Gut dysbiosis impairs intestinal renewal and lipid absorption in Scarb2 deficiency-associated neurodegeneration.

Scavenger receptor class B, member 2 (SCARB2) is linked to Gaucher disease (GD) and Parkinson's disease (PD). Deficiency in the SCARB2 gene causes progressive myoclonus epilepsy (PME), a rare group of inherited neurodegenerative diseases characterized by myoclonus. We found that Scarb2 deficiency in mice leads to age-dependent dietary lipid malabsorption, accompanied with vitamin E deficiency. Our investigation revealed that Scarb2 deficiency is associated with gut dysbiosis and an altered bile acid pool, leading to hyperactivation of FXR in intestine. Hyperactivation of FXR impairs epithelium renewal and lipid absorption. Patients with SCARB2 mutations have a severe reduction in their vitamin E levels and cannot absorb dietary vitamin E. Finally, inhibiting FXR or supplementing vitamin E ameliorates the neuromotor impairment and neuropathy in Scarb2 knockout mice. These data indicate that gastrointestinal dysfunction is associated with SCARB2 deficiency-related neurodegeneration, and SCARB2-associated neurodegeneration can be improved by addressing the nutrition deficits and gastrointestinal issues.

Genome-Wide Identification, Classification, and Expression Analyses of the CsDGAT Gene Family in Cannabis sativa L. and Their Response to Cold Treatment.

Hempseed is a nutrient-rich natural resource, and high levels of hempseed oil accumulate within hemp seeds, consisting primarily of different triglycerides. Members of the diacylglycerol acyltransferase (DGAT) enzyme family play critical roles in catalyzing triacylglycerol biosynthesis in plants, often governing the rate-limiting step in this process. As such, this study was designed to characterize the Cannabis sativa DGAT (CsDGAT) gene family in detail. Genomic analyses of the C. sativa revealed 10 candidate DGAT genes that were classified into four families (DGAT1, DGAT2, DGAT3, WS/DGAT) based on the features of different isoforms. Members of the CsDGAT family were found to be associated with large numbers of cis-acting promoter elements, including plant response elements, plant hormone response elements, light response elements, and stress response elements, suggesting roles for these genes in key processes such as development, environmental adaptation, and abiotic stress responses. Profiling of these genes in various tissues and varieties revealed varying spatial patterns of CsDGAT expression dynamics and differences in expression among C. sativa varieties, suggesting that the members of this gene family likely play distinct functional regulatory functions CsDGAT genes were upregulated in response to cold stress, and significant differences in the mode of regulation were observed when comparing roots and leaves, indicating that CsDGAT genes may play positive roles as regulators of cold responses in hemp while also playing distinct roles in shaping the responses of different parts of hemp seedlings to cold exposure. These data provide a robust basis for further functional studies of this gene family, supporting future efforts to screen the significance of CsDGAT candidate genes to validate their functions to improve hempseed oil composition.

Simultaneous production of hydrogen-methane and spatial community succession in an anaerobic baffled reactor treating corn starch processing wastewater.

Corn starch processing wastewater (CSPW) is a high-strength organic wastewater and biological treatment is considered as the dominant process. The present work investigated the effects of pH on the bioenergy production and spatial succession of microbial community in an anaerobic baffled reactor (ABR) treating CSPW. The results showed that above 90.5% of COD removal and above 16.6 L d-1 of methane were achieved at the influent pHs of 8.0 and 7.0 under organic loading rate of 4.0 kg COD·m-3·L-1 condition. Further decreasing the influent pH to 6.0 resulted in the COD removal decreased to 89.7%. Besides, 9.2 L d-1 of hydrogen and 13.0 L d-1 of methane were obtained. There was significant difference in the volatile fatty acids profiles during the variation of pH. Illumina Miseq sequencing showed that Clostridium, Ethanoligenens, Megasphaera, Prevotella and Trichococcus with relative abundance of 2.1%∼28.1% were the dominant hydrogen-producing bacteria in C1. Methanogens (Methanothrix and Methanobacterium) dominated in the last three compartments. Function predicted analysis revealed that the abundance of metabolic-related gene families containing carbohydrate, amino acids and energy in the last three compartments were higher than that in C1. A deduced biodegradation model of CSPW in ABR revealed that the anaerobic sludge in C1 mainly produced hydrogen. Microbial population in C3 was responsible for COD removal and methane production. The redundancy analysis revealed that hydrogen production was highly correlated with some hydrogen-producing bacteria in C1, whereas methane production was positively correlated with microbial group in C2∼ C4.

Characterization and comparison of lignin derived from corncob residues to better understand its potential applications.

