Metabolic profiles of Fufang Xiling Jiedu capsule in rats by ultra-performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry.

Fufang Xiling Jiedu capsule (FXJC), a traditional Chinese medicine that evolved from "Yinqiao Powder", is widely used for the treatment of cold and influenza. However, due to a lack of in vivo metabolism research, the chemical components responsible for the therapeutic effects still remain unclear. Hence, this study aimed to describe the metabolic profiles of the FXJC in rat plasma, urine, and feces. A combined data mining strategy based on ultra-performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry was employed and 201 xenobiotics, including 117 prototype components and 84 metabolites were detected. Phenolic acids, flavonoids, triterpenes, and lignans were prominent ingredients absorbed in vivo, and the major metabolic pathways of the detected metabolites were glucuronidation, sulfation, methylation, and oxidation. This is the first systematic study on the metabolism of the FXJC in vivo, providing valuable information for future studies on the efficacy, toxicity, and mechanism of the FXJC.

Study on quality analysis of different species of Coptidis rhizome based on fingerprint-effect relationship.

INTRODUCTION: The quality evaluation of Coptidis rhizome (CR) is attributed to the origin and processing method, and this strategy of ignoring the bioactive components usually leads to biased quality analysis, which is difficult to indicate the clinical efficacy. OBJECTIVES: In order to evaluate the quality level of different species of CR, we collected 20 batches of CR and investigated the fingerprint-effect relationship. METHODS: High-performance liquid chromatography (HPLC) fingerprints of CR were established, and the fingerprint-effect relationship was explored using cluster analysis, principal component analysis, Pearson correlation analysis, grey relation analysis, and partial least squares regression. RESULTS: We have identified a total of 10 common peaks (1-10) with similarity scores above 0.96. The study on the relationship between spectra and potency further showed that the contents of peaks 8, 9, and 10 are potential key components. And based on a previous study, a method of one measurement and multiple evaluations of CR was established to achieve the goal of simplifying the analytical process and reducing costs. CONCLUSION: Through a combination of fingerprint analysis, antioxidant activity evaluation, fingerprint-efficacy relationship analysis, and simultaneous quantification of multiple components, a CR quality control index and method have been selected and established, which can also provide a more comprehensive quality evaluation for traditional Chinese medicine.

Quality study of animal-derived traditional Chinese medicinal materials based on spectral technology: Calculus bovis as a case.

INTRODUCTION: Calculus bovis (C. bovis) is a typical traditional Chinese medicine (TCM) derived from animals, which has a remarkable curative effect and high price. OBJECTIVES: Rapid identification of C. bovis from different types was realized based on spectral technology, and a rapid quantitative analysis method for the main quality control indicator bilirubin was established. METHODS: We conducted a supervised and unsupervised pattern recognition study on 44 batches of different types of C. bovis by five spectral pretreatment methods. Three variable selection methods were used to extract the essential information, and the partial least squares regression (PLSR) quantitative model of bilirubin by near-infrared (NIR) spectroscopy was constructed. RESULTS: The partial least squares discriminant analysis (PLS-DA) model could achieve 100% accuracy in identifying different types of C. bovis. The R2 of the NIR quantitative model was 0.979, which is close to 1, and the root mean square error of calibration (RMSEC) was 2.3515, indicating the good prediction ability of the model. CONCLUSION: The study was carried out to further improve the basic data of quality control of C. bovis and help the high-quality development of TCM derived from animals.

Separation and purification of quinolyridine alkaloids from seeds of Thermopsis lanceolata R. Br. by conventional and pH-zone-refining counter-current chromatography.

