthritis in a Rat Model through Regulation of Complement and Coagulation Cascades.

The limitations of current medications for treating rheumatoid arthritis (RA) emphasize the urgent need for the development of new drugs. This study aimed to investigate the potential anti-RA mechanism of amygdalin using tandem mass tag (TMT)-based quantitative proteomics technology. First, the anti-RA activity of amygdalin was evaluated in a Complete Freund's adjuvant (CFA)-induced rat model. Then, the roles and importance of proteins in the extracted rat joint tissue were evaluated using TMT-based quantitative proteomics technology. A bioinformatics analysis was used to analyze differentially abundant proteins (DAPs). A proteomics analysis identified 297 DAPs in the amygdalin group compared with the model group, of which 53 upregulated proteins and 51 downregulated proteins showed opposite regulatory trends to the DAPs produced after modeling. According to enrichment analyses of the DAPs, the signaling pathways with a high correlation degree were determined to be the complement and coagulation cascades. Furthermore, western blotting and molecular docking were used to further validate the key node proteins, e.g., complement C1s subcomponent (C1s), component C3 (C3) and kininogen 1 (Kng1). These results suggest that amygdalin may be a promising agent for treating RA by regulating the complement and coagulation cascades.

Insight of "Yin-Jing" medical property ofLigusticum chuanxiong Hort. via pharmacokinetics and tissue distributions by ultra-performance liquid chromatography-triple quadrupole mass spectrometry.

ETHNOPHARMACOLOGICAL RELEVANCE: Ligusticum chuanxiong Hort. (Chuanxiong, LC), as an important traditional Chinese medicine (TCM), can not only be used as a monarch herb but also be used as a classic "Yin-Jing" () medicine in compound prescriptions, e.g., Buyang Huanwu Decoction (BHD). Although LC has the effect of guiding components into the brain in BHD, there is still a lack of scientific evidence on this "Yin-Jing" effects. Herein, we used pharmacokinetics and tissue distributions to investigate "Yin-Jing" effects of LC. To simplify the study, four major constituents in BHD, i.e., Calycosin (CA), astragaloside IV (AI), paeoniflorin (PA), and amygdalin (AM) were combined to form a simple compound (abbreviated as CAPA here) to replace the original BHD in this paper. The Yin-Jing medical property of LC was confirmed by the compatibility of CAPA with LC or its different fractions (Fr. A âˆ¼ Fr. F). AIM OF THE STUDY: To explore the "Yin-Jing" medical property of LC via pharmacokinetics and tissue distributions by ultra-performance liquid chromatography-triple quadrupole mass spectrometry (UPLC-QQQ-MS). MATERIALS AND METHODS: The contents of CA, AI, PA, and AM were simultaneously determined by the established and validated UPLC-QQQ-MS method in different rat tissues and plasma after administration of CAPA with the combination of LC or Fr. A âˆ¼ Fr. F. The pharmacokinetic parameters, e.g., Tmax, Cmax, AUC0-t and MRT0-t, were calculated to evaluate the efficiency of "Yin-Jing". RESULTS: The Cmax and AUC0-t of CA, AI, PA, and AM were remarkably increased in rat brain tissues compared with those of the control group after compatibility of LC. This demonstrated that LC has the Yin-Jing effects on brain tissues. Additionally, Fr. B or Fr. C might be the material basis by specifically studying the distributions of CA, AI, PA, and AM in brain tissue based on mutual compatibility. The effects of Fr. B and Fr. C on distributions of these constituents in other tissues or plasma was also studied to verify the effects of Yin-Jing of LC. The results showed that the same upward trend is found in heart, liver and plasma, but the intensity is insignificant as that in brain tissue. Furthermore, the Cmax and AUC0-t of some analytes in the rat spleen, lung, and kidney were significantly decreased compared with the control group (P < 0.05 or 0.01). CONCLUSIONS: LC has the function of Yin-Jing, especially guiding the components into the brain tissue. Moreover, Fr. B and Fr. C is suggested to be the pharmacodynamic material basis for the effect of Yin-Jing of LC. These finding explained that it was recommended to add LC into some prescriptions for treating cardiovascular and cerebrovascular diseases caused by Qi deficiency and blood stasis. This has laid a certain foundation for the research on the Yin-Jing efficacy of LC to better clarify the theory of TCM and guide the clinical application of Yin-Jing drugs.

