Advances in Mechanism Research on Polygonatum in Prevention and Treatment of Diabetes.

Diabetes mellitus is a fast-growing disease with a major influence on people's quality of life. Oral hypoglycemic drugs and insulin are currently the main effective drugs in the treatment of diabetes, but chronic consumption of these drugs has certain side effects. Polysaccharides, saponins, flavonoids, and phenolics are the primary secondary metabolites isolated from the rhizomes of Polygonatum sibiricum Redouté [Asparagaceae], Polygonatum kingianum Collett & Hemsl [Asparagaceae], or Polygonatum cyrtonema Hua [Asparagaceae], which have attracted much more attention owing to their unique therapeutic role in the treatment and prevention of diabetes. However, the research on the mechanism of these three Polygonatum spp. in diabetes has not been reviewed. This review provides a summary of the research progress of three Polygonatum spp. on diabetes and its complications, reveals the potential antidiabetic mechanism of three Polygonatum spp., and discusses the effect of different processed products of three Polygonatum spp. in treating diabetes, for the sake of a thorough understanding of its effects on the prevention and treatment of diabetes and diabetes complications.

Remediation of deterioration in microbial structure in continuous Pinellia ternata cropping soil by crop rotation.

Pinellia ternata is a traditional Chinese herb that suffers from continuous cropping (CC), which significantly decreases both yield and quality. The influence of CC on the microbiome in P. ternata rhizosphere and the effects of remediation on microbiota by rotational cropping (CR) were assessed by Illumina high-throughput sequencing technology. CC tends to decrease the α-diversities as a function of cultivation time, whereas CR tends to increase them. Differentially abundant analysis showed that microbial structure was important in maintaining the health status of P. ternata rhizosphere. Results suggested that CC soils were mainly enriched for Pseudomonas, Rhizobium, and Streptophyta operational taxonomic units (OTUs), while the CR soils were mainly enriched for Rhizobium, Pseudomonas, Flavobacterium, Sphingomonas, Rhizobacter, and Arthrobacter OTUs. On the basis of the community dissimilarities, we grouped all sample replicates into three post hoc clusters in which soils were defined as healthy, health-suppressed, and health-depressed soils. The three soil types represented different soil physicochemical properties. The activities of the microbiome features, including ammonia oxidizer, sulfate reducer, nitrite reducer, dehalogenation, xylan degrader, sulfide oxidizer, nitrogen fixation, atrazine metabolism, chitin degradation, degraded aromatic hydrocarbons, and chlorophenol degradation, were also considerably different among the three soils.

[Effect of different fertilization treatments on yield and secondary metabolites of Codonopsis pilosula].

The research studies the effect of different fertilization treatments on yield and accumulation of secondary metabolites of Codonopsis pilosula by using single factor randomized block design, in order to ensure reasonable harvesting time and fertilization ratio, and provide the basis for standardized cultivation of C. pilosula. According to the clustering results, the nitrogen fertilizer benefitted for the improvement of root diameter and biomass of C. pilosula. The phosphate fertilizer could promote the content of C. pilosula polysaccharide. The organic fertilizers could increase the content of lobetyolin. With the time going on, C. pilosula's yield, polysaccharide and ehanol-soluble extracts increased while the content of lobetyolin decreased. According to various factors, October is a more reasonable harvest period. Organic fertilizers are more helpful to the yield and accumulation of secondary metabolites of C. pilosula.

[Preparation and vitality detection of protoplast in Salvia miltiorrhiza Bunge].

We prepared protoplasts from Salvia miltiorrhiza Bunge suspension culture cells. Then, the protoplasts' vitality and functions were tested by fluorescein diacetate staining method and Fluo-3/AM flourescent probe. The optimal condition of protoplast isolation was Cellulase R-10 1.5%, Pectinase Y-23 0.3%, Macerozyme R-10 0.5%, 40 r/min 12 h, 600 r/min 5 min, and the protoplasts yield was 1.1x10(6) cells/g FW, the vitality was more than 95% by using fluorescein diacetate staining method. It has been confirmed that calcium fluorescent probe Fluo-3/AM can be successfully loaded into protoplasts.