Lignocellulosic biomass is regarded as an ideal renewable resource for replacing traditional fossil fuels. Corncob residues (CRs) generated after hemicelluloses pre-extraction of corncobs were potential feedstocks to produce lignin-based products. Three lignin fractions, i.e., dioxane lignin (DL), milled-wood lignin (MWL), and enzymatic mild acidolysis lignin (EMAL), were isolated from CRs and characterized by HPLC, GPC, FT-IR, NMR, and TGA to reveal characteristics of CRs lignin to better understand its potential applications. The results showed that the yield of DL, MWL and EMAL were 12.8%, 14.7% and 15.6%, respectively. The molecular weight of DL, MWL and EMAL were 2001, 5391, and 2629 g/mol, respectively. The EMAL and MWL had similar structural features including guaiacyl and syringyl units, ß-O-4' aryl ether, p-coumaric acids, ferulates, and phenylcoumarans, while the structure of DL was relatively simple. It is expected that this work will provide some useful information for the utilization of CRs in view of biorefinery.

Qualitative analysis of the roots of Salvia miltiorrhiza and Salvia yunnanensis based on NIR, UHPLC and LC-MS-MS.

Near infrared spectroscopy (NIR) was applied to discriminate the roots of salvia miltiorhiza Bunge (Danshen for short) and Salvia yunnanensis C. H. Wright (Zidanshen for short) by means of principal component analysis (PCA), improved and simplified K nearest neighbors (IS-KNN). Furthermore, an ultra-high performance liquid chromatographic (UHPLC) coupled with photodiode-array detector was developed for building fingerprints of lipophilic components of Danshen and Zidanshen, respectively. Basing on NIR information, both PCA and IS-KNN method classified the two kinds of Chinese medical herbs with 100% accuracy. The chromatographic fingerprints of the lipophilic components of Danshen and Zidanshen have 10 and 12 common peaks, respectively. Liquid chromatography coupled with mass spectroscopy (LC-MS-MS) was applied to identify these peaks. Among these, three small peaks in the fingerprints of Zidanshen are not found in Danshen, one of which was identified as α-lapachone, and the other two compounds were not yet identified; a small peak after tanshinone IIA in the fingerprints of Danshen was not found in Zidanshen, which was identified as miltirone. The two herbs have 10 common lipophilic components. The similarity between the two reference chromatograms of Zidanshen and Danshen is 0.902, but the mean similaritie between Zidanshen (or Danshen) fingerprints and its own reference chromatogram is 0.973 (or 0.976). The contents of main lipophilic components are significantly lower in Zidanshen than in Danshen (P < 0.01 or P < 0.05). The results indicate that the two Chinese medical materials are not only different in NIR spectra, but also different in species and quantities of lipophilic components. NIR spectra analysis can identify Danshen and Zidanshen rapidly and accurately. UHPLC coupled with MS analysis demonstrates the detail differences between the two herbs both in species and contents of their lipophilic components.

[Analysis of lipophilic components of Salvia miltiorrhiza roots and S. yunnanensis roots by UPLC and LC-MS/MS].

Fingerprints of lipophilic components in the roots of Salvia miltiorrhiza and S.yunnanensis were analyzed by UPLC-DADand UPLC coupled with mass spectroscopy to evaluate the differences and similarities of the lipophilic components in the two kinds of herbs.The UPLC analysis of 18 batches of S.miltiorrhiza and 16 batches of S.yunnanensis was performed on a 25℃Thermo Accucore C_(18)column(2.1 mm×100 mm,2.6µm)by Shimadzu LC-20AD;mobile phase was 0.026%phosphoric acid(A)-acetonitrile(B)with gradient elution;flow rate was 0.4 m L·min~(-1);detection wavelength was set at 270 nm;injection volume was 2µL.The molecular structures of the lipophilic components were analyzed on a 25℃Thermo Accucore C_(18)column(2.1 mm×100 mm,2.6µm)by Thermo U3000 UPLC Q Exactive Orbitrap LC-MS/MS with a mobile phaseconsisting of 0.1%formic acid water(A)and 0.1%formic acidacetonitrile(B).The mass spectrometry was acquired in positive modes using ESI.There are 10 common peaks in the lipophilic components of S.miltiorrhiza.The similarity between the 16 batches of S.miltiorrhiza and their own reference spectra was greater than 0.942,and the average similarity was 0.973.There are 12 common peaks in the lipophilic components of S.yunnanensis.The similarity between the 18 batches of S.yunnanensis and their own reference spectra was greater than 0.937,and the average similarity was 0.976.The similarity between the reference chromatograms of S.miltiorrhiza and S.yunnanensis was only 0.900.There are three lipophilic components in S.yunnanensis,which are not found in S.miltiorrhiza,and one of which isα-lapachone.There is a lipophilic component in S.miltiorrhiza not found in S.yunnanensis,which may be miltirone.The two herbs contain 8 common lipophilic components including dihydrotanshinoneⅠ,cryptotanshinone,tanshinoneⅠ,tanshinoneⅡ_A,nortanshinone in which the content of tanshinoneⅡ_A,dihydrotanshinoneⅠand cryptotanshinone of S.yunnanensisis significantly lower than that of S.miltiorrhiza(P<0.01),and the contents of tanshinoneⅠand nortanshinone are significantly lower than that of S.miltiorrhiza too(P<0.05).There are significant differences in the types and contents of lipophilic components between the roots of S.miltiorrhiza and S.yunnanensis,and the similarity between the fingerprints of interspecies is much lower than that between the same species.Therefore,the roots of S.miltiorrhiza and S.yunnanensis are two kinds of herbs which are quite different in chemical compounds and compositions.

