Optimization of Enzymatic Extraction Process of Polysaccharides from Pseudostellaria heterophylla Fibrous Roots by Response Surface Methodology and Its Pilot Application

[Objectives]To study and optimize the process conditions of enzymatic hydrolysis technology for extracting polysaccharides from Pseudostellaria heterophylla fibrous roots and its application in workshop pilot tests.[Methods]P.heterophylla fibrous roots were taken as the matrix material,and Box Behnken design was used to analyze the extraction time,composite enzyme addition amount,and liquid-solid ratio for response surface optimization experiments,and then applied to the pilot extraction of P.heterophylla fibrous roots.[Results]Response surface analysis showed that all factors had a significant impact on the experimental indicators.The optimal extraction process conditions for polysaccharides from P.heterophylla fibrous roots were extraction time of 2.7 h,compound enzyme addition of 2.5%,and liquid-solid ratio of 32.The yield of polysaccharides from P.heterophylla fibrous roots was 4.83%.The water extraction process of P.heterophylla fibrous roots extraction pilot was used as the control group for response surface optimization of the pilot experiment.The optimization results showed that the extraction time was 3 h,the amount of composite enzyme added was 2.5%,and the liquid-solid ratio was 28.The polysaccharide yield was 4.75%,an increase of 4.63%compared to the control group.[Conclusions]This paper could provide feasibility for the innovation of enzy-matic hydrolysis technology for P.heterophylla fibrous roots and its workshop pilot practice application,as well as a reference for the industrial application of its medicinal resources.

Study on Pathogen and Bio-control of Dampingoff Disease in Pseudostellaria heterophylla in Qiandongnan Prefecture of Guizhou Province

Fungal diseases occur seriously in Pseudostellaria heterophylla in Qian- dongnan Prefecture of Guizhou Province from March to May every year. By field in- vestigation, tissue isolation and microscopic identification, the main pathogen was iden- tified as Rhizoctonia solanL the pathogen of damping-off disease. At the same time, Alternaria alternate was also isolated from the sampled diseased soil, and it has pathogenicity to Pseudostellaria heterophylla seedlings. The bio-control test showed that GH18 strain （Bacillus spp.） had significant inhibitory effects on Rhizoctonia solani and Alternaria alternate.

Inorganic Elements in Tuber of Chinese Herbal Medicine Radix Pseudostellariae from Guizhou Province

By analyzing the contents of inorganic elements in Radix Pseudostellariae from Guizhou Province, the aim was to assess the present quality of Radix Pseu- dostellariae, set limited standards of heavy metals, establish element fingerprints, and find out the characteristic elements. ICP-MS was used to measure the content of inorganic elements and map the element fingerprints. Additionally, WM/T2-2004 was applied to evaluate the quality of heavy metal elements, and the characteristic elements were determined by principal component analysis. The results showed that the contents of inorganic elements in Radix Pseudostellariae were between 0.057 and 959 mg/kg with the coefficients of variation ranging from 0.134 to 1.478, and the contents of Cd, As, Pb, and Hg were below the Standard of WM/T2-2004 in 90% of Radix Pseudostellariae. The standard limits of heavy metals in Radix Pseu- dostellariae were Cr≤6.5 mg/kg, Cu≤10 mg/kg, As≤2.0 mg /kg, Cd≤0.3 mg/kg, Hg≤0.15 mg/kg, and Pb≤4.0 mg/kg. The features of the inorganic eJements finger- prints could provide theoretical basis of identifying the quality of Radix Pseudostel- lariae and distinguishing Radix Pseudostellariae from other Chinese herbal medicines. The characteristic inorganic elements of Radix Pseudostellariae were found to be Cd, Cu, Co, Zn, Fe, Ca, Mg, and AI. Radix Pseudostellariae from Guizhou Province contained abundant inorganic elements, and the contents of heavy metals were below the evaluation criterion. The study provided a reference for the future development of the limiting values of heaw metals in Radix Pseudostellariae.