Structure of a highly branched galacturonoglucan from fruits of Schisandra chinensis (Turcz.) Baill.

A novel galacturonoglucan, named SCP-1, is isolated and purified from Schisandra chinensis fruits. The structure of SCP-1 is systematically investigated by a combination of monosaccharide compositions, absolute Mw, methylation analysis, partial acid hydrolysis, isoamylase degradations, and nuclear magnetic resonance spectroscopy. The structure of SCP-1 is theoretically described as follows: (i) Glc and GalA in a molar ratio of 17:3; (ii) â†’ 4)-α-Glcp-(1→, →4,6)-α-Glcp-(1→, →3,4,6)-α-Glcp-(1→, α-Glcp-(1→, →4)-α-GalAp-6-OMe-(1→, α-GalAp-6-OMe-(1→, ß-Glcp-(1→, →6-)-ß-Glcp-(1 â†’ and →3,4)-ß-Glcp-(1 â†’ in a molar ratio of 48:5:3:3:10:5:12:5:9; (iii) a repeating unit of →4)-α-Glcp-(1 â†’ as a backbone with branched points at C-3 and C-6, substituted by different types of acidic and neutral side chains to form multiple branches; and (iv) a rigid rod configuration deduced from α value of 1.26 in Mark-Houwink equation ([η] = kMα). Anti-tumor assay investigated the effects of SCP-1 on human HepG2 cancer cell lines in vitro. This is for the first time to report a galacturonoglucan in S. chinensis fruits.

Structure of an unprecedent glucuronoxylogalactoglucomannan from fruit bodies of Auricularia auricula-judae (black woody ear).

An unprecedent glucuronoxylogalactoglucomannan (GXG'G″M), ME-2 (Mw, 2.60 × 105 g/mol; O-acetyl % = 16.7 %), was isolated and purified from water extracts of Auricularia auricula-judae (black woody ear). Firstly, due to much higher O-acetyl contents, we prepared its fully deacetylated products (dME-2; Mw, 2.13 × 105 g/mol) for convenient structure survey. The repeating structure-unit of dME-2 was readily proposed based on Mw determination, monosaccharide compositions, methylation analysis, free-radical degradation and 1/2D NMR spectroscopy. The dME-2 was identified as a highly branched polysaccharide with an average of 10 branches per 10 sugar backbone units. The backbone was only repeating →3)-α-Manp-(1→ residues, substituted at the C-2, C-6 and C-2,6 positions. The side chains included ß-GlcAp-(1→, ß-Xylp-(1→, α-Manp-(1→, α-Galp-(1→ and ß-Glcp-(1→. Secondly, the complex substituted positions of O-acetyl groups in ME-2 were determined to be at C-2, C-4, C-6 and C-4,6 in the backbone and at C-2 and C-2,3 in some side chains. Finally, the anti-inflammatory activity of ME-2 was preliminarily explored on LPS-stimulated THP-1 cells. The above date not only provided the first example for structural studies of GXG'G″M type polysaccharides, but also facilitated development and application of black woody ear polysaccharides as medicinal agents or functional dietary supplements.

Microwave assisted free radical degradation of Schisandra polysaccharides: Optimization, identification and application.

In order to establish structural-fingerprinting of polysaccharides for improvement of quality assessment, a sample preparation method based on microwave assisted free radical degradation (MFRD) of plant polysaccharides was proposed to produce oligosaccharides and small Mw polysaccharides. As a case study of Schisandra chinensis and S. sphenanthera fruit polysaccharides (SCP and SSP), the MFRD condition (i.e., 100 °C, 30 s and 80 W) was confirmed to be optimal. The potential structures of the MFRD products of SCP and SSP were further discussed by combinations of HILIC-ESI--QTOF-MSE and HILIC-ESI--Q-OT-IT-MS/MS. As followed, multivariable statistical analysis shows a clear separation of SCP and the SSP in PCA and OPLS-DA plots based HILIC-ESI--QTOF-MSE data. The VIP plot unveils several key Q-markers (e.g., peaks 3, 8, 9, 10, 15, 25, 26, 28, 29 and 30) with significant differences and stable emergences. Furthermore, a low-polymerization compositional fingerprinting was successfully constructed for SCP and SSP using a high-performance anion-exchange chromatography with pulsed amperometric detection. Compared to the conventional sample preparation methods, the MFRD took only a few thousandth of the time to accomplish degradations of plant polysaccharides. It significantly improves sample preparations and is generally applicable to various polysaccharide samples.

