Determination of harpagide and harpagoside contents in Scrophularia ningpoensis from 15 origins / 药学实践杂志

Objective To establish an HPLC method for the simultaneous determination of harpagide and harpagoside content in Scrophularia ningpoensis (SN). Methods An Eclipse C18 column was used for determination of methanol extract of S. ningpoensis with a HPLC-PDA method and mobile phase of acetonitrile-0.03% phosphate solution in a gradient elution manner. The flow rate of mobile phase was 1.0 ml/min, and the detection wavelengths were 210 nm and 280 nm. Results Harpagide and harpagoside contents in SN showed good linear relationships within 0.1020-0.5100 mg/ml (r=0.9999) and 0.0340-0.1700 mg/ml (r=0.9999). Their average recovery rates were 97.44% and 97.08%.The RSDs were 0.93% and 1.24%.. There were significant differences in the contents of harpagide and harpagoside in SN from 15 origins (P<0.01). The content of harpagoside in Sichuan Long-dong, Zhejiang Lin-an, Zhejiang Pan-an and Henan Nan-feng is higher. Conclusion This method is stable, accurate and reproducible and can be used for the quality control of SN.

Chemical Constituents from the Roots of Scrophularia ningpoensis Hemsl and Their Anti-inflammatory Activities / 中国药学杂志

OBJECTIVE: To study the chemical constituents in the roots of Scrophularia ningpoensis Hemsl. METHODSE: Compounds were isolated by chromatographic techniques and their structures were elucidated by spectral methods. The anti-inflammatory activities of all isolated compounds were evaluated by lipopolysaccharides (LPS)-stimulated BV2 cells model. RESULTS: Eighteen compounds were isolated and identified as 2'''-acetyl angroside C (1), saccatoside (2), 6-O-α-L-(2″-O-feruloyl)rhamnopyranosylcatalpol (3), scrophularioside (4), harprocumbide A (5), 6″-O-β-D-glucopyranosylharpagoside (6), harpagoside (7), 8-O-(p-coumaroyl)-harpagide (8), 8-O-feruloylharpagide (9), 6-O-α-D-galactopyranosylharpagoside (10), 6'-O-cinnamoylharpagide (11), angoroside B (12), angoroside C (13), scrophuloside B1 (14), 2-(3-hydroxy-4-methoxyphenyl)ethyl-O-α-L-arabinopyranosyl-(1→6)-O-α-L-rhamnopyranosyl-(1→3)]-β-D-glucopyranoside (15), darendoside B (16), 6-O-caffeoyl-β-D-fructofuranosyl-(2→1)-α-D-glucopyranoside (17), and sibirioside A (18). CONCLUSION: Compound 1 is new compound, and compounds 2-4, 12, 14, and 17 are isolated from this plant for the first time. Compounds 1, 12, and 13 show significant inhibition against nitric oxide production in LPS-stimulated BV2 cells model.

Comparative study on distribution of endophytic fungi in Scrophularia ningpoensis from different habitats / 中草药

Objective: To analyze the distribution of endophytic fungi in Scrophularia ningpoensis from different habitats. Methods: The endophytic fungi in S. ningpoensis from six localities (Pan’an of Zhejiang Province, Ankang of Shaanxi Province, Wuxue of Hubei Province, Shaodong of Hunan Province, Dazhou of Sichuan Province, and Bozhou of Anhui Province) were isolated, identified, and analyzed according to the morphology and ITS sequences. Results: A total of 3 052 strains of endophytic fungi were isolated from S. ningpoensis in six localities. A total of 84 fungal taxa were identified according to the morphology and ITS sequences, these strains were belong to 25 genera. Thereinto Diaporthe, Fusarium, Nigrospora, Alternaria, Phoma, Corynespora, Epicoccum, and Cladosporium were common genera to S. ningpoensis from six localities, but the dominant genus was different: Fusarium was the dominant genus from Anhui, Hubei, Hunan, and Shaanxi; Alternaria was the dominant genus from Sichuan; Diaporthe was the dominant genus from Zhejiang. According to the similarity coefficient, the composition of the endophytic fungi was distinctly different between six localities. The Shannon-wiener diversity index and Simpson index of endophytic fungi in S. ningpoensis from Zhejiang were found higher than others. The diversity of endophytic fungi in S. ningpoensis from Sichuan was lower than others. Conclusion: Systematic studying the diversity and community structure of endophytic fungi in S. ningpoensis and clarifying their distribution regularity in plant tissues can offer basic data and scientific basis for their development and utilization.