In this work, the preparative separation of quinolyridine alkaloids from seeds of T. lanceolata by conventional and pH-zone-refining counter-current chromatography. Traditional counter-current chromatography separation was performed by a flow-rate changing strategy with a solvent system of ethyl acetate-n-butanol-water (1:9:10, v/v) and 200 mg sample loading. Meanwhile, the pH-zone-refining mode was adopted for separating 2.0 g crude alkaloid extracts with the chloroform-methanol-water (4:3:3, v/v) solvent system using the stationary and mobile phases of 40 mM hydrochloric acid and 10 mM triethylamine. Finally, six compounds, including N-formylcytisine (two conformers) (1), N-acetycytisine (two conformers) (2), (-)-cytisine (3), 13-ß-hydroxylthermopsine (4), N-methylcytisine (5), and thermopsine (6) were successfully obtained in the two counter-current chromatography modes with the purities over 96.5%. Moreover, we adopted nuclear magnetic resonance and mass spectrometry for structural characterization. Based on the obtained findings, the pH-zone-refining mode was the efficient method to separate quinolyridine alkaloids relative to the traditional mode.

A Novel Variable Selection Method Based on Binning-Normalized Mutual Information for Multivariate Calibration.

Variable (wavelength) selection is essential in the multivariate analysis of near-infrared spectra to improve model performance and provide a more straightforward interpretation. This paper proposed a new variable selection method named binning-normalized mutual information (B-NMI) based on information entropy theory. "Data binning" was applied to reduce the effects of minor measurement errors and increase the features of near-infrared spectra. "Normalized mutual information" was employed to calculate the correlation between each wavelength and the reference values. The performance of B-NMI was evaluated by two experimental datasets (ideal ternary solvent mixture dataset, fluidized bed granulation dataset) and two public datasets (gasoline octane dataset, corn protein dataset). Compared with classic methods of backward and interval PLS (BIPLS), variable importance projection (VIP), correlation coefficient (CC), uninformative variables elimination (UVE), and competitive adaptive reweighted sampling (CARS), B-NMI not only selected the most featured wavelengths from the spectra of complex real-world samples but also improved the stability and robustness of variable selection results.

An efficient high-speed counter-current chromatography method for the preparative separation of potential antioxidants from Paeonia lactiflora Pall. combination of in vitro evaluation and molecular docking.

Paeonia lactiflora Pall., one of the most famous classical herbal medicine, has been used to treat diseases for over 1200 years. In this research, the functional ingredients were purified by online-switch two-dimensional high-speed counter-current chromatography combined with inner-recycling and continuous injection mode. The antioxidant activity was evaluated by investigating the 2,2'-azobis (2-amidinopropane) dihydrochloride-induced oxidant damage in vitro and confirmed through molecular docking. n-Butanol/ethyl acetate/water (2:3:5, v/v) solvent system was used for the first-dimensional separation and optimized the sample loading. Two pure compounds and a polyphenol-enriched fraction were separated. The polyphenol-enriched fraction was separated with a solvent system n-hexane/ethyl acetate/methanol/water (2:8:4:6, v/v) with continuous injection mode. Five compounds were successfully separated, including gallic acid (1), methyl gallate (2), albiflorin (3), paeoniflorin (4), and ethyl gallate (5). Their structures were identified by mass spectrometry and NMR spectroscopy. The results from the antioxidant effect showed that albiflorin had stronger antioxidant activity. Molecular docking results indicated that the affinity energy of the identified compounds ranged from -3.79 to -8.22 kcal/mol and albiflorin showed the lowest affinity energy. Overall, all those findings suggested that the strong antioxidant capacity of albiflorin can be potentially used for the treatment of diseases caused by oxidation.

Anti-pandemic influenza A (H1N1) virus potential of Xilingjiedu capsule in vitro.

This study is aimed to investigate the effect of Xilingjiedu capsule (XLC), one of a preparation of traditional Chinese medicine, on influenza A (H1N1) virus as well as its preliminary mechanism. The median cell mortality (TC50) to A549 cells and half effective inhibition concentration (IC50) of influenza A (H1N1) virus of XLC were determined by MTT assay. Reed-Muench method was used to calculated the 50% tissue culture infective dose (TCID50) of H1N1 virus to A549 cells. In mechanism research, the mRNA expression levels of MyD88, TLR4, TLR7 and TRAF6 and the protein expression level of MyD88 were detected by using RT-PCR and Western blot, respectively. The results suggested that XLC showed good anti influenza A (H1N1) virus activity. The antiviral mechanism of XLC was related to the Toll-like signaling pathway. It could drown regulate the mRNA expression level of MyD88 and TLR4 and the protein level of MyD88. This research provides reference for the application of XLC in anti influenza virus.

