Cloning of transcription factor PcFBA-1 in Pogostemon cabin and its interaction with FPPS promoter / 中国中药杂志

Farnesyl diphosphate synthase(FPPS) is a key enzyme at the branch point of the sesquiterpene biosynthetic pathway, but there are no reports on the transcriptional regulation of FPPS promoter in Pogostemon cabin. In the early stage of this study, we obtained the binding protein PcFBA-1 of FPPS gene promoter in P. cabin. In order to explore the possible mechanism of PcFBA-1 involved in the regulation of patchouli alcohol biosynthesis, this study performed PCR-based cloning and sequencing analysis of PcFBA-1, analyzed the expression patterns of PcFBA-1 in different tissues by fluorescence quantitative PCR and its subcellular localization using the protoplast transformation system, detected the binding of PcFBA-1 protein to the FPPS promoter in vitro with the yeast one-hybrid system, and verified its transcriptional regulatory function by dual-luciferase reporter gene assay. The findings demonstrated that the cloned PcFBA-1 had an open reading frame(ORF) of 1 131 bp, encoding a protein of 376 amino acids, containing two conserved domains named F-box-like superfamily and FBA-1 superfamily, and belonging to the F-box family. Moreover, neither signal peptide nor transmembrane domain was contained, implying that it was an unstable hydrophilic protein. In addition, as revealed by fluorescence quantitative PCR results, PcFBA-1 had the highest expression in leaves, and there was no significant difference in expression in roots or stems. PcFBA-1 protein was proved mainly located in the cytoplasm. Furthermore, yeast one-hybrid screening and dual-luciferase reporter gene assay showed that PcFBA-1 was able to bind to FPPS promoter both in vitro and in vivo to enhance the activity of FPPS promoter. In summary, this study identifies a new transcription factor PcFBA-1 in P. cabin, which directly binds to the FPPS gene promoter to enhance the promoter activity. This had laid a foundation for the biosynthesis of patchouli alcohol and other active ingre-dients and provided a basis for metabolic engineering and genetic improvement of P. cabin.

SSR loci information analysis in transcriptome of Andrographis paniculata / 中国中药杂志

To study the SSR loci information and develop molecular markers, a total of 43 683 Unigenes in transcriptome of Andrographis paniculata were used to explore SSR. The distribution frequency of SSR and the basic characteristics of repeat motifs were analyzed using MicroSAtellite software, SSR primers were designed by Primer 3.0 software and then validated by PCR. Moreover, the gene function analysis of SSR Unigene was obtained by Blast. The results showed that 14 135 SSR loci were found in the transcriptome of A. paniculata, which distributed in 9 973 Unigenes with a distribution frequency of 32.36%. Di-nucleotide and Tri-nucleotide repeat were the main types, accounted for 75.54% of all SSRs. The repeat motifs of AT/AT and CCG/CGG were the predominant repeat types of Di-nucleotide and Tri-nucleotide, respectively. A total of 4 740 pairs of SSR primers with the potential to produce polymorphism were designed for maker development. Ten pairs of primers in 20 pairs of randomly picked primers produced fragments with expected molecular size. The gene function of Unigenes containing SSR were mostly related to the basic metabolism function of A. paniculata. The SSR markers in transcriptome of A. paniculata show rich type, strong specificity and high potential of polymorphism, which will benefit the candidate gene mining and marker-assisted breeding.

Comparison of development condition of different Liriope spicata seedlings / 中国中药杂志

The study identified the main morphological index of the seedlings classification including seedling age,the root width and number of newborn buds and coarse roots, according to the local agricultural production techniques and assessment of Liriope spicata's growth and development condition. After carrying on K cluster analysis of the morphological, we separated the seedlings into two levels. The first level (Ⅰ)： the new talent with the root width exceeding two point five millimeters, the new born buds exceeding three, and with the coarse root exceeding one. The second level (Ⅱ)： the old talent with the root width below one millimeters, the newborn buds below two and without coarse root. The study surveyed the plants' growth index dynamics, as well as the yield and quality of the tuberous root. The experimental results suggested that the growth condition of seedling Ⅰwas better, the yield of earthnut higher, the quality of earthnut more excellent. The study lied the foundation of L. spicata's grading standards and standardized production.