An alternative strategy based on ultra-high-performance supercritical fluid chromatography for full monosaccharide compositional analysis of polysaccharides in Schisandra chinensis fruits.

Due to green and environment-friendly characteristics, ultra-high-performance supercritical fluid chromatography has been widely used in analytical fields in recent years, but until now few reports are available for monosaccharide compositional analysis of macromolecule polysaccharides. In this study, an ultra-high-performance supercritical fluid chromatography technology with an unusual binary modifier is used to determine the monosaccharide compositions of natural polysaccharides. Each carbohydrate herein is simultaneously labeled as 1-pheny-3-methyl-5-pyrazolone and acetyl-derivative via pre-column derivatizations aiming to increase UV absorption sensitivity and decrease water solubility. Ten common monosaccharides are fully separated and detected on ultra-high-performance supercritical fluid chromatography combined with a photo-diode array detector by systematic optimization of multiple relevant parameters, for example, column stationary phases, organic modifiers, additives, flow rates, and so on. Compared with carbon dioxide as a mobile phase, the addition of a binary modifier increases the resolution of analytes. Additionally, this method has the advantages of small consumption of organic solvent, safety, and being environmental-friendly. It has been successfully applied for full monosaccharide compositional analysis of heteropolysaccharides from Schisandra chinensis fruits. To sum up, a new alternative approach is provided for monosaccharide compositional analysis of natural polysaccharides.

Direct acetylation for full analysis of polysaccharides in edible plants and fungi using reverse phase liquid chromatography-multiple reaction monitoring mass spectrometry.

It is vitally important to characterize polysaccharides by monosaccharide composition method. In this study, a direct acetylation strategy combined with reversed-phase liquid chromatography electrospray tandem multiple reaction monitoring mass spectrometry (RPLC-ESI-MRM-MS) was developed for simultaneous determination of 8 aldoses (Glc, Gal, Man, Ara, Xyl, Rib, Rha and Fuc), a ketose (Fru), 2 alditols (Glc-ol and Man-ol) and 2 uronic acids (GlcA and GalA) on a high-pressure resistant reversed-phase column. Employing 1-MeIm as catalyst for direct acetylation, even though no DMSO was used to inhibit the transformation of configurations, each carbohydrate still produced a single chromatographic peak in RPLC conditions due to the ɑ- and ß- isomers merged together. Except for Fru and Man, all the other 11 carbohydrates were base-line separated in a 1.7 µm CYANO column. Therefore, correction factor method is further proposed to perfectly solve co-elution problem of Fru and Man because of occurrence of a specific Q3 ion for aldoses rather than ketose. The result was verified on a 1.7 µm Fluoro-Phenyl column with a full separation of Fru and Man. Herein, the established direct acetylation as followed RPLC-ESI-MRM-MS method was successfully applied for compositional analysis of complex polysaccharides from edible plants and fungi.

A nondestructive solution to quantify monosaccharides by ATR-FTIR and multivariate regressions: A case study of Atractylodes polysaccharides.

The quality evaluation of nature polysaccharides is a tough nut to crack because of its high Mw distributions and larger polarity property. It is well-known that infrared spectroscopy and multiple regression modeling have been used for quantitative examinations in multiple fields, but it has not been applied to the compositional analysis of polysaccharides. In this study, attenuated total reflectance-fourier transform infrared spectroscopy is used to simultaneously quantify aldoses, ketose and uronic acids in Atractylodes polysaccharides by a combination of multivariate regressions. After experience of different data processing pretreatments, the resulting spectrum contains maximum amount of information of monosaccharide contents in Atractylodes polysaccharides. In this case, different smoothing points, derivatives, SNV and MSC are used in the pre-modeling spectrum processing and VIP screening is used to reduce the number of variables to simplify the calculation of the model. All the most optimal prediction models have both good prediction ability (R2 ≥ 0.9 and RPD > 3) and no over fitting (RMSEP/RMSEC < 3). This strategy has opened a new possibility for the nondestructive determination of complex monosaccharide compositions of natural polysaccharides in a short detection time, low equipment requirement and high experimental safety.

Effects of various processed methods on chemical constituents and anti-hepatitis activity of Bupleurum scorzonerifolium Willd. by metabonomics and molecular docking.