[Effect of calcium on medium alkalinization induced by salicylic acid in Salvia miltiorrhiza suspension cultures].

We studied medium alkalinization in Salvia miltiorrhiza suspension cultures treated with salicylic acid and the effect of Ca2+ in this process through application of calcium channel antagonists (Verapamil, LaCl3, LiCl, 2-APB) and ionophore A23187. The results show that salicylic acid could induce significant medium alkalinization in S. miltiorrhiza culture. Verapamil and LaCl3 or LiCl and 2-APB, two different groups of calcium channel antagonist, significantly inhibited the medium alkalinization induced by salicylic acid. However, the suppression effect of verapamil or LaCl3 on medium alkalinization induced by salicylic acid was higher than that of LiCl or 2-APB. When two types of calcium channel inhibitor (LaCl3 and 2-APB) were used together, the medium alkalinization induced by salicylic acid was completely suppressed and even reduced the pH in medium. On the other hand, A23187 could promote the medium alkalinization. Based on the results above, we speculated that salicylic acid could induce significant medium alkalinization in S. miltiorrhiza culture, depending on the calcium from both extracell and intracell. Moreover, calcium from extracell plays a more dominant role in this process. Reveal of relationship in this research between Ca2+ and medium alkalinization can provide theory evidence for mechanism of the plant secondary metabolism.

[Effects of Ca2+ on salicylic-acid induced biosynthesis of salvianolic acid B in young seedlings of Salvia miltiorrhiza Bunge].

In order to study the effects of Ca2+ in the biosynthesis of salvianolic acid B (Sal B) induced by salicylic acid (SA) in the young seedlings of Salvia miltiorrhiza, we used confocal laser scanning microscopy and high performance liquid chromatography to measure the change of relative fluorescence intensity of Ca2+ and the contents of Sal B induced by SA before and after the application of extracellular calcium channel inhibitors (VP and LaCl3), intracellular calcium channel inhibitor (LiCl), as well as intracellular calmodulin antagonist (TFP). SA could induce the calcium burst, and the Ca2+ peak could last to 2-3 min in the guard cells of S. miltiorrhiza, which prompted the biosynthesis of Sal B after the Ca2+ burst. Both Vp or LaCl3, and LiCl or TFP could inhibit the burst of Ca2+ and the biosynthesis of Sal B. The above results demonstrated that Ca2+ from the extracellular and the intracellular calcium store regulate the biosynthesis of Sal B elicited by salicylic acid in S. miltiorrhiz young seedlings.

[Regulation effects of intracellular and extracellular Ca2+ on biosynthesis of rosmarinic acid induced by salicylic acid in young seedlings of Salvia miltiorrhiza].

OBJECTIVE: To investigate the effect of intracellular and extracellular Ca2+ on the biosynthesis of rosmarinic acid (RA) induced by salicylic acid in young seedlings of Salvia miltiorrhiza. METHOD: Young seedlings of S. miltiorrhiza were used to select an optimal concentration of salicylic acid (SA), and then use the optimal concentration of SA to investigate the effects of extracellular Ca2+ channel inhibitors Verapamil, LaCl3, intracelluar calmodulin antagonist TFP and intracelluar Ca2+ channel inhibitors LiCl on the biosynthesis of RA and related enzymes. RESULT: SA increased the accumulation of RA and the activities of PAL and TAT, especially the SA of 2 mmol x L(-1) after 24 h. SA improved the accumulation of RA to (40.51 +/- 2.16) mg x g(-1), which was 1.97 times than that of control, and the activities of PAL, TAT were 1.42 times and 1.29 times than those of the control. However, Vp, LaCl3, TFP, LiCl inhibited the effects of SA evidently. CONCLUSION: Ca2+ plays a key role in the regulation of the induction process.

[Hydrogen peroxide is involved in the signal transduction of salicylic acid-induced salvianolic acid B biosynthesis in Salvia miltiorrhiza cell cultures].