Optimization of steaming technology for decoction pieces of Scrophularia ningpoensis by D-optimal response surface methodology based on AHP- entropy weight method / 中草药

Objective :To determine the weight coefficient and optimize the steaming technology of decoction pieces of Scrophularia ningpoensis (SN). Methods The contents of harpagide, harpagoside, aucubin, acteoside, angoroside C, and cinnamic acid were simultaneously determined by HPLC. The weight coefficient of each component was evaluated by AHP-entropy (analyitc hierarchy process) method. With composite score as index, D-optimal response surface methodology was adopted to investigate the effects of soaking time, steaming time, and drying temperature on the quality of processed products and optimize the processing technology of decoction pieces of SN. Results :Optimal processing parameters were as follows: soaking time was 15.63 min, steaming time was 85 min, drying temperature was 60 ℃, and the synthetical mark was 97.20. Considering the actual situation, the optimum processing technology of SN was obtained by fine-turning the soaking time. The soaking time was 15 min, the steaming time was 85 min, and the drying temperature was 60 ℃. Also, the synthetical mark were 98.53, 99.39, 98.86, and its RSD was 0.47% through the obtained conditions in parallel with three batches of samples. Conclusion: The optimized steaming technology is simple and feasible, which can provide a reference for the steaming of decoction pieces of SN. The method established to simultaneously determine the contents of six components in decoction pieces of SN is rapid and reliable for controlling the quality of decoction pieces of SN.

Tissue Culture and Exercising Seedlings in Scrophularia ningpoensis / 中国实验方剂学杂志

Objective:Stems,petioles,stem sections with axillary and leaves of Scrophularia ningpoensis were taken as the material in vitro to screen out the suitable plantlet regeneration system and optimal exercising seedling conditions. Method:Different explants,hormones and concentrations on the induction and proliferation of cluster bud were studied by L16(45) orthogonal test. One factor analysis of variance (ANOVA) was made on the induction of adventitious buds rooted with different concentrations of hormones. At the same time,different substrates,watering cycles and transition modes were selected to optimize key technologies of exercising seedlings of S. ningpoensis. Result:Stem sections with axillary was the best explant,which was followed by stems,leaves and petioles. The suitable media for the induction of adventitious buds was MS+6-BA 0.5 mg·L-1+NAA 0.2 mg·L-1,with the induction rate of 100.0% and the proliferation multiple of 9.84.The suitable media for root induction was 1/2 MS+IBA 0.2 mg·L-1,with the rooting rate of 100.0% and the number of roots of 39.45.For matrix,they were transplanted with nutrient soil,vermiculite and perlite (5:2:1) as the media,to keep proper matching of fertility,permeability and water retention. The container seedlings can grow well,and the survival rate was more than 95% when they were watered every 2 days,the acclimatization of plantlets took 20 days indoor and 10 days in shaded greenhouses. Conclusion:The clonal propagation system of S. ningpoensis was established to provide an effective way for the efficient,rapid and steady plantlet regeneration,the breeding of high-quality seedlings and the suitable exercising seedling conditions of S. ningpoensis.