Qianliexin capsule exerts anti-inflammatory activity in chronic non-bacterial prostatitis and benign prostatic hyperplasia via NF-κB and inflammasome.

Qianliexin capsule (QLX) is a standardized traditional Chinese herbal preparation that has long been used to treat chronic non-bacterial prostatitis (CNP) and benign prostatic hyperplasia (BPH). This study investigated the anti-inflammatory activity of QLX in improving lower urinary tract symptoms (LUTS) associated with CNP and BPH. Rat models of CNP and BPH were induced by oestradiol or testosterone (hormonal imbalance) or chemical inflammation (carrageenan). QLX significantly relieved LUTS in CNP and BPH rat model by reducing prostate enlargement, epithelial thickness, pain response time, urine volume and bleeding time, and by improving prostatic blood flow. The expression of the pro-inflammatory cytokines interleukin (IL)-1ß and tumour necrosis factor (TNF)-α, the pro-inflammatory transcription factor nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), and inflammasome components (NLRP3, caspase-1 and ASC) in CNP and BPH tissues was reduced by QLX addition. QLX treatment was followed by reduced cellular malondialdehyde and increased superoxide dismutase, catalase and glutathione peroxidase activity, consistent with antioxidant activity. Increases in Beclin-1 expression and the LC3II/I ratio following QLX treatment indicated that autophagy had been induced. QLX relieved LUTS in CNP and BPH rat models by inhibiting inflammation. The underlying mechanisms included inhibition of inflammasome activation, NF-κB activation, oxidant stress and autophagy.

A metabolomics study of Qianliexin capsule treatment of benign prostatic hyperplasia induced by testosterone propionate in the rat model.

A metabolomics investigation of the treatment effect of Qianliexin (QLX) capsules was conducted on rats with benign prostatic hyperplasia (BPH) induced by testosterone propionate. Establishment of the BPH model was confirmed using the prostatic index. Hematoxylin and eosin (HE) staining for TGF-ß, EGFR, collagen, IL-1 ß, TNF-α was performed and changes in urine volume were measured. Urine and serum samples were collected from three groups, including a control group, a BPH model group and a QLX-treated group and subjected to metabolomics profiling based on ultrahigh-performance liquid chromatography-mass spectrometry. Pharmacodynamics analysis showed that the QLX group had significantly lower histopathological damage, fibrosis damage, and inflammation and higher urine output compared with the model group. Twenty-two potential biomarkers were identified in urine samples and 23 metabolites were identified in plasma samples. Alterations in metabolic patterns were evident in all sample types. The treatment effects of QLX appear to involve various metabolic pathways including lipid metabolism, fatty acid metabolism and purine generation and significantly reduced the pathological symptoms and related biochemical indicators of BPH and improved the level of potential marker metabolites. This comprehensive study suggested that differential markers provided insights into the metabolic pathways involved in BPH and the treatment effects of QLX.

The rapid identification of chemical constituents in Fufang Xiling Jiedu capsule, a modern Chinese medicine, by ultra-performance liquid chromatography coupled with quadrupole-time-of-flight tandem mass spectrometry and data mining strategy.