In Vitro Probiotic Effect of Pseudostellaria heterophylla Fibrous Roots on Feed Bacillus subtilis and Preliminary Study on Its Solid Fermentation

Traditional Chinese medicine Pseudostellaria heterophylla fibrous roots have been widely studied and applied in non-antibiotic clinical breeding of livestock and poultry. In vitro probiotic effect of P. heterophylla fibrous roots extract on feed probiotics Bacillus subtilis BD-K010 was studied, and the feasibility of solid fermentation of P. heterophylla fibrous roots was preliminarily evaluated. For in vitro probiotic effect, the increased concentration of P. heterophylla fibrous roots extract dependently increased the total biomass of B. subtilis BD-K010;1.0% P. heterophylla fibrous roots extract received the best effect, and the final p H of 1.0% experimental group was closer to neutral. Meantime, B. subtilis BD-K010 with optimum concentration of P. heterophylla fibrous roots showed significantly higher in vitro antibacterial effect than the control group(P<0.01),and the antibacterial effects on Escherichia coli and Staphylococcus aureus were improved by 51.99% and 63.16%, but it was ineffective to Salmonella. For solid fermentation, the profile of substrate complex and appendage flocculent structure on substrate surface at the end of fermenta-tion in experimental groups added with B. subtilis BD-K010 and cellulase plus BD-K010 were more complex;the live bacteria number, polysaccharide content and saponin content at fermentation end-point in two experimental groups were extremely higher than those in the control group(P<0.01).P. heterophylla fibrous roots extract had good in vitro probiotic effect on B. subtilis BD-K010 and promoted its antibacterial effect, and it is feasible to use probiotics for solid fermentation of P. heterophylla fibrous roots and to improve effective components. It is of great significance to further de-velop and utilize P. heterophylla fibrous roots resources in modern animal husbandry.

太子参(PSEUDOSTELLARIA HETEROPHYLLA)病毒病病原鉴定

太子参(Pseudostellaria heterophylla)是我国传统的中药材之一,其病毒病十分严重,田间发病率和种根带毒率可达100％,平均90％左右。自太子参病株分离得到的T-17、T-32、T-69和T-01四个病毒分离物,经鉴定,它们分别为芜菁花叶病毒(Turnip mosaicvirus,TuMV)、黄瓜花叶病毒(Cucumber mosaic virus,CMV)、蚕豆萎蔫病毒(Broad beanwilt virus,BBWV)和烟草花叶病毒(Tobacco mosaic virus,TMV)。其中以TuMV分布最广,为害最重,是太子参病毒病的主要病原。

HETEROPHYLLIN-A AND B,TWO CYCLOPEPTIDES,FROM THE ROOTS OF PSEUDOSTELLARIA HETEROPHYLLA

Two cyclopeptides,heterophyllin A and B,have been isolated from the roots of Pseudostellaria heterophylla.Their structures were elucidated by chemical, spectroscopic,and enzymatic methods.

Effects of Different Extract of Pseudostellaria Heterophylla on Immunological Function in Mice based on Meta-analysis and Network Meta-analysis

To evaluate the effects of different extract of Pseudostellaria heterophylla on immunological function in mice based on Meta-analysis and Network meta-analysis,the article retrieved domestic and foreign databases according to the"PICO"retrieval strategy,and used Stata and ADDIS software for Meta analysis.A total of 6 reports,10 randomised controlled trials(RCTs)were eventually involved.Meta analysis results showed that:compared with the control group,the experimental group of polysaccharide,saponin and crude extract were better than that of the control(P<0.05),which significantly improved the immunological function in mice.Network meta-analysis results showed that the saponin had the best effect on the increase of phagocytic index and the differences were statistically significant[MD=1.50,95%CI(0.71,2.28),P<0.05];The polysaccharide and saponin had better effect on the increase of spleen index than the control and crude extract,and the differences were statistically significant[MD=0.77,95%CI(0.27,1.31),P<0.05],[MD=0.71,95%CI(0.01,1.50),P<0.05];the polysaccharide had the best effect on the increase of thymus index and the differences were statistically significant[MD=0.70,95%CI(0.11,1.34),P<0.05].The rank probability showed that the saponin of Pseudostellaria heterophylla had the maximum probability to increase phagocytic index of mice;the probability for the components of Pseudostellaria heterophylla increasing spleen index of mice was in the order of crude extract>polysaccharide>saponin;the probability for the components of Pseudostellaria heterophylla increasing thymus index of mice was in the order of saponin>polysaccharide>crude extract.Based on the available evidence,the extract of Pseudostellaria heterophylla could improve the immunity of mice,and the clinical effect of polysaccharide and saponin was the best,which provided a more valuable scientific reference for evidencing that the polysaccharide and saponin of Pseudostellaria heterophylla was the effective components for improving immunological function,and also was conducive to the proper further development of Pseudostellaria heterophylla resources.