Isolation and identification of endophytic fungi producing harpagoside and harpagide from Scrophularia ningpoensis / 中国中药杂志

The endophytic fungi from root,main stem,branch and leaf of Scrophularia ningpoensis were isolated from Zhejiang,whether these strains could yield harpagide or harpagoside were tested by HPLC and LC-MS. According to the morphological characteristic and the similarity of the nucleotide sequence of internal transcribed spacer( ITS) between r DNAs,the strains producing harpagide or harpagoside were identified. The results showed that 210 strains were isolated from the samples,which were classified into 9 orders,13 families and 17 genera by morphological study. Harpagide was detected in endogenous fungi ZJ17 and harpagoside was detected in endogenous fungi ZJ25 by HPLC coupled with LC-MS. ZJ17 was identified as Alternaria alternate and ZJ25 was identified as A.gaisen by its morphology and authenticated by ITS( ITS4 and ITS5 regions and the intervening 5. 8 S rDNA region).

Study on the Effect Mechanism of Iridoid Glycosides of Scrophularia ningpoensis Inhibiting Cardiomyocytes Apoptosis in Myo- cardial Infarction Model Rats Based on Different Caspase Apoptosis Pathways / 中国药房

OBJECTIVE： To study the effect mechanism of iridoid glycosides extracted from Scrophularia ningpoensis inhibiting cardiomyocytes apoptosis in myocardial infarction model rats. METHODS： The male Wistar rats were randomly divided into sham operation group， model group and S. ningpoensis iridoid glycosides low-dose， medium-dose and high-dose groups， with 10 rats in each group. Myocardial infarction models were established by ligating the left anterior descending coronary artery of the rats， and sham operation group was only threaded without ligation. After the model was established， each administration group was given S. ningpoensis iridoid glycosides suspension intragastrically at three different doses of 50，100，200 mg/kg （by the amount of total glycosides extract） with 10 mL/time， twice a day， for consecutive 7 days. Sham operation group and model group were given constant volume of normal saline intragastrically with same method. The changes of S-T segment of lead ECG Ⅱ were recorded before， after and during 7 days of administration. Cardiac function of rats was examined. The serum levels of LDH， CK-MB， cTnⅠ， NT-pro BNP and TNF-α were determined by colorimetry， immunosuppression or ELISA. The apoptosis of myocardial cells was observed by TUNEL method. SOD activity and MDA content in cardiac myocytes were detected by colorimetry. The expressions of Bcl-2， Bax， Cyt C， Caspase-8， Caspase-9， Caspase-12， Caspase-3 and Calpain in cardiac myocytes were detected by ELISA， enzymolysis colorimetry or enzymatic fluorescence assay. RESULTS： Compared with sham operation， electrocardiogram S-T segment was significantly elevated and the left ventricular end-diastolic diameter and left ventricular end-systolic diameter were significantly increased in the model group； left ventricular ejection fraction and short axis shortening rate decreased significantly； serum levels of LDH， CK-MB， cTnⅠ， NT-pro BNP and TNF-α were increased significantly； there were a large number of yellow-brown apoptotic cells in myocardial tissue； the activity of SOD in myocardial tissue was significantly decreased while the content of MDA was significantly increased； the protein expression level of Bcl-2 and Bcl-2/Bax were significantly decreased， while the levels of Bax， Cyt C， Caspase-3， Caspase-8， Caspase-9， Caspase-12 and Calpain were significantly increased （P＜0.05 or P＜0.01）. Compared with model group， above indexes and pathological changes of myocardial tissue were improved significantly in administration group； the level of Bcl-2 and Bcl-2/Bax in cardiomyocytes increased significantly， while the levels of Bax， Cyt C， Caspase-3， Caspase-8， Caspase-9， Caspase-12 and Calpain decreased significantly （P＜0.05 or P＜0.01）. CONCLUSIONS： S. ningpoensis iridoid glycosides can inhibit the activation of Caspase-3 by inhibiting three apoptotic pathways related to Caspase-8， Caspase-9 and Caspase-12， and then inhibit the apoptosis of cardiomyocytes.