Rhodococcus daqingensis sp. nov., isolated from petroleum-contaminated soil.

A novel Gram-stain positive, aerobic, non-motile bacterial strain, designated Z1T, was isolated from a sample of petroleum-contaminated soil collected in Daqing, Heilongjiang province, China and characterised with a series of taxonomic approaches. The morphological and chemotaxonomic properties of the isolate were typical of those of members of the genus Rhodococcus. Phylogenetic analyses based on 16S rRNA gene sequences showed that strain Z1T belongs to the genus Rhodococcus and clustered with Rhodococcus maanshanensis DSM 44675T (99.2%, sequence similarity) and Rhodococcus tukisamuensis JCM 11308T (97.9%), respectively. However, the DNA-DNA hybridizations between strain Z1T and R. maanshanensis DSM 44675T and R. tukisamuensis JCM 11308T were both less than 70%. The optimal growth temperature and pH for strain Z1T were found to be at 28 °C and at pH 7.0. The peptidoglycan was found to contain meso-diaminopimelic acid; arabinose, galactose and glucose were detected as diagnostic sugars. The main polar lipids were identified as diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol, phosphatidylinositol mannoside and an unidentified lipid; MK-8(H2) was found as the major menaquinone. The major fatty acids were identified as C16:0, 10-methyl C18:0 and C18:1ω9c. Mycolic acids were found to be present. The G + C content of the genomic DNA was determined to be 66.7 mol%. Based on a comparative analysis of phenotypic and genotypic characteristics, in combination with DNA-DNA hybridization results, strain Z1T can be distinguished from the type strains of its two close neighbours as a novel species of the genus Rhodococcus, for which the name Rhodococcus daqingensis sp. nov. is proposed. The type strain is Z1T (= CGMCC 1.13630T = DSM 107227T).

Properties of pectin extracted from fermented and steeped hawthorn wine pomace: A comparison.

To investigate the characteristics of the pectins from fermented and steeped hawthorn wine pomace (FHP and SHP), they were acid-extracted, respectively, and compared detailedly. The results of esterification degree indicated that these two pectins were both defined as high methoxyl pectin, which was confirmed by Fourier transform infrared spectroscopy analysis. Molecular weight of SHP was higher than that of FHP. Differential scanning calorimetry analysis showed that the melt temperature of SHP was lower than that of FHP, and the molecular arrangement of SHP is more orderly than FHP. FHP and SHP solutions exhibited shear-thinning properties but SHP had a stronger anti-shear ability. FHP tended to be more elastic-solid as the concentration increased, which was not prominent in SHP. This study suggested that FHP and SHP powder could be new sources of pectin and both pectins could be potentially used as a thicker and stabilizer in food processing.

Exposure to PFDoA causes disruption of the hypothalamus-pituitary-thyroid axis in zebrafish larvae.

