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OBJECTIVE To explore the mechanism of Kuaisong yin in the prevention and treatment of constipation. METHODS Slow transit constipation （STC） model was established with Compound difenoxylate tablet in mice and rats. Two batches of mice were divided into blank group， model group， positive control group （Maren soft capsule， 0.64 g/kg）， Kuaisong yin low-dose， medium-dose and high-dose groups （3.2， 6.4， 12.8 g/kg）， with 10 mice in each group. The effect of Kuaisong yin on constipation in mice was evaluated by intestinal propulsion experiment and defecation experiment. Rats were divided into blank group， model group， positive control group （Maren soft capsule，0.36 g/kg）， Kuaisong yin low-dose and high-dose groups （2.4， 4.8 g/kg）， with 7 or 8 rats in each group. They were given relevant medicine once a day for 1 week. The metabonomics of serum and urine of rats were analyzed by UPLC-Q-TOF-MS/MS technology. RESULTS Compared with model group， the ink propulsion rate and 5 h defecation volume of mice in Kuaisong yin high-dose group were significantly increased （P＜0.05）； the first defecation time of mice in Kuaisong yin medium-dose and high-dose groups was significantly shortened， and the quality of defecation was significantly reduced within 5 h （P＜0.05 or P＜0.01）. Serum metabonomics screened 16 compounds （such as proline， propionylcarnitine， hemolytic phosphatidylcholine， etc.） and 6 metabolic pathways （such as sphingomyelin metabolism， arginine and proline metabolism， sphingolipid biosynthesis-lactose and neolactone series）. Urine metabonomics screened 20 different metabolites （such as prostaglandin A2， L-valine， phosphatidylcholine， sphingomyelin， etc.） and 8 metabolic pathways （such as valine， leucine and isoleucine biosynthesis， sphingomyelin metabolism， pyruvate metabolism， etc.）. CONCLUSIONS Kuaisong yin can play a role in improving constipation by regulating different metabolites such as hemolytic phosphatidylcholine， phosphatidylcholine， prostaglandin A2， L-valine， proline， and regulating metabolic pathways such as multiple amino acid metabolism， sphingomyelin metabolism， etc.
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OBJECTIVE To analyze the odor composition changes of two kinds of traditional Chinese medicine sachet （children type and adults type） with different placement time by using ultra-fast gasphase electronic nose technology. METHODS The change rule of sachet components at different storage times was analyzed by gas chromatography. At the same time， the qualitative results were obtained by combining electronic nose with Arochembase database. Discriminant factor analysis was used to analyze the overall odor composition differences of the two sachet samples. RESULTS A total of 10 odor compositions were identified in children-type sachet， including α-pinene and β-pinene as the functional index compositions； five odor compositions of children-type sachet disappeared after 0.25 days， and most of them disappeared after 7 days； the cumulative contribution rate of discriminant factor analysis was 99.225%. A total of 8 odor compositions were identified in adult-type sachets， including α-pinene and α-phellandrene as the functional index compositions； four odor components disappeared after the adult-type sachet was placed for 0.25 days； after 15 days of placement， the peak 6-8 disappeared， and the intensity of peak 5 decreased by 34.3% compared with 0 day of placement； the cumulative contribution rate of discriminant factor analysis was 91.965%. CONCLUSIONS With the extension of storage time， the smell and composition of the two traditional Chinese medicine sachets are decreasing. It is recommended that the use time of children-type sachet is 7 days， and that of adult-type sachet is 15 days.
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OBJECTIVE To study the mechanism of Compound zaoren granule in improving insomnia. METHODS Forty-nine mice were divided into blank group， model group， positive control group 1 （Estazolam tablets 0.5 mg/kg），control group 2 （Shumian capsule 0.6 g/kg）， Compound zaoren granule low-dose， medium-dose and high-dose groups （2.5， 5， 10 g/kg）， with 7 mice in each group. The insomnia model was established by chronic unpredictable mild stress combined with 4-chloro-DL- phenylacetic acid. The behavioral changes of mice were investigated through open field test and pentobarbital sodium synergistic hypnosis experiment， as well as the pathomorphology of mice hypothalamus tissue was observed by HE staining. The metabonomics analysis and multivariate statistical analysis of serum in mice were performed by UHPLC-Q-TOF-MS/MS， and the differential metabolites were screened out； the metabolic pathway analysis was conducted based on MetaboAnalyst 5.0 database. RESULTS Compared with blank group， the total travelling distance， the number of entering the central region and the moving distance in the central region of the model group were significantly reduced （P＜0.05）， the proportion of total rest time was significantly increased （P＜0.05）， the sleep duration of mice was significantly shortened （P＜0.05）， and hypothalamic nerve cells damaged and severely vacuolated. Compared with model group， the total travelling distance of Compound zaoren granule low-dose and medium-dose groups were increased significantly and the proportions of total rest time of those groups were decreased significantly （P＜0.05）， and the sleep duration of mice in Compound zaoren granule high-dose group was prolonged significantly （P＜0.05）； the hypothalamic nerve cells of mice in each administration group recovered to varying degrees， and the hypothalamus histiocytes of mice in the Compound zaoren granules high-dose group were closer to those in the blank group. A total of 18 differential metabolites （such as phenylalanine， taurine， norvaline， methionine） and 4 important amino acid metabolic pathways （L-phenylalanine， tyrosine and tryptophan biosynthesis； taurine and hypotaurine metabolism； L-phenylalanine metabolism； cysteine and methionine metabolism） were identified through metabolomics analysis. CONCLUSIONS Compound zaoren granules can normalize the disordered metabolism in vivo by regulating differential metabolites such as phenylalanine， taurine， and four amino acid metabolic pathways， so as to improve insomnia.