Preparation and quality standard of standard decoction of Scrophulariae Radix pieces / 中国中药杂志

The standard decoction of Chinese herbal decoction pieces is a standard reference substance to measure whether different dosage forms of Chinese medicine are basically consistent with those of clinical decoction,and provides new ideas and methods for effectively solving the problems of uneven quality in Chinese medicine dispensing granules. In this study,a systematic method for evaluating the quality of Scrophulariae Radix decoction was established from the perspective of " standard decoction",providing reference for the quality control of the Scrophulariae Radix dispensing granules. 15 batches of Scrophulariae Radix decoction pieces from different origins were collected,and 15 batches of standard decoctions were prepared according to the standardized process with water as solvent.Harpagide and harpagoside were used as quantitative detection indicators to determine the content,calculate the transfer rates and determine the extraction rate. The high performance liquid chromatography( HPLC) was used to establish a standard decoction fingerprint analysis method. The results showed that the transfer rates of harpagide and harpagoside in 15 batches of Scrophulariae Radix pieces standard decoction were( 70. 84±13. 39) % and( 48. 56±6. 40) % respectively; the extraction rate was( 57. 47±5. 89) %. Nine peaks were identified in the HPLC fingerprint,and the similarity was higher than 0. 97 between the fingerprints of 15 batches of standard decoction and the control fingerprint. In this study,the preparation process of standard decoction of Scrophulariae Radix pieces conformed to the traditional decoction preparation method. The sources of the samples were representative,and the established fingerprint method was stable and feasible,which can provide reference for the preparation and quality control of Scrophulariae Radix dispensing granules.

Identification and Analysis of Harpagide Metabolites in Rats in vivo / 中国药房

OBJECTIVE:To study the metabolites,distribution,metabolic type and the possible activity of harpagide which is the active component from Scrophularia ningpoensis in rats in vivo. METHODS:4 SD rats were divided into blank group (ul-tra-pure water) and administration group (harpagide reference solution),2 in each group,ig,160 mg/kg,twice a day,for 3 d. Urine and feces were collected every 12 h before administration and the first administration;sample blood 8 mL was taken after 0.5,1 h of last administration;heart,liver,spleen,lung,kidney,stomach and small intestine were taken. The blood,urine,fe-ces and other tissue solutions were prepared,HPLC-MS was conducted to detect and identify the harpagide metabolites in rats in vi-vo and presume metabolic pathways,and PharmMapper software was used to predict metabolites activity. RESULTS:12 harpagide metabolites were identified in rats in vivo,the form of prototypes and metabolites were distributed in heart,liver,spleen,lung, kidney,stomach and small intestine. The metabolic type mainly included hydrolysis,dehydration,reduction,methylation,sul-fation,glucuronic acid binding,grade A coumaric acid binding,etc. The 12 compounds may have activities in the treatment of epi-lepsy,amyotrophic lateral sclerosis,diabetes,stroke,etc. CONCLUSIONS:Harpagide may be effective in the form of prototypes and metabolites. The study has provided basis for attributing the origins of metabolite,studying the effective form of S. ningpoensis clarifying its pharmacological mechanism and processing mechanism.

Study on the Effects of Exogenous Abscisic Acid on Salt-alkali Tolerance of Scrophularia ningpoensis Seed-lings / 中国药房