Preliminary study on effect of Rhodiolae Crenulatae Radix et Rhizoma cell wall-broken decoction pieces on intestinal flora of mice / 中国中药杂志

This study aims to analyze and compare the effect of cell wall-broken decoction pieces, conventional decoction pieces and conventional powder of Rhodiolae Crenulatae Radix et Rhizoma on the intestinal flora of normal mice. The conventional bacterial culture and PCR-DGGE (polymerase chain reaction-denaturing gradient gel electrophoresis) were adopted for the mice after the oral administration for 14 days. According to the bacterial culture results, the 1/8 dose cell wall-broken decoction pieces group showed fewer Enterococcus and Escherichia coli bacillus but more Lactobacillus and Bifidobacterium than the conventional decoction pieces group and the traditional powder group (P <0.05). Meanwhile, on the basis of the PCR-DGGE results, the 1/8 dose cell wall-broken decoction pieces group revealed the highest Shannon-Wiener index (H) and species richness (S) among the seven groups, with extremely significant differences compared with the normal group (P <0.01), significant differences compared with the conventional decoction pieces group and the conventional powder group (P <0.05) and a high intra-group similarity. In conclusion, the long-term intake of 1/8 dose Rhodiolae Crenulatae Radix et Rhizoma cell wall-broken decoction pieces showed a certain effect in regulating intestinal tract by promoting the growth of Lactobacillus and Bifidobacterium. Furthermore, the intestinal flora community will become more stable.

Effects of Pb2+ stress on seed germination & seedling growth of Rabdosia rubescens / 中国中药杂志

The seeds of Rabdosia rubescens were as the materials to research the impacts of different lead (Pb2+) concentrations(0, 135, 270, 540, 1 080 mg x L(-1)) on seed germination and seedling growth. The results show that: Low concentration of lead had no obvious effect on early germination of the seed, the germination vigor and germination speed were lightly higher but not significantly differed at the level of Pb concentration 135 mg x L(-1) with control group; Mid-high concentration of Pb solution (270-1 080 mg x L(-1)) significantly inhibited the seed germination and seedling growth, which reduced the seed germination rate, germination vigor, germination index, embryo root length and shoot length, growth index with increasing of Pb concentrations. There was a inhibitory effect on embryo shoot length and root length at mid-high lead concentrations stress, and stronger inhibitory effect on root , which was more sensitive than shoot to Pb stress(P < 0.05). Pb bioaccumulation coefficient (BC) was 0.76-2.59, increased with concentration of Pb; Pb enrichment in seedling mainly caused the growth inhibition. The fitting model predictive analyses show, the critical concentration of Pb, which causes the germination rate and biomass fresh weight reducing 10%, is 195.18, 101.65 mg x L(-1).

Study of achieving multiple models of plasma treatment by remoulding B.BRAUN Diapact CRRT / 国际生物医学工程杂志

Objective To study how to use the machine of B.BRAUN's Diapact continuous renal replacement therapy (CRRT) to achieve multiple models of plasma treatment by analyzing the characteristics of the machine.Methods Based on the machine of B.BRAUN's Diapact CRRT,assisted by extra single pump,the pipeline connection of the machine was reformed and the appropriate parameter was reset.Results After the reconstruction,the new equipment could succeed in achieving special plasma treatment such as double filtration plasmpheresis (DFPP)and plasma adsorption (PA).Conclusions The reconstructed equipment can attain special plasma treatment.Compared with plasma exchange (PE),the equipment has some characteristics such as few syndromes and without external plasma.According to its safety and stability,this new method is suitable for clinical application.