Perfluorododecanoic acid (PFDoA), a kind of perfluorinated carboxylic acid (PFCA) with 12 carbon atoms, has an extensive industrial utilization and is widespread in both wildlife and the water environment, and was reported to have the potential to cause a disruption in the thyroid hormone system homeostasis. In this study, zebrafish embryos/larvae were exposed to different concentrations of PFDoA (0, 0.24, 1.2, 6 mg/L) for 96 h post-fertilization (hpf). PFDoA exposure caused obvious growth restriction connected with the reduced thyroid hormones (THs) contents in zebrafish larvae, strengthening the interference effect on the growth of fish larvae. The transcriptional level of genes within the hypothalamic-pituitary-thyroid (HPT) axis was analyzed. The gene expression levels of thyrotropin-releasing hormone (trh) and corticotrophin-releasing hormone (crh) were upregulated upon exposure to 6 mg/L of PFDoA, and iodothyronine deiodinases (dio2) was upregulated in the 1.2 mg/L PFDoA group. The transcription of thyroglobulin (tg) and thyroid receptor (trß) were significantly downregulated upon exposure to 1.2 mg/L and 6 mg/L of PFDoA. PFDoA could also decrease the levels of sodium/iodide symporter (nis) and transthyretin (ttr) gene expression in a concentration-dependent manner after exposure. A significant decrease in thyroid-stimulating hormoneß (tshß), uridinediphosphate-glucuronosyltransferase (ugt1ab) and thyroid receptor (trα) gene expression were observed at 6 mg/L PFDoA exposure. Upregulation and downregulation of iodothyronine deiodinases (dio1) gene expression were observed upon the treatment of 1.2 mg/L and 6 mg/L PFDoA, respectively. All the data demonstrated that gene expression in the HPT axis altered after different PFDoA treatment and the potential mechanisms of the disruption of thyroid status could occur at several steps in the process of synthesis, regulation, and action of thyroid hormones.

miR-219 Cooperates with miR-338 in Myelination and Promotes Myelin Repair in the CNS.

A lack of sufficient oligodendrocyte myelination contributes to remyelination failure in demyelinating disorders. miRNAs have been implicated in oligodendrogenesis; however, their functions in myelin regeneration remained elusive. Through developmentally regulated targeted mutagenesis, we demonstrate that miR-219 alleles are critical for CNS myelination and remyelination after injury. Further deletion of miR-338 exacerbates the miR-219 mutant hypomyelination phenotype. Conversely, miR-219 overexpression promotes precocious oligodendrocyte maturation and regeneration processes in transgenic mice. Integrated transcriptome profiling and biotin-affinity miRNA pull-down approaches reveal stage-specific miR-219 targets in oligodendrocytes and further uncover a novel network for miR-219 targeting of differentiation inhibitors including Lingo1 and Etv5. Inhibition of Lingo1 and Etv5 partially rescues differentiation defects of miR-219-deficient oligodendrocyte precursors. Furthermore, miR-219 mimics enhance myelin restoration following lysolecithin-induced demyelination as well as experimental autoimmune encephalomyelitis, principal animal models of multiple sclerosis. Together, our findings identify context-specific miRNA-regulated checkpoints that control myelinogenesis and a therapeutic role for miR-219 in CNS myelin repair.

Geodermatophilus daqingensis sp. nov., isolated from petroleum-contaminated soil.

A novel Gram-positive actinobacterium, designated WT-2-1T, was isolated from a sample of petroleum-contaminated soil collected in Daqing, Heilongjiang province, China and characterised using a polyphasic taxonomic approach. The optimal growth for strain WT-2-1T was found to be at 25-35 °C and at pH 6.0-9.0 and with 0-4% (w/v) NaCl, forming blackish green-coloured colonies. Chemotaxonomic and molecular characteristics of the isolate match those described for members of the genus Geodermatophilus. The peptidoglycan was found to contain meso-diaminopimelic acid; galactose, glucose and xylose were detected as diagnostic sugars. The main phospholipids were identified as diphosphatidylglycerol, phosphatidylcholine, phosphatidylinositol, phosphatidylethanolamine and phosphatidylglycerol; MK-9(H4) was the dominant menaquinone present. The major cellular fatty acids were identified as iso-C16:0 and iso-C15:0. 16S rRNA gene sequence analysis showed that strain WT-2-1T is a member of the genus Geodermatophilus, with high sequence similarities to Geodermatophilus aquaeductus BMG801T (98.4%), Geodermatophilus saharensis CF5/5T (98.4%), Geodermatophilus bullaregiensis BMG841T (98.3%) and Geodermatophilus normandii CF5/3T (98.3%). Based on the phenotypic characteristics, phylogenetic data and DNA-DNA hybridization results, the isolate is concluded to represent a novel species of the genus Geodermatophilus, for which the name Geodermatophilus daqingensis sp. nov. is proposed. The type strain is WT-2-1T (=CGMCC 4.7381T = DSM 104001T).

Evaluation of the effects of active fractions of chinese medicine formulas on IL-1ß, IL-6, and TNF-α release from ANA-1 murine macrophages.