Hydrogen peroxide (H2O2), one of reactive oxygen species, is widely generated in many biological systems, and it mediates various physiological and biochemical process in plants. To investigate the role of H2O2 as a signaling molecule in the process of salicylic acid (SA)-induced Salvianolic acid B (Sal B) accumulation, we separately inspected the cultured cells of Salvia miltiorrhiza with SA, H2O2, catalase (CAT), 2-(4-carboxy-2-phenyl)-4,4,5,5-tetramethylimidazoline-l-oxyl-3-oxide (DMTU) and Imidazole (IMD) to investigate the influence on the activity of phenylalanine ammonia-lyase (PAL) and tyrosine aminotransferase (TAT) and the accumulation of Sal B. Treatment of S. miltiorrhiza cells with SA resulted in an increase of H2O2, the increase of PAL and TAT and accumulation of Sal B. Exogenous application of 10-30 mmol/L H2O2 was found to effectively increase PAL and TAT activity as well as the Sal B content. CAT, a H2O2 scavenger, eliminated the Sal B-accumulating effects of exogenous H2O2 and SA. These indicated that H2O2 may serve as an upstream signaling molecule in the SA-induced accumulation of Sal B signal transduction pathway. Disposed by DMTU, a chemical trap for H2O2, as observed to be effective in inhibiting SA-induced accumulation of Sal B. IMD strongly inhibits the activity of NADPH oxidase, which is one of the main sources of H2O2 formation in plant cells. IMD treatment strongly inhibited the accumulation of Sal B in cultured cells of S. miltiorrhiza, but the effects of IMD, can be partially reversed by the exogenous SA. The accumulation of Sal B was blocked once the generation of H2O2 by NADPH oxidase was inhibited, and H2O2 served as signaling molecule mediated the SA-induced Sal B accumulation.

[Effects of salicylic acid on synthesis of rosmarinic acid and related enzymes in the suspension cultures of Salvia miltiorrhiza].

Rosmarinic acid (RA), a phenolic acid, is one of the important secondary metabolites produced in Salvia miltiorrhiza. To observe the influence of salicylic acid (SA), an elicitor, on the synthesis of RA and related enzymes, we treated the cell suspension cultures of S. miltiorrhiza with SA and L-a-aminooxy-beta-phenylpropionic acid (AOPP), a competitive inhibitor of tyrosine aminotransferase (TAT). Under this condition, the activities of related enzymes, such as phenylalanine ammonia-lyase and TAT were traced and assayed; the accumulative amount of RA was measured. The results showed that the PAL activity reached the peak at 4 h, 124% higher than that of the control, and the content of RA reached its maximum ((5.914 +/- 0.296) mg/g dry weight) at 8 h, after treated by 6.25 mg/L SA on day 6 of the suspension culture. The results of treatment with 0.1 micromol/L AOPP showed that AOPP affected little on the TAT activity, while the PAL activity was significantly influenced, with 44% lower than that of the control at 6 h. Meanwhile, the reduced accumulation of RA ((4.709 +/- 0.204) mg/g dry weight) paralleled with the decrease in PAL activity. The co-treatment by 0.1 micromol/L AOPP and 6.25 mg/L SA relieved the restriction imposed by AOPP on PAL, and made the cell cultures accumulate more RA than sole treatment with AOPP, indicated that SA induced the accumulation of RA in suspension cell culture of S. miltiorrhiza, and the rate-limiting effect of PAL was stronger than TAT.

[Effects of calcium on synthesis of rosmarinic acid and related enzymes in suspension cultures of Salvia miltiorrhiza].