OBJECTIVE:To study the effects of exogenous abscisic acid(ABA)on salt-alkali tolerance of Scrophularia ning-poensis seedlings,and to provide theoretic evidence for cultivating S. ningpoensis in salt-alkali soil. METHODS:50 S. ningpoensis seedlings were divided into control group,salt-alkali group(75 mmol/L),salt-alkali(75 mmol/L)+ABA low-concentration,medi-um-concentration and high-concentration groups(10,50,100 μmol/L),with 10 plants in each group. 20 days after transplanting, the plants were sprayed with drugs every 4 days. Growth indexes of the plants(length of stem,fresh weight,dry weight,survival rate),physiological indexes [the contents of chlorophyll,soluble sugar (SS),soluble protein (SP) and free proline (Pro)],the contents of MDA and H2O2,the activities of antioxidase [the activities of SOD,POD,CAT,GR] and the contents of Na+ and K+in stem and root were determined in each group 2 weeks later. RESULTS:Compared with control group,stem height and survival rate of S. ningpoensis seedlings,SP,the contents of Na+ and K+ in the stem,the contents of Na+,CAT and GR in the roots were all decreased significantly in salt-alkali group;while chlorophyll,Pro,MDA and H2O2 contents,the content of K+ in the roots and the activities of SOD and POD were all increased significantly(P<0.05 or P<0.01). Compared with salt-alkali group,stem height of S. ningpoensis seedlings,fresh weight and chlorophyll content were increased significantly in salt-alkali+ABA medium-concentra-tion group,while MDA and the content of Na+ in the stem were decreased significantly;dry weight,the contents of SS and SP, the activities of SOD and CAT were increased significantly in salt-alkali+ABA low-concentration and medium-concentration groups, while the SS content was increased in salt-alkali+ABA high-concentration group;stem height and dry weight were decreased signifi-cantly in salt-alkali+ABA high-concentration group;Pro content of salt-alkali+ABA low-concentration and high-concentration groups were decreased significantly;H2O2,Na+ content in the roots were decreased significantly in salt-alkali+ABA low-concentra-tion,medium-concentration and high-concentration groups;while GR activity,K+ content in the stem and roots were increased sig-nificantly;POD activity were decreased significantly in salt-alkali+ABA medium-concentration and high-concentration groups. CON-CLUSIONS:The addition of a certain concentration of exogenous ABA can effectively increase salt-alkali tolerance of S. ningpoen-sis seedlings and strengthen the ability of the plant adapting to salt-alkali environment.

Clinical efficacy of Scrophularia ningpoensis granules combined with fosinopril for congestive heart failure / 药学实践杂志

Objective To evaluate the efficacy of Scrophularia ningpoensis granules combined with fosinopril in treatment of congestive heart failure.Methods 60 congestive heart failure patients were randomly divided into 2 groups, 30 patients in the treatment group and 30 patients in the control group.Besides the conventional therapy, the control group was treated with fosinopril and the treatment group received Scrophularia ningpoensis granules plus fosinopril for 90 days.Left ventricular end diastolic diameter (LVEDD), left ventricular end systolic diameter (LVESD), left ventricular ejection fraction (LVEF) and serum B-type brain natriuretic peptide (BNP) were measured before and after treatment.Results The total effective rate in the treatment group and the control group were 90%, but significantly effective number in the treatment group was higher than the control group.The levels of BNP, LVEDD and LVESD in both groups were decreased, while LVEF was increased after the treatment.The difference was statistically significant (P<0.01).The degree of improvement of heart function in the treatment group was better than that in the control group (P<0.05).Conclusion The combination therapy of Scrophularia ningpoensis granules and fosinopril is efficacious in treating CHF.

Spectroscopic Study of Interaction of Harpagoside and Human Serum Albumin / 分析化学

Harpagoside (HAR) is believed to be a main compound in Scrophularia ningpoensis which possess a broad of biological activities.Human serum albumin (HSA) has important physiological roles in transportation, distribution and metabolism of many endogenous and exogenous substances in body.It is great significance in pharmacology to investigate the interaction mechanism of HAR and HSA.In this work, the interaction between HAR and HSA was investigated by fluorescence and ultraviolet absorption spectroscopy at different pH (pH=4.0, 7.4, and 9.0) and temperatures (297, 310 and 323 K).The experimental results showed that the HAR could cause the fluorescence quenching of HSA through a static quenching procedure, showing that the HAR regularly quenched the intrinsic fluorescence of HSA, and a decrease in the quenching constant was observed with an increase in temperature.Under different conditions, all the magnitude of binding constants (KA) was larger than 105 L/mol and the number of binding sites (n) in the binary system were approximate to 1.Base on the magnitude of enthalpy and entropy changes, the negative values of ΔG, ΔH and ΔS revealed that the binding of HAR with HSA was spontaneous and exothermic process, and the main interaction forces of the HAR with HAR were van der Waals forces and/or hydrogen bonding interaction.The binding distance (r) between the HAR and HSA was calculated to be about 4.2 nm based on the theory of F(o)rster′s nonradiation energy transfer, which indicated that the energy transfer from HSA to HAR occurred with high possibility.What was more, the synchronous florescence spectroscopy confirmed the conformational changes of HSA during the binding reaction.