ETHNOPHARMACOLOGICAL RELEVANCE: Yaotongning (YTN) is a traditional Chinese Medicine (TCM) that contains ten component medicinal materials (CMMs) and uses Chinese rice wine as a vehicle to enhance its efficacy. YTN has been used for rheumatoid arthritis (RA) treatment in China for decades and has been reported to have anti-inflammatory and analgesic effects, as well as to strengthen the immune system. AIM OF THE STUDY: The present work quantitatively evaluated the in vitro effects of active fractions from the ten CMMs that make up YTN and eight additional herbs commonly used in TCM formulas for RA treatment, as well as different combinations of these active fractions, on cellular immune response; the findings were used to determine which active fractions are responsible for promoting an immune response, and to assess whether YTN is superior to other similar formulas and whether YTN can be improved by simplifying its formula from the point of its cellular immunomodulatory activity. MATERIALS AND METHODS: Using the YTN formulation principles and some concepts in combinatorial chemistry, 27 TCM samples were designed by combining some or all of the active fractions of YTN and other eight herbs used for RA treatment. Release of interleukin-1ß (IL-1ß), interleukin-6 (IL-6), and tumor necrosis factor-alpha (TNF-α) from ANA-1 murine macrophages was measured using an enzyme-linked immunosorbent assay (ELISA). The immunoregulatory effects of the TCM samples were evaluated by comparing their half-effective concentrations (EC50) for stimulating the release of these cytokines. RESULTS: Among the investigated active fractions, the flavonoids from Carthamus tinctorius (Fct), Davallia mariesii (Fdm), and Cinnamomum cassia Twig volatile oils (Vca) from the eight selected herbs effectively promoted IL-1ß and IL-6 release from ANA-1 cells. Saponins from the YTN CMM Glycyrrhiza uralensis (Sgu) were the most potent promoters of IL-1ß and TNF-α release. The aqueous extract of YTN CMM Eupolyphaga sinensis (Ves) strongly enhanced the release of IL-1ß, IL-6, and TNF-α from ANA-1 cells. The EC50 values for stimulating the secretion of IL-1ß, IL-6, and TNF-α could be determined for only six samples. The full YTN formula and the sample containing 50% Glycyrrhiza uralensis saponins, 25% of the mixture of alkaloids, and 25% of the mixture of all flavonoids exhibited good comprehensive cellular immunomodulatory activity. The immunomodulatory activity of the complete YTN formula was better than that of the sample containing all active fractions of YTN without Chinese rice wine (the YTN vehicle). CONCLUSIONS: Sgu and Ves are the primary active fractions of YTN involved in stimulating immune responses. The YTN prescription was reasonably effective at promoting cellular immune responses. Chinese rice wine, the YTN vehicle, strengthened the immunoregulatory activity of YTN. The results of this study demonstrate that the YTN recipe could be improved by reducing the number of CMMs and altering some active fractions without reducing its activity to promote cellular immune responses.

[Exploration of rapidly determining quality of traditional Chinese medicines by (NIR) spectroscopy based on internet sharing mode].

Because of the numerous varieties of herbal species and active ingredients in the traditional Chinese medicine(TCM),the traditional methods employed could hardly satisfy the current determination requirements of TCM.The present work proposed an idea to realize rapid determination of the quality of TCM based on near infrared(NIR)spectroscopy and internet sharing mode. Low cost and portable multi-source composite spectrometer was invented by our group for in-site fast measurement of spectra of TCM samples. The database could be set up by sharing spectra and quality detection data of TCM samples among TCM enterprises based on the internet platform.A novel method called as keeping same relationship between X and Y space based on K nearest neighbors(KNN-KSR for short)was applied to predict the contents of effective compounds of the samples. In addition,a comparative study between KNN-KSR and partial least squares(PLS)was conducted. Two datasets were applied to validate above idea:one was about 58 Ginkgo Folium samples samples measured with four near-infrared spectroscopy instruments and two multi-source composite spectrometers,another one was about 80 corn samples available online measured with three NIR instruments. The results show that the KNN-KSR method could obtain more reliable outcomes without correcting spectrum.However transforming the PLS models to other instruments could hardly acquire better predictive results until spectral calibration is performed. Meanwhile,the similar analysis results of total flavonoids and total lactones of Ginkgo Folium samples are achieved on the multi-source composite spectrometers and near-infrared spectroscopy instruments,and the prediction results of KNN-KSR are better than PLS. The idea proposed in present study is in urgent need of more samples spectra, and then to be verified by more case studies.

Efficacy of poly-unsaturated fatty acid therapy on patients with nonalcoholic steatohepatitis.