In this study, a plant metabonomics technique, utilizing ultra-high performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight mass spectrometry (UPLC-ESI--QTOF), was used to clarify the differences of various processed Bupleurum scorzonerifolium Willd. Principal component analysis (PCA) and orthogonal partial least squares discriminant analysis (OPLS-DA) were used to determine the differences in raw and different processed B. scorzonerifolium samples, including vinegar stir-fry, wine stir-fry, honey stir-fry, honey bran stir-fry and charcoal stir-fry. Thus, 39 significant compounds, e.g., saponins, free fatty acids, fatty acid pentitol glycosides, organic acid and linalool glycosides, were clearly or tentatively identified by UPLC-ESI--QTOF-MS/MS fragmentation pathways and by comparison with available reference standards. Most importantly, fatty acid pentitol glycosides were discovered and identified in B. scorzonerifolium for the first time. Furthermore, a HepG2 hepatitis model induced by TNF-α was used to measure the anti-hepatitis effect of raw and processed B. scorzonerifolium in vitro. Molecular docking was used to understand the interaction of key Q-markers with the active sites of the target protein. The results show that the UPLC-QTOF-MS metabolomics coupled with molecular docking is a powerful tool to quickly identify quality control characteristics of B. scorzonerifolium and its products.

Exploring the effects of different processing techniques on the composition and biological activity of Platycodon grandiflorus (Jacq.) A.DC. by metabonomics and pharmacologic design.

ETHNOPHARMACOLOGICAL RELEVANCE: Platycodon grandiflorus (Jacq.) A.DC. (PG) is a common natural medicine with a history of thousands of years. The processing products were mainly recorded as raw, honey-processed, wine-fried, yellow-fried, and bran-fried PG, which were respectively used for different clinical purposes. Therefore, it is necessary to study the chemical composition and pharmacological activity of PG after processing. AIM OF THE STUDY: To explore the effects of different processing methods on the composition and biological activity of PG using metabonomics and pharmacologic design. MATERIALS AND METHODS: UPLC-QTOF-MS combined with multivariate statistical analysis was used to identify different metabolites before and after the processing of PG. Network pharmacology was used to construct the metabolite-target-disease network. CCK-8 assay, flow cytometry, and western blotting were used to detect cell viability, apoptosis, and the expression of related proteins, respectively. RESULT: A total of 43 differentially expressed metabolites (VIP >10) were detected and identified in the analyzed groups. Based on their chemical nature, these metabolites were divided into five categories, namely, saccharolipids, flavonoid glycosides, alkynes, saponins, and lipids (including fatty acids, phospholipids, fatty aldehydes, and sterols). The content of lipids in the five processed groups (CH, FC, JZ, MZI, and MZG) was found to be higher than that in raw PG. In particular, the processing approaches explored herein increased the contents of many phospholipids, such as, glycerophosphoinositols, phosphatidic acids, and lysophosphatidyle·thanolamines. The 8 metabolites were found by venn diagram to distinguish different processed products (metabolites 2, 6, 19, 20, 21, 26, 28, and 38). The results of network pharmacology analysis showed that the primary anti-cancer targets of 43 metabolites of PG processing products are PIK3CA, Akt, and STAT3, and based on CCK-8 assay, MZI has a significant killing effect on A549 cells, compared to other processing techniques. Moreover, flow cytometry analysis showed that the cells treated with MZI exhibit significantly increased cell apoptosis, and that the effect is dose-dependent. Finally, the western blots performed herein demonstrated that the MZI effectively inhibits the expression of p-Akt and p-STAT3, which is consistent with the network pharmacology results. CONCLUSION: Depending on the processing technique, the contents of 43 different metabolites in PG were varied significantly. Specifically, the contents of phospholipids and fatty acids increase, whereas the contents of large Mw saponins decrease. Compared to the other investigated processing methods, MZI increases the potential of PG in inducing cell apoptosis and inhibiting cell proliferation by affecting the Akt and STAT3 signaling pathways. The increased levels of 3-O-ß-glucopyranosyl polygalacic acid and platycoside F after honey-frying confirm these results.

Structure and immunological activity of an arabinan-rich acidic polysaccharide from Atractylodes lancea (Thunb.) DC.