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OBJECTIVE：To esta blish the m ethod for identifying Schizonepeta tenusfolia from different habitats based on odor information. METHODS ：The odor of S. tenusfolia from different habitats were identified by Heracles Ⅱ ultra-fast gas phase electronic nose. Qualitative analysis was conducted according to obtained chromatographic information combined with AroChemBase database and Kovats retention index qualitative database. Principle component analysis （PCA）and discriminant factor analysis （DFA）were conducted by using Alpha Soft V 14.2 software，and cluster analysis （CA）was performed with SPSS 22.2 software. RESULTS ：There were 16 common peaks in 15 batches of S. tenusfolia from different habitats. After comparison with AroChemBase database and Kovates retention index qualitative database ，a total of 13 possible components were obtained. The possible components and sensory description information of S. tenusfolia from different habitats were basically the same ，but only the content was different. The chromatographic peak intensities of common peak No. 2 were in descending order as Anhui ＞ Gansu＞Henan＞Hebei＞Jiangsu，the chromatographic peak intensities of common peak No. 6 were in descending order as Anhui ＞ Hebei＞Gansu≈Henan＞Jiangsu，the chromatographic peak intensities of common peak No. 9 were in descending order as Anhui ＞ Gansu＞Henan＞Jiangsu＞Hebei，the chromatographic peak intensity of common peak No. 13 were in descending order as Anhui ≈ Gansu＞Hebei＞Jiangsu＞Henan，which represented the chromatographic peak intensity of methyl formate （peak No. 2），α-pinene （peak No. 6），3-nonone（peak No. 9）and α-terpineol（peak No. 13）were significantly different due to the change of habitats. PCA results showed that the cumulative contribution rate of the first two principal components was 96.807％. Results of DFA showed that contribution rates of discriminant factor 1 and discriminant factor 2 were 92.089％ and 3.982％. CA results showed that when the distance was 10，15 batches of samples could be clustered into 3 categories，B1-B5 and J 1-J3 into one category ，A1-A3 into one category ，G1，G2，N1 and N 2 into one category. The results were basically consistent with those of PCA and DFA. CONCLUSIONS：Ultra-fast gas phase electronic nose technology can be used to identify S. tenusfolia from different habitats rapidly. Methyl formate ，α-pinene，3-nonone and α-terpineol may be the key factors to distinguish S. tenusfolia from different habitats.
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Objective] To amplify and sequence the light chain of anti idiotypic monoclonal antibody NP30 of Schistosoma japonicum. [Methods] By comparing the conserved regions at each end of the nucleotide sequences of murine germ line genes enco ding FR1 and FR4 regions of immunoglobulin light chain variable regions, we designed a set of primers for amplification of V L gene. The hybridoma cells secreting anti idiotypic monoclonal antibody NP30 of Schistosoma japonicum were cultured and their genome DNAs were extracted and used as templates for PCR. The PCR product was then cloned into pUC19 vector. The recombinants were sequenced by Sanger′s method. The V L gene was compared with GenBank and published mouse V L genes. [Results] The full length of V L gene was 318 bp. The V L gene was a member of mouse Ig ? light chain subgroup IV and generated from rearrangement of germ line V and J? 4 genes. The V L gene sequence has been registered by GenBank(accession No. AF206720). [Conclusion] The obtained V L gene was a potentially functional gene of anti idiotypic monoclonal antibody NP30 of Schistosoma japonicum .
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Objective To construct single chain antibody specific to membrane protein of Schistosoma japonicum by gonetic engineering technique. Methods The V\-H (heavy-chain variable region) and V\-L (light-chain variable region) genes were amplified by PCR from the genomic DNA of NP11-4 cell line, and sequenced by Sanger's method. The ScFv was constructed in pTHA90 vector using V\-H and V\-L genes, then expressed by IPTG. Results The V\-H and V\-L genes were obtained through PCR. The DNA sequences showed that V\-H and V\-L were new variable region genes of antibody. They were registered by GenBank. A ScFv gene with (Gly4Ser) 3 intralinker in the pTHA90 vector was successfully constructed. The ScFv was expressed as thioredoxin-fused proteins about 36^2 kDa. Conclusion A specific ScFv against the membrane protein of Schistosoma japonicum was constructed and expressed.