A high throughput coupled with high performance liquid chromatography-tandem mass spectrometry method for determination of aflatoxin B1, B2, G1, G2 in 10 traditional Chinese medicines / 中国中药杂志

As the dilution procedure was applied, a simple, rapid and cost-effective high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for determination of aflatoxin B1, B2, G1, and G2 was successfully by performed in a total 83 samples of 10 traditional Chinese medicines (TCMs), which were collected from 5 different hospital pharmacies and 5 different medical stores in Guangzhou city. Matrix effects of these 10 TCMs were ranged from 80.23% to 115.5% in low, intermediate and high concentration levels, indicating that the negative effect was overcome in this study. Meanwhile, the analysis method was proved to be stable and reliable during the whole analysis using Semen Armeniacae Amarum spiked 3 concentration levels of standard solution as quality control samples and the RSD < 6.6% was obtained. The contamination levels of 83 investigated samples were 13.89% and 17.02% in hospital pharmacies and medical stores, respectively. The result was presented to provide relevant reference and supplement to those researchers in TCMs analysis and screening.

Cross-validation of the osmotic pressure based on Pitzer model with air humidity osmometry at high concentration of ammonium sulfate solutions / 药学学报

The osmotic pressure of ammonium sulfate solutions has been measured by the well-established freezing point osmometry in dilute solutions and we recently reported air humidity osmometry in a much wider range of concentration. Air humidity osmometry cross-validated the theoretical calculations of osmotic pressure based on the Pitzer model at high concentrations by two one-sided test (TOST) of equivalence with multiple testing corrections, where no other experimental method could serve as a reference for comparison. Although more strict equivalence criteria were established between the measurements of freezing point osmometry and the calculations based on the Pitzer model at low concentration, air humidity osmometry is the only currently available osmometry applicable to high concentration, serves as an economic addition to standard osmometry.

Determination of aflatoxin B1, B2, G1, G2 in armeniacae semen amarum by high-performance liquid chromatography-tandem mass spectrometry / 中国中药杂志

A simple, rapid and cost-effective high-performance liquid chromatography-tandem mass spectrometry (LC-MS/ MS) method was established for simultaneous determination of aflatoxins (AFB1, AFB2, AFG1, AFG2) in Armeniacae Semen Amarum and the application was performance in 11 samples collected from different markets, medical stores and hospitals. The sample was extracted with 84% acetonitrile/water and 250 microL extraction was directly injected into a LC-MS/MS system without further purification procedure after being redissolved with methanol. The LC separation was performed on a C18 column with a linear gradient elution program of 4 mmol x L(-1) NH4 Ac-0.1% formic acid solution and menthol as the mobile phase. Selected reaction monitoring (SRM) was used for selective determination of the four aflatoxins on a triple quadruple mass spectrometer, which was operated in positive ionization modes. All the four aflatoxins showed a good linear relationship with r > 0.999 0, the average recoveries were between 87.88% and 102.9% and the matrix effect was ranged from 90.71% to 99.30% in low, intermediate and high levels. Furthermore, the higher recovery was obtained by the method reported in this study, comparing to the cleanup procedure with the Mycosep 226 purification column. Eleven samples collected were detected and the contamination levels of the AFB1 were between 1.590-2.340 microg x kg(-1) and the AF (B1 + B2 + G1 + G2) was ranged from 2.340 to 3.340 microg x kg(-1). In summary, the developed method was suitable to detect and screen AFB1, AFB2, AFG1, AFG2 in Armeniacae Semen Amarum.

Separation and molecular identification of fungal contamination on surface of 15 Chinese herbal medicines / 中国中药杂志

<p><b>OBJECTIVE</b>To evaluate fungal contamination on the surface of Chinese herbal medicines and explore an appropriate method for fast and efficient identification of contaminant fungi.</p><p><b>METHOD</b>Chinese herbal medicines were first washed and the washing solution was plated onto potato dextrose agar (PDA) to obtain the pure isolates. For molecular identification, two new pairs of specific primers were designed according to ITS region of fungi genome sequences. The strains were identified through polymerase chain reaction (PCR) and sequence analysis.</p><p><b>RESULT</b>Fifty fungal strains were obtained from the surface of 15 Chinese herbal medicines with the percent of contaminated samples of 93.3%. Twenty-seven strains among them were successfully identified.</p><p><b>CONCLUSION</b>Fungal contamination on the surface of Chinese herbal medicines is quite common. Although different fungal species were isolated, the genus Aspergillus was the predominant. The primer pairs developed in this study are compatible and can be used to identify fungal species from the surface of Chinese herbal medicines.</p>