Fufang Xiling Jiedu capsule is an effective Chinese medicine widely used for the treatment of cold and influenza. However, its chemical constituents had not been determined, which entailed a huge obstacle to further pharmacological studies, clinical-safe medication administration, and quality evaluation. To identify the chemical constituents in Fufang Xiling Jiedu capsule, an efficient and systematic approach using ultra-high-performance liquid chromatography coupled with a quadrupole time-of-flight mass spectrometry in conjunction with a data mining strategy was adopted in this study. As a result, 145 compounds were qualitatively identified, including 26 phenolic acids, 46 flavonoids, 39 triterpenes, and 34 other compounds, among which 6 were potentially new and 144 were being reported from Fufang Xiling Jiedu capsule for the first time. This research not only provides useful information for quality control of Fufang Xiling Jiedu capsule and its involved single herbs but also serve as basis data for further study of Fufang Xiling Jiedu capsule in vivo. Moreover, it provides a reference for the characterization of the chemical constituents of other Chinese medicine preparations.

[A new aurone with anti-inflammatory activity from Cleistocalyx operculatus flower buds].

A new compound(Z)-6-hydroxy-4-methoxy-5,7-dimethylaurone was isolated from Cleistocalyx operculatus flower buds. Its structure was identified by spectroscopic data including MS, ¹H-NMR, ¹³C-NMR HSQC and HMBC. A known compound, 2',4'-dihydroxy-6'-methoxy-3'5'-dimethylchalcone (DMC), was also isolated and identified,and used as material to synthesize (Z)-6-hydroxy-4-methoxy-5,7-dimethylaurone.Anti-inflammatory activities of the two compounds were tested in vitro. The results showed that (Z)-6-hydroxy-4-methoxy-5,7-dimethylaurone possesses much stronger PGE2 inhibitory activity (IC50 6.12 nmol·L⁻¹) than the positive control ibuprofen （68.66 nmol·L⁻¹ï¼‰.

Discovery of acylguanidine oseltamivir carboxylate derivatives as potent neuraminidase inhibitors.

In search of novel anti-influenza agents with higher potency, a series of acylguanidine oseltamivir carboxylate analogues were synthesized and evaluated against influenza viruses (H1N1 and H3N2) in vitro. The representative compounds with strong inhibitory activities (IC50 <40nM) against neuraminidase (NA) were further tested against the NA from oseltamivir-resistant strain (H259Y). Among them, compounds 9 and 17 were potent NA inhibitors that exhibited a 5 and 11-fold increase in activity comparing with oseltamivir carboxylate (2, OC) against the H259Y mutant, respectively. Furthermore, the effect against influenza virus H259Y mutant (H1N1) replication and cytotoxicity assays indicated that compounds 9 and 17 exhibited a 20 and 6-fold increase than the parent compound 2, and had no obvious cytotoxicity in vitro. Moreover, the molecular docking studies revealed that the docking modes of compounds 9 and 17 were different from that of oseltamivir, and the new hydrogen bonds and hydrophobic interaction were formed in this case. This work provided unique insights in the discovery of potent inhibitors against NAs from wild-type and oseltamivir-resistant strains.

[Simultaneous determination of eleven components in Ginkgo biloba leaves by high performance liquid chromatography method].

To study Ginkgo biloba leaves in different producing area, we establish an HPLC method for the simultaneously determination of seven flavonoids glycosides and four biflavonoids in G. biloba leaves. The analysis was performed on an Agilent ZORBAX SB-C18 column(4.6 mm×250 mm, 5 µm) wich acetonitrile, and 0.4% phosphoric acid as mobile phase at flow rate of 1 mL•min⁻¹ in a gradient edution, and the detection was carried out at 254 nm.The calibration curves of the seven flavonoids glycosides and four biflavonoids had a good linearitiy with good recoveries. The established HPLC method is simple, rapid, accurate, reliable, and sensitive, and can be applied to the identification and quality control of G. biloba leaves.

[Simultaneous determination of thirteen components in Tong'an injection by LC-MS/MS].

