Chemical Composition of Rubus pirifolius Smith

[Objectives]This paper aimed to study the chemical composition of Rubus pirifolius Smith[Methods]A preliminary experiment was carried out using the qualitative reactions of various chemical constituents,in order to roughly detect the types of chemical constituents that may be contained in R.pirifolius Smith The ethanol extract of R.pirifolius Smith were separated and purified using silica gel column chromatography,silica gel thin-layer chromatography and recrystallization,and the chemicals obtained were identified using mass spectrometry,nuclear magnetic resonance and literature reference.[Results]The main constituents of R.pirifolius Smith are saponins,tannins,flavonoids,phytosterols,triterpenes and anthraquinones.A total of five compounds were isolated from the ethanol extract of R.pirifolius Smith,and they were identified asβ-sitosterol(1),arjunolic acid(2),stigmasterol-3-O-β-D-glucopyranoside(3),euscaphic acid(4)and 3,3′,4-trimethyle llegic acid(5).[Conclusions]Compounds 1,2,3,4 and 5 were isolated from R.pirifolius Smith for the first time.

Content Determination of Flavonoids in Radix Millettiae Speciosae from Different Areas of Guangxi

[Objectives] This study aimed to investigate the quality of Radix Millettiae Speciosae at different planting sites in Guangxi. [Methods] The content of flavonoids in Radix Millettiae Speciosae was determined by HPLC. The chromatographic conditions were as follows: column, ZORBAX SB-C_(18); mobile phase, acetonitrile-0.1% glacial acetic acid(42∶58); detection wavelength, 250(formononetin) and 310(maackiain) nm; flow rate, 1.0 mL/min; and column temperature, 30℃. [Results] Under the chromatographic conditions above, formononetin and maackiain could be completely separated from impurities. The standard curve had a good linear relationship(R^2>0.999 9). The precision and stability met analysis requirements. The average recovery rates were 97.53% and 98.54%, respectively, and the RSD values were 2.03% and 1.88%, respectively, indicating that the established method has good reproducibility, durability and accuracy. The content of formononetin was highest in the Radix Millettiae Speciosae from Yongning District, Nanning City, and the content of maackiain was highest in the Radix Millettiae Speciosae from Xichang Town, Hepu County. [Conclusions] The HPLC method established in this study is simple, accurate and stable. It can be used as a quality control method for Radix Millettiae Speciosae. The contents of flavonoids in Radix Millettiae Speciosae from Yongning District, Nanning City and Xichang Town, Hepu County were higher than those from other planting bases.

Determination of Chlorogenic Acid in Stems of Mussaenda pubescens Ait.f.by High Performance Liquid Chromatography

[Objectives] This study was conducted to establish a method for determination of chlorogenic acid in Mussaenda pubescens Ait.f.[Methods]HPLC used SHISEIDO C_(18) MG Ⅱ column( 5 μm,4.6 mml.D.× 250 mm) as chromatographic column and acetonitrile-0.4% phosphoric acid solution( 9∶91) as mobile phase.The separation was performed at a flow rate of 1.0 ml/min and a column temperature of 30 ℃,and the detection wavelength was set at 327 nm.[Results]Chlorogenic acid had a good linear relation in the range of 0.254 7-2.547 5 μg( R2= 0.999 9).The recovery rate was 97.8%,RSD = 1.82%( n = 9).[Conclusions]The content of chlorogenic acid in M.pubescens was determined by ultrasonic extraction and HPLC.The method was simple,stable and reliable and could be used for the quality control of M.pubescens.

Bioinformatic Analysis of ARF7 in Huperzia serrata

[Objectives]This paper aims to study the function of the ARF7 gene family of Huperzia serrata.[Methods]Based on the full-length transcriptome sequencing results of H.serrata,the unigenes of 17 H.serrata ARF7 candidate genes were analyzed using biogenetics technology.[Results]The molecular weight of the ARF7 proteins ranges from 11.28 to 608.54 kDa.HsARF7-4,HsARF7-5,HsARF7-8,HsARF7-9,HsARF7-10,HsARF7-13,HsARF7-16 and HsARF7-17 are basic proteins;all the proteins are unstable;except that HsARF7-6 and HsARF7-15 are hydrophilic,the rest are lipophilic proteins;there are 197 types of cis-acting elements,and 17 of the proteins contain as more as 20 acting elements such as GTGANTG10,CBFHV,GT1CONSENSUS,NODCON2GM,CAATBOX1,MYBCORE,GATABOX,WRKY71OS and RAV1AAT;HsARF7-1,HsARF7-2,HsARF7-4,HsARF7-6,HsARF7-8,HsARF7-10,HsARF7-12,HsARF7-14 and HsARF7-16 all have the 6 types of motif predicted;the proteins have domains such as PABP-1234 super family,RRM_SF super famliy,PRK10263 super family,AUX_IAA super family and MFS super family;the proteins all can cross cell membrane and have no signal peptide,and they are all located in the nucleus.On the level of secondary structure,the proportion ofβ-sheet is the smallest.In HsARF7-7 and HsARF7-9,α-helix has the largest proportion,and in the remaining proteins,random coil has the largest proportion;and tertiary structure prediction found that HsARF7-1,HsARF7-2,HsARF7-4,HsARF7-14 and HsARF7-17 have a tertiary structure of polyadenylate-binding protein,and HsARF7-7 has a tertiary structure of auxin-induced protein iaa4.[Conclusions]The above analysis results can provide a certain theoretical basis for further research on the biological function and regulatory mechanism of the ARF gene of H.serrata in the future,and provide a reference for the study of the growth and development of H.serrata.

