[Cloning and expression analysis of U6 promoters in Panax quinquefolius].

The U6 promoter is an important element driving sgRNA transcription in the CRISPR/Cas9 system. Seven PqU6 promo-ter sequences were cloned from the gDNA of Panax quinquefolium, and the transcriptional activation ability of the seven promoters was studied. In this study, seven PqU6 promoter sequences with a length of about 1 300 bp were cloned from the adventitious roots of P. quinquefolium cultivated for 5 weeks. Bioinformatics tools were used to analyze the sequence characteristics of PqU6 promoters, and the fusion expression vectors of GUS gene driven by PqU6-P were constructed. Tobacco leaves were transformed by Agrobacterium tumefaciens-mediated method for activity detection. The seven PqU6 promoters were truncated from the 5'-end to reach 283, 287, 279, 289, 295, 289, and 283 bp, respectively. The vectors for detection of promoter activity were constructed with GUS as a reported gene and used to transform P. quinquefolium callus and tobacco leaves. The results showed that seven PqU6 promoter sequences(PqU6-1P to PqU6-7P) were cloned from the gDNA of P. quinquefolium, with the length ranged from 1 246 bp to 1 308 bp. Sequence comparison results showed that the seven PqU6 promoter sequences and the AtU6-P promoter all had USE and TATA boxes, which are essential elements affecting the transcriptional activity of the U6 promoter. The results of GUS staining and enzyme activity test showed that all the seven PqU6 promoters had transcriptional activity. The PqU6-7P with a length of 1 269 bp had the highest transcriptional activity, 1.31 times that of the positive control P-35S. When the seven PqU6 promoters were truncated from the 5'-end(PqU6-1PA to PqU6-7PA), their transcriptional activities were different in tobacco leaves and P. quinquefolium callus. The transcriptional activity of PqU6-7PA promoter(283 bp) was 1.59 times that of AtU6-P promoter(292 bp) when the recipient material was P. quinquefolium callus. The findings provide more ideal endogenous U6 promoters for CRISPR/Cas9 technology in ginseng and other medicinal plants.

Seven new 2-(2-phenylethyl) chromone derivatives from agarwood of Aquilaria agallocha with inhibitory effects on nitric oxide production.

Seven new 2-(2-Phenethyl) chromone derivatives (1-7), including four 2-(2-Phenethyl) chromones (1-4), one 6, 7, 8 trihydroxy-2-(2-Phenethyl) chromone (5), one acetylated 5, 6, 7, 8-tetrahydroxy-2-(2-Phenethyl) chromone (6), and one chlorine-containing 5, 6, 7, 8-tetrahydro-2-(2-Phenethyl) chromone (7), along with eight known compounds (8-15), were isolated from agarwood originating from Aquilaria agallocha Roxb.. Their structures were determined mainly by high-resolution electrospray ionization mass spectrometry (HR-ESI-MS) and nuclear magnetic resonance (NMR) analysis. The absolute configurations of 3-7 were resolved by electronic circular dichroism (ECD) calculations. Nearly all compounds were evaluated for their anti-inflammatory activities in RAW264.7 cells. Compounds 1 and 7-11 displayed significant anti-inflammatory activities with IC50 values ranging from 3.71 to 32.04 µM.

Three new C-glycosyflavones with acetyl substitutions from Swertia mileensis.

Three new acetylated C-glycosylflavones, 3″,6″-di-O-acetylswertiajaponin (1), 4″,6″-di-O-acetylswertiajaponin (2), and 6″-O-acetylswertiajaponin (3), together with six known compounds were isolated from the whole herb of Swertia mileensis. Their structures were elucidated on extensive NMR experiments and mass spectrometry studies. 1H and 13C NMR data exhibited doublet signals at room temperature. Variable temperature 1H NMR experiments were carried out to investigate the presence of rotational isomerism of C-glycosylflavones. All compounds showed potential antioxidant activities against apoptosis of H2O2-induced human embryo liver L02 cells.

Extractions of oil from Descurainia sophia seed using supercritical CO2, chemical compositions by GC-MS and evaluation of the anti-tussive, expectorant and anti-asthmatic activities.

Descurainia sophia is widely distributed in China and is one of the most troublesome annual weeds. It has diverse medicinal usage. D. sophia has abundant oil, making it an important oil plant in China. The main goal of this study was to obtain the maximum yield of the oil by an optimal selection of supercritical fluid extraction parameters. According to the central composite design and response surface methodology for supercritical fluid extraction method, a quadratic polynomial model was used to predict the yield of D. sophia seed oil. A series of runs was performed to assess the optimal extraction conditions. The results indicated that the extraction pressure had the greatest impact on oil yield within the range of the operating conditions studied. A total of approximately 67 compounds were separated in D. sophia seed oil by GC-MS, of which 51 compounds represented 98.21% of the total oils, for the first time. This study was also aimed at evaluating the anti-asthmatic, anti-tussive and expectorant activities in vivo of D. sophia seed oil which supplied for further research on bioactive constituents and pharmacological mechanisms.

A new kaempferol trioside from Silphium perfoliatum.

A new apiose-containing kaempferol trioside, kaempferol-3-O-α-L-rhamnosyl-(1‴ → 6″)-O-ß-D-galactopyranosyl-7-O-ß-D-apiofuranoside, along with 16 known compounds, were isolated from 50% acetone extract of Silphium perfoliatum L. Their structures were elucidated by acid hydrolysis and spectroscopic techniques including UV, IR, MS, ¹H, ¹³C, and 2D-NMR. In addition, the pharmacological activity of compound 1 was tested with HepG2 and Balb/c mice (splenic lymphocytes and thymic lymphocytes) in vitro, and it exhibited inhibitory effect on the proliferation of HepG2 cells and showed the immunosuppressive activity.