An arabinan-rich acidic polysaccharide, named ALP4-2 ([α]20 D = +197.8 (c 1.0 mg/mL, H2O); and Mw = 5.59 × 103 g/mol), was obtained from Atractylodes lancea (Thunb.) DC. ALP4-2 is mainly comprised of Ara along with a small amount of GalA, Gal, Rha, Glc and Xyl. The structure was decorated by glycosidic linkages of α-Araf-(1→, →3)-α-Araf-(1→, →5)-α-Araf-(1→, →3,5)-α-Araf-(1→, →2,4)-α-Rhap-(1→, α-GalAp-(1→, →4)-α-GalAp-6-OMe-(1→, →4)-α-GalAp-6-OMe and ß-Galp-(1→ with a ratio of 6:1:7:5:5:1:7:1:4. The structure, configuration and microstructure of ALP4-2 was proposed by comprehensive considerations of results from SEC-MALLS-RID, SEC-HRMS, GC-MS, and 1D/2D NMR spectroscopy. Except for a high methyl ester in full pectin regions, an abundant arabinan moiety is observed in ALP4-2 with highly complex and branched characteristics. The immunoactivity displayed that ALP4-2 can significantly promote phagocytosis of macrophage without cytotoxicity, and stimulate nitric oxide and cytokines (TNF-α, IL-6 and IL-10) release on RAW 264.7.

A novel LC-MS/MS method for complete composition analysis of polysaccharides by aldononitrile acetate and multiple reaction monitoring.

Carbohydrate analysis has always been a challenging task due to the occurrence of high polarity and multiple isomers. Aldoses are commonly analyzed by gas liquid chromatography (GLC) following aldononitrile acetate derivatization (AND). However, the GLC technique cannot be applied for the simultaneous determination of aldoses, ketoses, and uronic acids. In this study, a new method based on the combination of liquid chromatography-tandem mass spectrometry (LC-MS/MS) and AND is developed for the complete characterization of monosaccharide composition (i.e., aldoses, ketoses, alditols, amino sugars, and uronic acids) in plant-derived polysaccharides. In addition to discussing the possible byproducts, the study optimizes the multiple reaction monitoring (MRM) parameters and LC conditions. The final separation of 17 carbohydrates is performed on a BEH Shield RP18 column (150 mm × 2.1 mm, 1.7 µm) within 25 min, without using any buffer salt. Notably, the complex polysaccharides extracted from Ligusticum chuanxiong, Platycodon grandiflorum, Cyathula officinalis Kuan, Juglans mandshurica Maxim, and Aralia elata (Miq.). Seem bud can be successfully characterized using the developed method. Overall, the results demonstrated that the newly established LC-MS/MS MRM method is more effective and powerful than the GLC-based methods reported previously, and it is more suitable for the analysis of highly complex natural polysaccharides, including complex pectins, fructosans, and glycoproteins.

Low-polymerization compositional fingerprinting for characterization of Schisandra polysaccharides by hydrophilic interaction liquid chromatography-electrospray mass spectrometry.

A hydrophilic interaction liquid chromatography-negative electrospray-mass spectrometry (HILIC-ESI--MS) coupled with microwave assisted mild acid (MAMA) depolymerization is proposed here for unusual discrimination and characterization of plant polysaccharides: a case study of fruit polysaccharides in Schisandra chinensis and S. sphenanthera (SCP and SSP). The optimized MAMA hydrolysis procedure was proposed for sample preparations of low-polymerization saccharides (Mw < 5000 Da) released in SCP and SSP. In addition, HILIC-MS/MS was employed for elucidation of isomeric glycosidic linkages in terms of 18O labelling. The MAMA hydrolysates showed that the amount of neutral →(4Hex1)n→ moiety is confirmed to be more bigger than that of acidic →(4HexA1)n → in SCP, whereas the amount of acidic →(4HexA1)n→ moiety seems to be more bigger than that of neutral →(4Hex1)n→ in SSP. The resulting low-polymerization compositional fingerprinting (LCF) showed the performance on rapid visualization of SCP and SSP by HILIC-MIM-MS. Principal components analysis (PCA) and hierarchical cluster analysis (HCA) further unveils several key Q-markers (e.g., m/z 503, 369, 665, 827, 989, 1151 and 735) for rapid discrimination of SCP and SSP. This practical study showed that the LCF with PCA and HCA could effectively reflect structural differences and could rapidly achieve discrimination of SCP and SSP.

Identification and comparison of triterpene saponins in Aralia elata leaves and buds by the energy-resolved MSAll technique on a liquid chromatography/quadrupole time-of-flight mass spectrometry.