Optimization of Steam Explosion Process Condition for Extracting Polysaccharides from Pseudostellaria heterophylla by Response Surface Methodology

【Objective】To optimize the steam explosion process condition for extracting polysaccharides from Pseudostellaria heterophylla.【Method】The effects of steam pressure,pressure-maintaining time and material moisture content on the extraction of polysaccharides from Pseudostellaria heterophylla were studied by response surface methodology based on Box-Behnken design.【Result】The findings showed that each factor could significantly affect the test index,and the optimum condition was as follows:steam pressure 1.50 Mpa,pressuremaintaining time 46 s and material moisture content 46%.Under this condition,the verified experimental value of polysaccharides from Pseudostellaria heterophylla was 39.32%,indicating a relative standard deviation of 2.73%from the predictive value.Meanwhile,scanning electron microcopy(SEM)images showed that the surface physical structure of Pseudostellaria heterophylla was irregularly broken and cracked,which means the physical structure of Pseudostellaria heterophylla was changed and destroyed at the cellular level.【Conclusion】This experiment provides a new approach for the extraction of polysaccharides from Pseudostellaria heterophylla,as well as a reference for the resource utilization of Pseudostellaria heterophylla.

Reevaluation on Antioxidant Activity of Tonic Chinese Herbs Pseudostellaria heterophylla

Tonic Chinese herbs Pseudostellaria heterophylla has complex components,which has both medicinal and nutritional effects,and the clinical research on anti-oxidation of P.heterophylla has become a hotspot.By searching domestic and foreign databases,the antioxidant literatures of P.heterophylla were searched,consulted,sorted,classified and analyzed,and systematic reevaluation of antioxidant function was conducted from three aspects of P.heterophylla extract in vitro chemical method,biological method and animal test method according to evaluation criteria for antioxidant activity of traditional Chinese medicine and Meta analysis statistical methods.The results would provide systematic reference for further research and development of P.heterophylla,especially for forage value excavation and development of P.heterophylla in the field of modern animal breeding.

Investigation and Green Prevention and Control Strategies of Pseudostellaria heterophylla Diseases in Genuine Producing Area in Zherong County

Zherong County of Fujian Province is a genuine producing area of Pseudostellaria heterophylla,and disease has become a main factor restricting the healthy development of P.heterophylla industry in the county.In order to scientifically understand the occurrence status of agricultural diseases and serve the GAP of P.heterophylla,P.heterophylla diseases in Zherong County were investigated,and the information and regularity of diseases were summarized.P.heterophylla diseases in Zherong County were infectious,and caused by fungi and viruses.The diseases occurred frequently in spring and summer,and symptoms were manifested in pathological changes of leaf and tuber.Eventually,green prevention and control strategies of diseases were put forward:cultivating new disease-resistant varieties,scientifically administrating drugs according to symptoms,administrating drugs in optimum period,strengthening biological control,strictly requiring drug safety period and paying attention to green prevention and control during the planting process.The occurrence of P.heterophylla diseases in Zherong County was investigated and summarized,and the corresponding strategies were put forward,so as to scientifically serve the agricultural industry in P.heterophylla genuine producing area,and to provide a basis for the sustainable development of modern pollution-free planting industry of P.heterophylla.

Effects of Panax ginseng,Codonopsis pilosula and Pseudostellaria heterophylla on Sexual Maturity of SD Female Rats

[Objectives] To study the effects of Panax ginseng,Codonopsis pilosula and Pseudostellaria heterophylla on the growth and development of SD female rats. [Methods] The rats were randomly divided into control group,P. ginseng group,C. pilosula group,and P. heterophylla group,intragastrically administered with physiological saline,P. ginseng,C. pilosula,and P. heterophylla for 2 weeks,daily vaginal smear inspection was performed,estradiol( E_2) level before and after intervention was detected,rats were dissected,took out ovary and uterus,and calculated the ovarian index and uterus index. [Results]In 4 groups of rats,there was no significant difference in the time of sexual maturity and ovarian index; compared with the control group,E_2 level of C. pilosula group and P. ginseng group were higher( P < 0. 05);compared with the control group and P. heterophylla group,the ovarian index of P. ginseng group was higher( P < 0. 05); compared with the control group,the body weight of three administration groups increased much more( P < 0. 05). [Conclusions] P. ginseng and C. pilosula may lead to sexual maturity of SD female rats,while P. ginseng,C. pilosula and P. heterophylla can promote increase of body weight of SD female rats.