A quantitative method for simultaneous assay of seven active ingredients with one marker in Scrophularia ningpoensis root / 中国中药杂志

To establish a new quantitative method for simultaneous determination of multi-components in Scrophularia root by using high performance liquid chromatography (HPLC) and validate its feasibilities．Meanwhile,using catalpol as one chemical reference substance to establish the relative correct factors and relative retention values of aucubin,harpagide,acteoside,angoroside C,harpagoside and cinnamic acid．Then using the quantitative analysis of multi-components by single-marker (QAMS)model,the six analytes can be quantitatively determined in Scrophularia root．The method was evaluated by comparison of the quantitative results to external standard method．No significant differences were observed between the quantitative results of the two methods．The obtained RCFs were credible．It is feasible and suitable to evaluate the quality of Scrophularia root.

Transcriptome characterization for Scrophularia ningpoensis based on high-throughput sequencing technology and related genes for synthesis of terpenoid compounds / 中国中药杂志

To investigate the profile of gene function and search for SSR, a new technology of high-throughput Solexa/Illumina sequencing was used to generate the root transcriptome of Scrophularia ningpoensis, and 65 602 036 raw reads were obtained. Based on the bioinformatics analysis and Trinity, 73 983 unigenes were obtained with an average length of 823 bp. The comparison of sequence homology in database showed that 56 389 unigenes had different degrees of homology. A total of 520 metabolic pathways related genes and 191 relDODO transcription factors were identified by the Swiss-Prot, GO, KEGG and COG.The 11 659 SSRs were found by MISA and the highest frequency was AG/CT. In this study, we obtained numerous SSRs to provide references for the study of functional gene cloning and genetic diversity of S. ningpoensis. The key genes involved in the secondary metabolism are the basis for the study of biosynthesis and regulatory mechanism of the secondary metabolites.

Antifungal effects of three medicinal crops on Phytophthora nicotianae / 中国中药杂志

Tobacco black shank is one of the most harmful soil-borne diseases infected by Phytophthora parasitica. In order to probe the control method to this disease, in this study, the mycelial growth rate method was employed to investigate the antifungal effects of extracts from stem-leaf and root, root exudates, and their combination of Scrophularia ningpoensis, Chuanmingshen violaceum and Pinellia ternata The results showed that: ①Stem-leaf and root extracts of S. ningpoensis, C. violaceum and P. ternata exhibited different antifungal activities, and the inhibition increased with the increase of extract concentration. The antifungal effect of S. ningpoensis extracts at 0.5 g•mL⁻¹ was the strongest than other medicinal plants, the inhibition rate of steam-leaf and root extracts reached 74.88%, 69.27%, respectively. The inhibitory effect of C. violaceum and P. ternata was relatively lower, however, there is a significant gain effect after combination of steam-leaf and root extracts of C. violaceum. ②The root exudates of S. ningpoensis, C. violaceum and P. ternata showed fungistasis to Phytophthora nicotianae, and fungistasis was enhanced with the increase of root exudate concentration. The antifungal effect in the order of C. violaceum > S. ningpoensis > P. ternata. ③The antifungal activity of combination of extract and root exudate from S. ningpoensis was similar with the effect of C. violaceum, they were both stronger than P. ternata, and the antifungal activity for three combination were located between the antifungal activity of their extracts and root exudates. S. ningpoensis and C. violaceum can be potentially applied to prevent and control the tobacco black shank.