AIM: To examine whether poly-unsaturated fatty acid (PUFA) therapy is beneficial for improving nonalcoholic steatohepatitis (NASH). METHODS: In total, 78 patients pathologically diagnosed with NASH were enrolled and were randomly assigned into the control group and the PUFA therapy group (added 50 mL PUFA with 1:1 ratio of EHA and DHA into daily diet). At the initial analysis and after 6 mo of PUFA therapy, parameters of interest including liver enzymes, lipid profiles, markers of inflammation and oxidation, and histological changes were evaluated and compared between these two groups. RESULTS: At the initial analysis, in patients with NASH, serum levels of alanine aminotransferase (ALT) and aspartase aminotransferase (AST) were slightly elevated. Triglyceride (TG), total cholesterol (TC) and low-density lipoprotein cholesterol levels, markers of systemic inflammation [C-reactive protein (CRP)] and oxidation [malondialdehyde (MDA)], as well as fibrosis parameters of type IV collagen and pro-collagen type III pro-peptide were also increased beyond the normal range. Six months later, ALT and AST levels were significantly reduced in the PUFA group compared with the control group. In addition, serum levels of TG and TC, CRP and MDA, and type IV collagen and pro-collagen type III pro-peptide were also simultaneously and significantly reduced. Of note, histological evaluation showed that steatosis grade, necro-inflammatory grade, fibrosis stage, and ballooning score were all profoundly improved in comparison to the control group, strongly suggesting that increased PUFA consumption was a potential way to offset NASH progression. CONCLUSION: Increased PUFA consumption is a potential promising approach for NASH prevention and reversal.

[Preliminary study on suitability of ozone sterilization in traditional Chinese medicine and its preparation].

Microbial contamination, growth and reproduction have a great influence on the quality of traditional Chinese medicine (TCM) and its preparation. TCM may be polluted by microbial in the production process due to ambient air, facilities and appliances and operating personnel, which affects the quality and efficacy of the final product. The GMP certification inspection standard of TCM manufacturing enterprises clearly established: "Sterilization methods of Chinese herbal medicine, intermediate products, finished products should follow the principle whether or not to change its quality", "TCM powder that used as medicine directly should follow the principle whether to do microbiological examinations". So it's particularly important for product quality, corporate energy consumption and its efficiency to choose the scientific and effective sterilization techniques and methods. Ozone is a kind of safe, environmentally friendly, efficient and no residue emerging sterilization technology. It has been widely applied in various fields of medical and health care and production and living. This paper mainly analyzed the ozone sterilization technology of TCM, aiming to explore the principle of ozone sterilization, the advantages, application status and existing problems and so on. The management regulations and implementation rules of ozone sterilization were summarized to make sterilization of Chinese medicine in the production, management, quality control and other aspects standardized, reasonable and scientific, and to provide the theoretical reference of the ozone sterilization technology for TCM and its preparation.

Quantitative evaluation of in vitro effects and interactions of active fractions in a Chinese medicinal formula (Yaotongning Capsule) on rat chondrocytes.

ETHNOPHARMACOLOGICAL RELEVANCE: Yaotongning Capsule (YTNC) is a Traditional Chinese Medicinal (TCM) formula that has been demonstrated to be effective for osteoarthritis (OA) treatment in clinical use. Many compounds and 10 component medicinal materials (CMMs for short, i.e., the fundamental elements used in TCM formulas) in YTNC are challenging to study the pharmacological effects and interactions of the CMMs. Besides, it is difficult to know whether the YTNC formula is reasonable, and if YTNC formula could be improved without comparing YTNC with other TCM formulas of treating OA. Based on different combinations of the active fractions from the 10 CMMs of YTNC and eight additional herbs frequently used in the TCM formulas of treating OA, the present study evaluated systematically the in vitro effects of these active fractions and the interactions among the active fractions from YTNC on rat chondrocytes to find possible solutions of the above questions. MATERIALS AND METHODS: Based on the formulation of YTNC and the concept of combinatorial chemistry, the active fractions were applied to form the whole YTNC prescription (i.e., the combination of all YTNC active fractions and the extract of YTNC׳s vehicle), five disassembled formulas of YTNC (i.e., the combinations of some active fractions in YTNC) and 21 TCM samples consisted of different kinds of active fractions. The degenerated chondrocytes were induced with interleukin-1ß (IL-1ß), and then the half-effective concentration (EC50) value of the proliferation activity was analyzed to evaluate the 27 TCM samples. Nine samples were screened for the following evaluation on glycosaminoglycan (GAG) synthesis. Rat articular cartilage was obtained from six Sprague-Dawley rats (seven days of age), and then chondrocytes were isolated through enzymatic digestion with 0.2% Collagenase II. Proliferations of chondrocytes were examined through Cell Counting Kit-8 assay, when the intracellular levels of GAG were detected by 1,9-Dimethylmethylene blue staining. The interactions between the active fractions in YTNC were evaluated by comparing experimental EC50 values of the YTNC formulas with their additive EC50 values. The effects of every active fraction were estimated by comparing the EC50 values of the TCM sample containing the active fraction with that of the initial sample without the active fraction. RESULTS: The whole formula of YTNC was very good at promoting the proliferation and GAG synthesis among all the 27 TCM samples. The vehicle of YTNC (Chinese rice wine) strengthened the two activities of YTNC. Refer to promoting the proliferation in chondrocytes, Davallia mariesii flavonoids (not belong to YTNC) were more potent than Glycyrrhiza uralensis flavonoids in YTNC, while the saponins, volatile oils and polysaccharides of YTNC were more potent than those from the eight additional herbs. Some samples including fewer active fractions were as good as YTNC. The YTNC formula and its disassembled formulas exhibited good activities both in promoting the proliferation and GAG synthesis, and the whole formula was most potent among the six YTNC formulas. CONCLUSIONS: The YTNC formula is reasonable and has advantage in promoting the proliferation and GAG synthesis in IL-1ß induced chondrocytes. YTNC׳s vehicle Chinese rice wine plays an important role in strengthening the activity of YTNC. YTNC may have the potential activity on treating chondrocytes degeneration caused by OA. However, the formula still can be simplified based on the combination of alkaloids, flavonoids and 50% of saponins from Glycyrrhiza uralensis to improve its quality controllability and safety. The present study can be a quite purposeful work for developing new YTNC-based formulas with maximal therapeutic efficacy and minimal adverse effects.