Effects of Different Concentrations of Radix pseudostellariae Fibrous Root Extracts on Serum Cytokines in Mice and Their Correlation Analysis

To investigate the effects of Radix pseudostellariae fibrous root extracts(RPFRE) on serum cytokines in normal and immunosuppressed mice and the correlation between indexes, KM male mice were randomly divided into four groups, which were given 0(control), 0.1, 0.2, 0.4 g/kg RPFRE by gavage. The levels of IL-2, IL-4, IL-6, and IFN-γ in the serum of mice were measured 14 d after gavage. On the 15day, the remaining mice in the experimental group were intraperitoneally injected with cyclophosphamide(CY), and samples were collected on the 18day to measure serum cytokines levels. The results showed that RPFRE improved the serum cytokine level in normal mice. 0.1 g/kg RPFRE significantly increased the content of IL-2and IFN-γ(P<0.05), while 0.4 g/kg RPFRE significantly increased the content of IL-4 and IL-6(P<0.05). Correlation analysis showed that serum cytokines IL-4 and IL-6 in normal mice had an extremely significant positive correlation with RPFRE concentration(P<0.01), IL-2 had a significant or extremely significant positive correlation with IL-4, IL-6, and IFN-γ(P<0.01or P<0.05), and IL-4 had an extremely significant positive correlation with IL-6(P<0.01). After CY injection, serum cytokine levels in mice decreased and immune function was inhibited. Compared with the control group, the experimental group recovered the level of cytokines to some extent. 0.1 g/kg RPFRE significantly increased the content of IL-2 in immunosuppressed mice(P<0.05), and 0.4 g/kg RPFRE significantly increased the content of IL-4, IL-6, and IFN-γ in immunosuppressed mice(P<0.05). Correlation analysis showed that the serum cytokines in immunosuppressed mice were significantly positively correlated with RPFRE concentration(P<0.01), and there was a certain dose dependence. Moreover, the correlation coefficient between IL-2 and IL-4, IL-6, and IFN-γ, IL-4 and IL-6, and IL-6 and IFN-γ reached an extremely significant level(P<0.01). To sum up, RPFRE could improve the serum cytokines level of mice, promote the secretion of cytokines, and have a certain immune protective effect. There was a significant positive correlation between the RPFRE concentration and the serum cytokines in mice, which provided basic data support for the further study of RPFRE immune function.

Key Points for High-yield Cultivation Management of Pseudostellaria heterophylla in Weihai Area

[Objectives] To explore the scientific cultivation mode of Pseudostellaria heterophylla in Shandong Province. [Methods] According to the ecological characteristics of P. heterophylla, the cultivation techniques and management points of P. heterophylla in Weihai are summarized. [Results] The scientific cultivation techniques and management points of P. heterophylla are introduced from the aspects of land selection, land preparation, seed selection, seedling nursing, timely sowing and transplanting, field management and disease and pest control. [Conclusions] This study will provide theoretical guidance for the high-quality, high-yield and high-efficiency cultivation of P. heterophylla in Shandong area.

Saponins Content in Fibrous Roots of Pseudostellaria heterophylla and Optimization of the SEP Extraction Technology

[Objectives]To explore the saponins resources in fibrous roots of Pseudostellaria heterophylla and optimize the SEP technology for extracting saponins.[Methods]Saponins contents in fibrous roots and root tubers of P.heterophylla were compared;with the fibrous roots as host materials,this experiment applied Box-Behnken design to analyze the influence of steam pressure,maintenance pressure time,moisture content of host materials on the content of saponins extracts.[Results]Saponins content was higher in fibrous roots than in tuberous roots of P.heterophylla;the response surface optimization showed that SEP(steam explosion pretreatment)promoted the contents of saponins and extracts in fibrous roots,and the optimal extracting conditions were stream pressure 1.50 Mpa,maintenance pressure time 54 s,and moisture content of host materials 30%.Under this extracting conditions,the verification value of saponins content experiment was 0.441%,36.53%higher than the untreated control group.[Conclusions]This research offered a new method of exploring saponins resources from fibrous roots of P.heterophylla,and provided a reference for developing new medicinal parts of P.heterophylla.