Effects of Fu Shen ke li on myocardial cell apoptosis and endoplasmic reticulum stress-related protein expressions in rats with adriamycin induced heart failure / 中国医师杂志

Objective To explore the contribution of endoplasmic reticulum stress to cardiac myocyte apoptosis in mouse congestive heart failure induced by Doxorubicin.Methods Doxorubicin (DOX)was administered intraperitoneally to rats,2 mg/kg doses over 8 weeks.Echocardiographic and hemodynamics measurements were obtained at 8 weeks after treatment.Then left ventricular (LV) samples were collected.The successfully established rats were divided into 3 groups (Fu Shen Ke Li 40 mg/kg,60 mg/kg,and 80 mg/kg).After 5 weeks post the operation,an echocardiographic was performed on all the rats under narcotism,Western blot to test the B-cell lymphoma-2 (Bcl-2),myeloid cell leukemia-1 (Mcl-1),Bcl-2-Associated X (Bax),Caspase-3,C/EBP homologous protein (CHOP),and 78-kDa glucose-regulated protein(GRP78),and reverse transcription-polymerase chain reaction (RT-PCR) to test the mRNA expression levels of Mcl-1,and NOXA.Results The left ventricular end-diastolic diameter (LVEDD),left ventricular end systolic diameter (LVESD),left ventricular ejection fraction (LVEF) prompted modeling success The left ventricular end-diastolic diameter (LVEDD),left ventricular ejection fraction (LVEF),and fractional shortening (FS) have been improved in high and middle groups of Fu Shen Ke Li.Compared to the model group,higher expression levels of Bcl-2,and Mcl-1 were,lower expressions of Bax,NOXA,CHOP,and GRP78 were (P ＜ O.05) in 3 groups of Fu Shen Ke Li.Compared to the model group,there were higher mRNA expression level of Mcl-1 was,lower mRNA expression of NOXA was (P ＜ 0.05) in 3 groups of Fu Shen Ke Li.Conclusions Fu Shen Ke Li depresses endoplasmic reticulum stress,which can reduce the apoptosis of myocardial cells,and ultimately achieve the purpose of treatment of heart failure.

Identification and phylogenetic analysis of endophytic fungi isolated from Scrophularia ningpoensis / 中国中药杂志

The endophytic fungi from root, main stem, branch and leaf of Scrophularia ningpoensis were isolated and identified from Wulong and Chongqing, and the population diversity analysis and phylogenetic analysis were followed. The result indicated that, as to population diversity index, S. ningpoensis from Wulong： leaf>main stem=branch>root, branch from Chongqing>branch from Wulong. Fifty-eight endophytic fungi were obtained, most of which were the pathogens of the plant. Colletotrichum was the prevailing genus, of which C. gloeosporioides and C. boninense were the prevailing strains. Leaf and seedlings might be the main path of infection. Endophytic fungi and pathogen might convert to each other, influenced by such factors as environment, genotype et al.

LC-LTQ-Orbitrap analysis on chemical constituents in Scrophulariae Radix extract and their metabolites in rat plasma / 中国中药杂志

Chemical constituents in extract of Scrophulariae Radix and their metabolites in rat plasma after oral administration were identified by HPLC-LTQ-Orbitrap. Samples were separated by a Venusil MP C₁₈ column using a binary gradient elution. The information on the total ion chromatogram, the extraction chromatogram and the mass spectrogram in a negative mode were synthetically analyzed by comparing the retention time, MS and MS/MS spectra with literature data and some of reference standards to conduct a qualitative study on constituents of Radix Scrophulariae extract in vivo and in vitro. Totally 37 compounds from Scrophularia ningpoensis extract were detected including 12 iridoid glycosides, 20 phenylpropanoids and 5 unknown compounds. In vivo, harpagide, harpagoside and angoroside C were confirmed to enter into the blood in prototype forms. Besides, another 2 prototype compounds and 2 metabolites were detected in rat plasma after oral administration of S. ningpoensis extract. The results are beneficial for the determination of bioactive substances of S. ningpoensis and significant for further studies on S. ningpoensis.