Quantitative evaluation of the in vitro effect and interactions of active fractions in Yaotongning-based formulae on prostaglandin E2 production.

ETHNOPHARMACOLOGICAL RELEVANCE: Traditional Chinese medicine (TCM) formula of Yaotongning Capsules (YTNC) is a common remedy to treat rheumatism (RA) in China and possesses diverse biological activities including anti-inflammation. However the effects of component material medicines (CMMs) in YTNC and different combinations of the CMMs on the efficacy of YTNC and the interactions of these CMMs have been being unclear due to ten CMMs and too many compounds involved in YTNC. Moreover, many TCM formulae are available for treating RA according to TCM theory. It is unknown if the YTNC prescription is better than other TCM formulae for treating RA or better efficacy could be obtained when some CMMs in YTNC are replaced by other herbs. Quantitatively investigate the in vitro effect of active fractions from the CMMs of YTNC and other eight herbs commonly used in the TCM formulae for RA treatment on anti-inflammatory activity of different combinations of the active fractions, the interactions of the active fractions to evaluate the reasonability, advantage (or disadvantage) of the YTNC prescription and to see if the prescription could be improved from the point of anti-inflammation. MATERIALS AND METHODS: Twenty-six active fractions, which were categorized as alkaloids, flavonoids, saponins, volatile oils/aqueous extracts and polysaccharides were prepared to design TCM samples by combining some of the active fractions, based on the YTNC formulating principle, combination chemistry concept and the importance of the active fractions in YTNC. The anti-inflammatory activities of the samples were evaluated by their half-maximal inhibitory concentration (IC50) values that inhibiting the production of prostaglandin E2 (PGE2) in ANA-1 murine macrophages (ANA-1 cells). The cells plated in 96-well plates were classified into blank group and test sample group. Each group was stimulated with lipopolysaccharides (LPS, 1 mg/mL) for 2h. ANA-1 cells were pretreated with different concentrations of test samples prior to the addition of arachidonic acid (10 µmol/L). The supernatants were collected and measured using PGE2 ELISA Kit, and the cytotoxicity was assayed by cell counting Kit-8 (CCK8)-based test. The interactions of the active fractions in YTNC were evaluated by comparing the experimental IC50 values of the samples derived from YTNC to their corresponding additive IC50 values. The effect of each active fraction on cellular anti-inflammation-PGE2 secretion inhibition activity, and the reasonability, advantages (or disadvantages) of YTNC were evaluated based on the comparison of IC50 values of the samples. RESULTS: The disassembled formulae consisted of some active fractions of YTNC and the whole prescription of YTNC consisted of the all active fractions from YTNC all demonstrate cellular anti-inflammatory activity, and there were no significant differences between these formulae. The vehicle of YTNC Chinese rice wine exhibits the ability to enhance the cellular anti-inflammation of YTNC. Synergistic effect exerts in the combination of alkaloids, flavonoids and saponins of YTNC, antagonistic or additive effects occur in the other combinations of active fractions from YTNC. The anti-inflammatory activities of some TCM samples which include some active fractions from the eight selected herbs are significantly higher than the samples derived from YTNC. The flavonoids of Carthamus tinctorius, the volatile oils of Cinnamomum cassia and Angelica pubescens perform better in cellular anti-inflammation than the flavonoids and volatile oils in YTNC. CONCLUSIONS: The prescription of YTNC is reasonable in the view of anti-inflammation. The saponins and polysaccharides from the CMMs of YTNC have better anti-inflammatory activities than the saponins and polysaccharides from the other eight herbs. Reducing the varieties of YTNC CMMs and replacing the flavonoids and volatile oils of YTNC with the flavonoids of Carthamus tinctorius and the volatile oils of Cinnamomum cassia (or Angelica pubescens) would improve the safety and anti-inflammatory activity of YTNC. Synthetically evaluating various pharmacological activities of TCM formulae designed in the present work may lead to develop more effective and safer TCM using YTNC as prototypes.