Analysis of the mechanism of Radix Astragali-Radix Pseudostellariae in the treatment of chronic heart failure based on network pharmacology

Objective:To explore and analyze the effective component,target and mechanism of action of Radix Astragali-Radix Pseudostellariae in the treatment of chronic heart failure(CHF)by network pharmacological research methods.Methods:With the help of TCMSP database,the effective chemical constituents and targets of Radix Astragali-Radix Pseudostellariae were screened out,and CHF-related proteins were obtained by Gene Cards and OMIM database.The target protein interaction network(PPI)was constructed on the String database platform,and using DAVID database to conduct the GO function and KEGG pathway enrichment analysis on the target of the obtained astragalus-pseudostellariae for the treatment of CHF.Results:Radix Astragali-Radix Pseudostellariae were selected as 28 effective chemical components and 104 therapeutic targets for CHF.There were obtained 49 GO functional annotations and 86 KEGG signaling pathways.Conclusion:Radix Astragali-Radix Pseudostellariae can treat CHF through multi-target and multi-pathway.

Immunoprotection Effects of Radix Pseudostellariae Fibrous Root Extraction (RPFRE) on Mice and Its Relationship with Antioxidant Function

[Objective] The paper was to explore the immunoprotection effects of Radix Pseudostellariae fibrous root extraction(RPFRE) on experi-mental mice and its relationship with antioxidant function. [Method] KM male mice were randomly divided into 4 groups based on RPFRE gradient concentration, and the immune function indexes and antioxidant indexes of mice were determined at 14 d post intragastric administration of RPFRE.The remaining mice in the experimental group were intraperitoneally injected with cyclophosphamide(CY) from the 15^(th) to 17^(th) day, and samples were collected to determine the above indexes on the 18^(th) day. [Result] Intragastric administration of RPFRE for 14 d improved the spleen index,thymus index, T/B cell stimulation index and total antioxidant capacity(T-AOC) of normal mice, and extremely increased the thymus index and TAOC(P<0.01). The T-AOC had extremely positive correlation with the spleen index, thymus index and B cell stimulation index( P<0.01), and had significantly positive correlation with the T cell stimulation index(P<0.05). From the 15^(th) to 17^(th) day of the experiment, the immune function indexes and antioxidant indexes of mice decreased and the malondialdehyde(MDA) content increased. The experimental groups of 0.1, 0.2 and 0.4 g/kg RPFRE promoted the immunoprotection and antioxidant effect of mice. RPFRE concentration had extremely positive correlation with 4 indexes of immune function and antioxidant indexes T-AOC and SOD(P<0.01), but had extremely negative correlation with MDA(P<0.01). There was a significantly positive correlation between immune function indexes and antioxidant indexes of mice after CY intervention( P<0.05 or P<0.01). [Conclusion] RPFRE can improve the immune function indexes and antioxidant indexes of normal mice and immunosuppressed mice and has immunopro-tection effect on immunosuppressed mice, and there is a significant correlation between immune function indexes and antioxidant indexes of mice.

Study on the Mechanism of Pseudostellariae Radix in Regulating Angiogenesis Based on Network Pharmacology and a Dual-screening System