We studied the influence of the concentration of Ca2+ (0-50 mmol/L) in culture medium on the synthesis of rosmarinic acid (RA) and related enzymes in Salvia miltiorrhiza suspension cultures. Using verpamil (VP, a calcium channel antagonist) and ionophore A23187, we studied the mechanism of secondary metabolites of Salvia miltiorrhiza suspension cultures influenced by the concentration of Ca2+ in the culture medium. The synthesis of intracellular RA in 6-day incubation was significantly dependent on the medium Ca2+ concentration. At the optimal Ca2+ concentration of 10 mmol/L, a maximal RA content of 20.149 mg/g biomass dry weight was reached, which was about 37.3% and 20.4% higher than that at Ca2+ concentrations of 1 and 3 mmol/L, respectively. The variation of the activity of PAL and TAT, two key enzymes of the two branches of RA, could be affected by the concentration of Ca2+ in culture medium. The change of their activity occurred prior to the accumulation of RA, which suggested both of the key enzymes be involved in the synthesis of RA. Meanwhile, the enzymatic action of PAL was more distinct than TAT. The treatment of VP and A23187, respectively, indicated that the influence of RA affected by the concentration of Ca2+ in the culture medium was accomplished by the intracellular Ca2+, and the flow of Ca2+ from the extracellular to the intracellular environment could also participate in this process.

[Distributional difference of fat-soluble compounds in the roots, stems and leaves of four Salvia plants].

To illustrate distribution of fat-soluble compounds in the roots, stems and leaves of four Salvia plants, the methods of Histochemistry and HPLC were adopted to analyze different parts of the four Salvia plants in this paper. The results showed that distribution was differential, and following as this: the roots, stems and leaves of four Salvia plants contained fat-soluble compounds, moreover, the fat-soluble compounds of the roots located in periderm and the stems and leaves in epidermis. The main components of the fat-soluble compounds were Tanshinone IIA, Tanshinone I and Dihydrotanshinone I in the toots of Salvia miltiorrhiza Bunge and Salvia miltiorrhiza bge. f. alba, yet there were only Tanshinone IIA in the roots of Salvia japonica and Salvia officinalis. And fat-soluble compounds were not Tanshinone IIA, Tanshinone I and Dihydrotanshinone I in the stems and leaves of four Salvia plants. The type and content of fat-soluble compounds related to the species and introduction regions, they changed with the species and introduction regions. The conclusion clarified the accurate distribution of fat-soluble compounds in the different parts of four Salvia plants, and provided some theoretical basis for the application of Chinese herbs.

[Effect of water stress on growth and accumulation of active components of Isatis indigotica].

OBJECTIVE: To study the effect of water stress on the growth and accumulation of active components of Isatis indigotica. METHOD: By pot culture experiment,the characteristic indexes of water requirement, chlorophyll content, transpiration rate, photosynthesis, biomass and effective ingredient indirubin were studied under normal and water stress conditions, respectively. RESULT: With the increase of the extent and duration of water stress, the quantity of water requirement, chlorophyll content, respiration rate, photosynthesis and biomass of I. indigotica were decreased, while water use efficiency increased. CONCLUSION: It is a critical period of water requirement in July, high water content in soil and extreme water stress are not propitious to the accumulation of indirubin, appropriate water stress can stimulate indirubin accumulatio. The harvest of leaves of I. indigotica is optimal in July, while the harvest of root is appropriate in October. Yield and quantity of I. indigotica are improved when the field capacity is between 45% and 70%.

[Effective constitutents contents in indigowood roots and leaves from different regions].

By the method of HPLC, and with indigo and indirubin as indicative constituents, this paper determined the effective constituents contents in indigowoad roots and leaves collected from different regions. The results showed that the total content of effective constituents in indigowoad root was the highest (16.27 mg x kg(-1)) from Hanyin of Shaanxi Province, followed by Longde of Ningxia Autonomous Region (15.67 mg x kg(-1)), Bozhou of Anhui Province (14.90 mg x kg(-1)), and Linquan of Anhui Province and Jiamusi of Heilongjiang Province (13.97 mg x kg(-1)), and that in indigowoad leaf had the same order, with the values being 698.32, 683.68, 680.11, 654.19 and 642.23 mg x kg(-1), respectively. The HPLC chromatograms of leaf samples collected from different regions were differed, not only in the test constituents, but also in the other constituents manifested as various peaks in the chromatogram. It was suggested that besides meteorological factors, the selection of geo-authentic region should be also involved in the cultivation of Isatis indigotica.