Cloning and characterization of squalene synthase and cycloartenol synthase from Siraitia grosvenorii

Mogrosides and steroid saponins are tetracyclic triterpenoids found in Siraitia grosvenorii.Squalene synthase(SQS) and cycloartenol synthase(CAS) are key enzymes in triterpenoid and steroid biosynthesis.In this study,full-length cDNAs of SgSQS and SgCAS were cloned by a rapid amplification of cDNA-ends with polymerase chain reaction(RACE-PCR) approach.The SgSQS cDNA has a 1254 bp open reading frame(ORF) encoding 417 amino acids,and the SgCAS cDNA contains a 2298 bp ORF encoding 765 amino acids.Bioinformatic analysis showed that the deduced SgSQS protein has two transmembrane regions in the C-terminal.Both SgSQS and SgCAS have significantly higher levels in fruits than in other tissues,suggesting that steroids and mogrosides are competitors for the same precursors in fruits.Combined in silico prediction and subcellular localization,experiments in tobacco indicated that SgSQS was probably in the cytoplasm or on the cytoskeleton,and SgCAS was likely located in the nucleus or cytosol.These results will provide a foundation for further study of SgSQS and SgCAS gene functions in S.grosvenorii,and may facilitate improvements in mogroside content in fruit by regulating gene expression.

Cloning and expression of SgCYP450-4from Siraitia grosvenorii

CYP450 plays an essential role in the development and growth of the fruits of Siraitia grosvenorii. However, little is known about the SgCYP450-4 gene in S. grosvenorii. Here, based on transcriptome data, a full-length c DNA sequence of SgCYP450-4 was cloned by reverse transcriptasepolymerase chain reaction(RT-PCR) and rapid-amplification of c DNA ends(RACE) strategies. SgCYP450-4 is 1677 bp in length(Gen Bank accession No. AEM42985.1) and contains a complete open reading frame(ORF) of 1422 bp. The deduced protein was composed of 473 amino acids, the molecular weight is 54.01 kDa, the theoretical isoelectric point(PI) is 8.8, and the protein was predicted to possess cytochrome P450 domains. SgCYP450-4 gene was highly expressed in root, diploid fruit and fruit treated with hormone and pollination. At 10 days after treatment with pollination and hormones, the expression of SgCYP450-4had the highest level and then decreased over time, which was consistent with the development of fruits of S.Grosvenorii. Hormonal treatment could significantly induce the expression of SgCYP450-4. These results provide a reference for regulation of fruit development and the use of parthenocarpy to generate seedless fruit, and provide a scientific basis for the production of growth regulator application agents.

The effect of 8-OH-DPAT and dapoxetine on gene expression in the brain of male rats during ejaculation

The 5-HT_(1A) receptor agonist 8-hydroxy-2-[di-n-propylamino] tetralin(8-OH-DPAT) promotes ejaculation of male rats, whereas dapoxetine delays this process. However, the gene expression profile of the brain at ejaculation following administrationof these two compounds has not been fully elucidated. In the present study, a transcriptomic Body Map was generated by conducting mRNA-Seq on brain samples of male Sprague–Dawley rats. The study included four groups: pre-copulatory control(CK) group,ejaculation(EJ) group, 0.5 mg/kg 8-OH-DPAT-ejaculation group(DPAT), and 60 mg/kg dapoxetineejaculation(DAP) group. The resulting analysis generated an average of approximately 47 million sequence reads. Significant differences in the gene expression profiles of the aforementioned groups were observed in the EJ(257 genes), DPAT(349 genes) and the DAP(207 genes) compared with the control rats. The results indicate that the expression of Drd1 and Slc6a3 was significantly different after treatment with 8-OH-DPAT, whereas the expression of Drd4 was significantly different after treatment with dapoxetine. Other genes, such as Wnt9b, Cdkn1 a and Fosb, exhibited significant differences in expression after the two treatments and are related to bladder cancer, renal cell carcinoma and sexual addiction. The present study reveals the basic pattern of gene expression that was activated at ejaculation in the presence of 8-OH-DPAT or dapoxetine, providing preliminary gene expression information during rat ejaculation.