The aim of this study was to develop a simple, sensitive ultra performance liquid chromatography mass spectrometry (UPLC-MS/MS) method for the determination of syringaresinol, N-trans-feruloyltyramine, chelerythrine chloride, sinomenine, coptisine chloride, sanguinarine, chelidonine, magnolflorine, allocryptopine, protopine, farrerol, stylopine and dihydrosanguin-arine in Tong'an injection (TAI), which could be used for the quality control of TAI. The UPLC analysis was performed on Agilent Zorbax SB-Aq column (2.1 mm×150 mm,3.5 µm), with 0.1% formic acid solution (A) -acetonitrile (B) as the mobile phase for gradient elution (0.01-2 min, 5%B; 2-8 min, 5%-30%B; 8-11 min, 30%-95%B; 11-13 min, 95%B; 13-13.1 min, 95%-5%B; 13.1-14 min, 5%B). The flow rate was 0.5 mL•min⁻¹, and the column temperature was 25 ℃; multiple reaction monitoring (MRM) was performed in electrospray ion source positive ion mode for quantitative determination. The calibration curves for the above thirteen compounds showed good linear relationship in corresponding mass concentration range (r>0.999 0). The average recovery rate of the compounds ranged from 95.70% to 104.8%, with RSD of less than 1.9%. The contents of thirteen active components in 10 batches of TAI were 0.021 2-0.029 0, 0.001 7-0.002 3, 0.000 9-0.001 3, 5.952-6.205 2, 0.195 4-0.240 5, 0.002 0-0.002 9, 0.693-0.798 2, 0.069 3-0.078 2, 0.089 29-0.102 9, 0.386 5-0.420 1, 0.014 3-0.015 9, 0.755 3-0.842 1, and 0.008 2-0.011 2 g•L⁻¹ respectively. Methodology validation proved that this method was simple, rapid, sensitive and accurate, which can be used to provide reference for the comprehensive evaluation of TAI quality. The determination results of 10 batches of TAI showed the content of each batch had no significant difference. The results may provide a basis for the quality control of TAI.

Potential lipase inhibitors from Chinese medicinal herbs.

CONTEXT: Obesity has become a major health concern, and it places both personal and economic burdens on the world's population. Traditional Chinese medicinal herbs are rich source of lead compounds and are possible drug candidates, which may be used to treat this condition. OBJECTIVE: This study screened potent pancreatic lipase inhibitors found in traditional Chinese medicinal herbs for ability to treat obesity. MATERIALS AND METHODS: A porcine pancreatic lipase inhibition assay was established, and the inhibitory activity of 35 traditional Chinese medicinal herbs was evaluated at a concentration of 200 µg/mL. Two elutions of herbal extracts with strong lipase inhibitory activity were further fractionated by preparative high-performance liquid chromatography into 22 sub-fractions each, and these sub-fractions were tested for anti-lipase activity. Sub-fractions, which exhibited strong lipase inhibitory activity, were continuously fractionated into individual compounds. Two active compounds with potent anti-lipase activity were finally isolated and identified from two traditional Chinese medicinal herbs, respectively. RESULTS: Among 35 traditional Chinese medicinal herbs, the 95% ethanol elutions of Panax notoginseng (Burk.) F.H. Chen (Araliaceae) and Magnolia officinalis Rehd. et Wils (Magnoliaceae) showed strong anti-lipase activity. Two compounds, including 20(S)-ginsenoside Rg3 and honokiol were identified using bioactivity-guided isolation with IC50 = 33.7 and 59.4 µg/mL, respectively. CONCLUSION: 20(S)-ginsenoside Rg3 and honokiol might be suitable candidates for the treatment of obesity.

[Effect of rat intestinal flora in vitro on metabolites of acteoside].

Acteoside was used for anaerobic incubation with rat intestinal flora in vitro. HPLC was used to detect the changes of acteoside at different incubation time points and HPLC-Q-TOF-MS was used to identify the metabolites of acteoside. The results showed that acteoside could be metabolized by rat intestinal flora in vitro and the metabolites were 3,4-dihydroxyphenyl acid, caffeic acid and 3-(3'-hydroxyphenyl) propionic acid.

[Effect of rat intestinal flora on in vitro metabolic transformation of pumiloside].