In this study, we identify the triterpene saponins (TSs) extracted from the leaves and buds of Aralia elata (Miq.) Seems using ultra-performance liquid chromatography and positive ionization electrospray quadrupole time-of-flight mass spectrometry (UPLC-ESI+-QTOF). The energy-resolved MSAll (erMSall) technique is applied in order to simultaneously collect the diverse precursors attributed to [M+H]+, [M + NH4]+ and [M + Na]+ ions. A practical and effective erMSall workflow is established to rapidly identify and compare the saponins in the analyzed samples. In total, 111 TSs of structure are estimated, including 44 new compounds that had not been identified previously in A. elata. Of the five aglycones detected in the samples, a sapogenin 3ß, 16α, 23-trihydroxyoleana-11,13-dien-28-oic acid (A5) that is identified for the first time in A. elata leaves. Compared to the buds, the leaves number twice as many hederagenin-type (A2) compounds. Although the number of other aglycones does not vary significantly between the buds and the leaves, A5 compounds are exclusively detected in the latter. Moreover, the C-3 sugar chains of TSs in A. elata leaves are mainly neutral (e.g., Hex+Hex, Hex+Hex+Hex and Hex+Hex+Hex+Hex), whereas those of bud TS compounds are primarily acidic (e.g., Pen+HexA, Hex+HexA and Hex+Pen+HexA). Some of the identified TS compounds, e.g., 27, 28, 32, 46, 54, 57, 71 and 105 can be used as indices to evaluate the quality of the plant leaves and buds. Overall, this study is of great significance for the comparative study of triterpenoid saponins in the leaves and buds of Aralia elata.

Discrimination and characterization of Panax polysaccharides by 2D COS-IR spectroscopy with chemometrics.

In this study, a novel two-dimensional correlation infrared spectroscopy (2DCOS-IR) is presented to rapidly characterize and discriminate polysaccharides in Panax ginseng (PGP), P. notoginseng (PNP), and P. quinquefolius (PQP) using attenuated total reflection Fourier transform infrared spectroscopy-based on single-characteristic temperature as the external disturbance (2D-sATR-FTIR). Compared with two existing 2DCOS-IR methods based on gradient heating pathways using KBr pellet (100 min; 2D-KBr-FTIR) and attenuated total reflection (30 min; 2D-gATR-FTIR), the new procedure took an average of just 2 min to finish a sample measurement, which resolved previously tedious and time-consuming dilemmas. It offered advantages in the quality evaluation of natural polysaccharides and featured nondestructive, high-throughput, and high-efficiency characteristics. An intuitive analysis of the 2D-sATR-FTIR demonstrated that PNP was first identified because it had fewer auto-peaks. Posteriorly, PGP and PQP were distinguished according to the ratio of the auto-peaks 6 and 9, with the former greater than 1 and the latter less than 1. Furthermore, characteristic auto-peaks 1, 5, and 6 were unambiguously determined as Quality-markers using PCA and PLS-DA for visualized identifications. LDA was successfully used to establish a predictive model of the PGP, PNP, and PQP based on the positions and intensity of these three characteristic auto-peaks.

Structural characterization of the metabolites of orally ingested hederasaponin B, a natural saponin that is isolated from Acanthopanax senticosus leaves by liquid chromatography-mass spectrometry.

Plant saponins are important natural product with biologically active. However, the metabolism of these compounds has rarely been studied due to their low bioavailability and the complexity of their metabolite structures. In this study, ultra-performance liquid chromatography/Fusion Lumos Orbitrap mass spectrometry was used to analyze the metabolites of hederasaponin B in vivo, and its possible metabolic pathways were proposed. After oral administration of the parent drug, a total of 47 metabolites are identified in rat feces (42), urine (11), and plasma (9) samples. These metabolites resulted from the metabolic processes in phases I and II reactions involved in deglycosylation, hydroxylation, acetylation, oxidation, gluconalciation and glycosylations. Deglycosylation is the main metabolic pathway (accounts for 52.46 % of all metabolites in feces samples). Among the identified metabolites, four were glycosylated (deprotonated precursors at m/z = 1335.7, 1365.7, 1467.9, and 1379.6) with higher molecular weight than the parent drug . These glycosylated compounds account for 11.55 % of the metabolites in rat feces according to the semi-quantitative chromatographic peak areas. To sum up, the results of this study provide a basis for further understanding the metabolism of plant saponins in vivo.