[Objectives]This study was conducted to clarify the action mechanism of Pseudostellariae Radix in regulating angiogenesis by using network pharmacology and a dual-screening system,and to provide a basis for its clinical treatment of cardiovascular diseases.[Methods]The TCMSP database was used for preliminary screening to obtain the active compounds of Pseudostellariae Radix and the protein targets of its action.GeneCards and OMIM databases were used to search for targets related to angiogenesis.Cytoscape 3.9.1 was used to construct a drug-target network and protein interaction network of Pseudostellariae Radix in angiogenesis.The GO enrichment analysis and KEGG pathway analysis of the targets of Pseudostellariae Radix in angiogenesis were carried out on Metascape platform.The effects of the screened active compounds were verified using a dual-screening system.[Results]Six active components of Pseudostellariae Radix,luteolin,acetin,beta-sitosterol,linarin,schottenol and 1-monolinolein,were screened by TCMSP database;and the six active components were predicted with 78 common target proteins related to angiogenesis,of which 19 were core targets.Pseudostellariae Radix mainly intervened in angiogenesis through domain specific binding,ubiquitin-like protein ligase binding,kinase binding and other molecular functions to regulate biological processes such as membrane microdomain,plasma membrane raft and caveola.The results of KEGG enrichment indicated that pathways in cancer,lipid and atherosclerosis,hepatitis B,apoptosis,toxoplasmosis and other key pathways might be the mechanism for the intervention of angiogenesis.The results of the dual-screening system showed that luteolin,acacetin,beta-sitosterol and linarin protected HUVECs and promoted zebrafish angiogenesis.[Conclusions]This study preliminarily demonstrated that luteolin,acacetin,beta-sitosterol and linarin could intervene in angiogenesis through multiple targets and multiple pathways,providing ideas and a scientific basis for the treatment of cardiovascular diseases.

Pseudostellaria heterophylla:Antioxidant Activity and Its Mechanism of Action

The tonic traditional Chinese medicine Pseudostellaria heterophylla both medicinal and nutritional effects.It has been applied to research in the field of modern aquaculture,and its anti-oxidant clinical trials have also become a hot spot.Through searching domestic and foreign literature databases to sort out related literature,the anti-oxidation property of P.heterophylla was reviewed systematically from the three levels of in vitro chemical method,biological method and animal experiment method.On this basis,the functional mechanism of the anti-oxidant activity of P.heterophylla explored,in order to provide a reference for the follow-up in-depth research on the anti-oxidation of P.heterophylla and the discovery and development of feed and health care value of P.heterophylla the field of modern animal breeding.

世界孩儿参属(Pseudostellaria Pax)的分类纲要及地理分布

在标本和文献综合分析的基础上,对世界范围内孩儿参属(Pseudostellaria Pax)植物的主要形态特征进行了总结,依据Mizushima的分类系统编制了孩儿参属的分类检索表,对5个存疑种进行了初步分类处理,并对孩儿参属各种类的分布状况进行了绘图和分析。结果表明:具有肉质块根、绝大多数种具有闭锁花以及开放花花瓣常呈2浅裂是孩儿参属区别于石竹科(Caryophyllaceae)其他属的典型特征。初步确认全世界的孩儿参属植物共22种,被分成Sect.Pseudostellaria和Sect.Mamillatae 2个组,其中,前者仅包含石生孩儿参〔P.rupestris(Turcz.)Pax〕1种,而后者则被进一步分成Ser.Mamillatae和Ser.Distantes 2个系。本研究结果不支持将P.sierrae Rabeler et R.L.Hartm.和P.oxyphylla(B.L.Rob.)R.L.Hartm.et Rabeler归入孩儿参属的观点,并建议将矮小孩儿参〔P.maximowicziana(Franch.et Sav.)Pax〕、须弥孩儿参〔P.himalaica(Franch.)Pax〕和异花孩儿参〔P.heterantha(Maxim.)Pax〕分别独立成种。从分布区域看,孩儿参属呈北温带间断分布,由北美洲、地中海地区和东亚3个相互隔离的地理区域组成,并且,东亚为其多样化分布中心。

Determination of Polysaccharide in Radix pseudostellariae Extract by Size-Exclusion High-Performance Liquid Chromatography

A simple size-exclusion high-performance liquid chromatographic method was developed for rapid molecular mass screening to determine the average molecular mass, polydispersity, and quantity of Taizishen polysaccharides extract. The screening used a TOSOH TSK-GEL G3000SW high-performance liquid chromatographic column with 0.1 mol/L NaNO3 buffer as the mobile phase and refractive index detector. The molecular mass calibration curve was linear for polysaccharide standards from 10 to 100 kDa with a correlation coefficient of 0.9991. The method can be used to analyze the quantity, average molecular mass, and polydispersity of polysaccharides. In addition, the screening method is suitable for quality control of polysaccharide preparations in Chinese medicines and pharmaceutical products.