To study the metabolic transformation of pumiloside by rat intestinal flora in vitro and identify its metabolites. Pumiloside was incubated in the rat intestinal flora in vitro. HPLC was used to monitor the metabolic process, and HPLC-Q-TOF-MS was used to identify the structures of biotransformation products. In vitro, pumiloside was easily metabolized by rat intestinal flora, and with the prolongation of metabolic time, pumiloside was transformed into several metabolites. Three metabolites were initially identified in this experiment. The study indicated that pumiloside could be extensively metabolized in the rat intestinal flora in vitro.

[Research on Chinese medicine pairs effects of Panax notoginseng and Bletilla striata before and after compatibility on contents of notoginsenosides].

To discuss the synergistic mechanism of compatible use of two medicinal herbs,Panax notoginseng and Bletilla striata, an HPLC was established to determine two ginseng saponins (20S)-ginseng saponin Rg3 and ginseng saponin Rh4 contained in single decoction of Panax notoginseng as well as in compound decoction of Panax notoginseng and Bletillastriata in different compatibility ratio (1∶0.5, 1∶1, 1∶2), followed by analyzing the impact of amount of notoginsenosides after compatibility. As a result, compared with the single decoction of Panax notoginseng, the contents of ginseng saponin Rg3 and ginseng saponin Rh4 in the compound decoction of Panax notoginseng and Bletillastriata were on the rise as the increasement of the amount of Bletillastriata. The contents of the notoginsengsaponin R1, ginseng saponin Rg1 and ginseng saponin Rb1 of Panax notoginseng single decoction were significantly decreased after compatibility. Therefore, after compatibility, it was more easy to produce (20S)-ginseng saponin Rg3 and ginseng saponin Rh4.This study can extend to a method of preparation of (20S)-ginseng saponin Rg3 and ginseng saponin Rh4. Furthermore, after compatibility, two ginseng saponins which had lipase inhibitory effect were both increased significantly, indicating that the compatibility of these two herb medicines may have effect on losing weight.

Anti-hyperuricemic and nephroprotective effects of rhein in hyperuricemic mice.

Hyperuricemia has been considered to be a key risk factor for kidney disease. The formation of uric acid crystals in the kidney further stimulates an intensive inflammatory response. Rhein possesses various pharmacological activities, including anti-inflammatory, antioxidative, antitumor, purgative effects, and so on. To our knowledge, no previous work has been reported about the therapeutic effect of rhein on urate nephropathy. In this study, a model of hyperuricemia and nephropathy induced by adenine and ethambutol in mice was established. Meanwhile, the potential beneficial effects and mechanisms of rhein on hyperuricemia and nephropathy were also investigated. The results demonstrated that rhein significantly decreased the serum uric acid level by inhibiting the xanthine oxidase activity and increasing the excretion of urinary uric acid. In addition, rhein also markedly improved kidney damage related to hyperuricemia. Further investigation indicated that rhein improved the symptoms of nephropathy through decreasing the production of proinflammatory cytokines, including interleukin 1ß, prostaglandin E2, and tumor necrosis factor-α and inhibiting the expression of transforming growth factor-ß1. The present study suggests that rhein may have a considerable potential for development as an anti-hyperuricemic and nephroprotective agent for clinical application.

[Study on chemical constituents from seed of Oroxylum indicum].

Oroxylum indicum was a traditional Chinese medicine. In order to study the chemical constituents from the seed of O. indicum, the chemical constituents of 80% methanol extract of seeds of O. indicum were subjected to chromatography on silica gel, Sephadex LH-20, and preparative HPLC, leading to the isolation of eleven compounds. The structures were identified by various spectroscopic data including ESI-MS, 1H-NMR and 13C-NMR data as oroxin B (1), chrysin (2), baicalein (3), neglectein (4), quercetin-3-O-ß-D-galactopy ranoside (5), quercetin-7-O-ß-D-glucopyranoside (6), 2α,3ß-dihydroxylluPeol (7), lupeol (8), rengyol (9), ß-sitostero (10), and stigmasterol (11). Among them, compound 5 were firstly obtained from O. indicum.