[Extraction technology optimization and quality analysis of volatile oil in Rhizoma Curcumae].

OBJECTIVE: To investigate the effect of technologies and conditions on volatile oil yield extracted from Rhizoma Curcumae. METHODS: Water Extraction coupling Rectification (WER) and Steam Distillation (SD) technologies were applied to extract the volatile oils based on orthogonal table L9 (3(3)) to find out optimized condition. RESULTS: Variance and range analysis of orthogonal experiment results showed that the best conditions of WER and SD were as follows: ultrasound 0 h, extract 12 h with 8 (or 12 fold water for SD) fold water amount. Paired T test on the yields of the oils indicated that the oil yields prepared by WER and SD were significantly different. GC-MS analysis characterized 12 common compounds,which occupied 97.19% (SD) and 92.25% (WER) of the ones identified, respectively. Moreover, the relative percentage of the common constituents were almost the same. CONCLUSION: Ultrasound is not good for extracting volatile oil from Rhizoma Curcumae. WER could not only increase the oil yield of Rhizoma Curcumae, but also keep the quality of the oils accord with that extracted by SD.

A strategy for evaluating antipyretic efficacy of Chinese herbal medicines based on UV spectra fingerprints.

ETHNOPHARMACOLOGICAL RELEVANCE: Quality analysis and control of Chinese herbal medicines (CHM) or herbal medicines (HM) are being more and more investigated based on fingerprint analysis, and there are also some researches on correlating fingerprints of CHM to their efficacy. Multi-component analysis methods together with fingerprints are considered potential useful tools to select candidate herbal drugs from extracts of herbs in pharmacological/bio-prospecting investigations. AIM OF THE STUDY: To explore a strategy for evaluating efficacy strength of CHM samples based on their spectra fingerprints and validate it. METHODOLOGY: Radix bupleuri (RB), a typical Chinese medicinal herb for relieving exterior syndrome, and Flos lonicerae (FL), Fructus forsythiae (FF), and Radix isatidis (RI) that are widely applied Chinese herbs for heat clearing and detoxifying, were selected as herbal sources. The aqueous extracts, volatile oils and mixtures of the extracts and oils of the four herbs, plus Ibuprofen suspension (IS), Shuanghuanglian oral liquid (SHL), mixture of SHL and the volatile oils of FL and FF, were used for subject samples to do antipyretic experiments on rats. Ultraviolet spectra were used as the spectra fingerprints to represent chemical characteristics of the samples. Principal component analysis (PCA) and canonical correlation analysis (CCA) were adopted as evaluation tools to establish the correlation between pharmacological and spectra data, from which a spectral index for evaluating antipyretic effects of CHM samples was constructed. Furthermore, four compound samples were designed by mixing 50% volatile oils and 50% aqueous extracts of the four herbs with different ratios to validate the strategy. RESULTS: Efficacy sequence of the 15 calibrating and 4 validating CHM samples, defined by the first canonical correlative variable U(1) of their UV spectra, was consistent with that given by pharmacological experiments. CONCLUSIONS: The strategy proposed in this study could be applied to evaluate efficacy strength of CHM and helpful for screening candidate herbal drugs from different herbs or prepared by different technologies.

[Water extraction coupling rectification for isolating volatile oils of Ligusticum chuanxiong].

To improve the oil yield of Chuanxiong volatile oils and keep their quality same as that of oils prepared by steam distillation (SD) for short), a new technology of coupling water extraction and rectification (WER for short) was applied to isolate Chuanxiong volatile oils. The oil yield, as well as GC-MS chromatogram of the volatile oils extracted by WER and SD were compared. The oil yield of WER (0.50%) was that of SD (0.20%). There were 21 common constituents in the two kinds of volatile oils, which occupied 98.94% of the oils extracted by SD, 98.80% of that by WER, respectively. Moreover, the relative percentage of the common constituents was almost the same. The results indicate that WER could not only increase the oil yield of Ligusticum chuanxiong, but also keep the quality of the oils accord with that extracted by SD.