Study on the Quality Standard of Liyan Mixture / 中国药房

OBJECTIVE:To establish the quality standard of Liyan mixture. METHODS:TLC was conducted to identify the Scrophularia ningpoensis,Terminalia chebula and Glycyrrhiza uralensis;HPLC was conducted to determine the content of chloro-genic acid. The column was Agilent Eclipse Plus C18 with mobile phase of methanol-0.2% phosphoric acid (18:82,V/V) at flow rate of 1.0 ml/min,the detection wavelength was 327 nm,column temperature was 30 ℃ and the injection volume was 20 μl. RE-SULTS:TLC showed S. ningpoensis,T. chebula and G. uralensis had clear spots and good separation. There was no interference for negative control. The linear range of chlorogenic acid was 0.16-1.6 μg(r=0.999 9);RSDs of precision,stability and reproduc-ibility tests were lower than 1.0%,recovery was 96.9%-101.4%(RSD=1.17%,n=9). CONCLUSIONS:The method is simple and reproducible,and can be used for the quality control of Liyan mixture.

Study on Resistance Physiological Responses of Seedling of Scrophularia ningpoensis to Saline-alkali Stress / 中国药房

OBJECTIVE:To study the resistance physiological responses of seedling of Scrophularia ningpoensis to the stress caused by saline and alkali at different concentrations. METHODS:The seedling of S. ningpoensis was placed in the illumination in-cubator,in which the temperature was maintained at(20±2)℃,light-dark cycle at 13 h/11 h and illumination intensity at 2 000 lx. The alkali (NaHCO3) of 50,100,150,200 and 250 mmol/L and the saline (NaCl) of 50,100,150,200 and 250 mmol/L were poured once every 4 days for 16 consecutive days. The growth indexes(plant height,fresh weight,dry weight),chlorophyll (Chl) content,antioxidative indexes [superoxide dismutase (SOD),catalase (CAT),peroxidase (POD),hydrogen peroxide (H2O2),malonaldehyde (MDA),membrane penetration (RMP)],and osmotic regulation physiological indexes [soluble sugar (SS),soluble protein(SP)and proline(Pro)] of the seedling were determined. There set a blank control group(distilled water). RESULTS:For saline concentration higher than 50 mmol/L and alkali concentration higher than 150 mmol/L,the seedling height, fresh weight and Chl content were lower;for saline concentration higher than 100 mmol/L and alkali concentration higher than 150 mmol/L,the dry weight of the seedling was lower. For saline concentration of 50 mmol/L and alkali concentration of 150 mmol/L, the activities of SOD and POD were the strongest;for saline concentration of 150 mmol/L and alkali concentration of 50 mmol/L, the activity of CAT was the strongest. The contents of H2O2 and MDA increased and the content of SP decreased with the increase in saline-alkali concentration. The level of RMP increased therewith,that is to say,membrane stability decreased. For saline concen-tration higher than 100 mmol/L and alkali concentration higher than 150 mmol/L,the accumulation of SS terminated;for saline and alkali concentrations higher than 100 mmol/L,the accumulation of Pro terminated,however,the content of Pro was markedly high-er in the above-said alkali concentration than in that saline concentration. CONCLUSIONS:Saline-alkali stress causes the tissues of S. ningpoensis to produce active oxygen and its cytoplasmic membrane system to be damaged. Chl,enzymatic antioxidants such as SOD,CAT and POD,and osmoprotectants such as Pro and SS can buffer saline-alkali stress to some degree. The maximum concen-trations of NaCl and NaHCO3 that seedling of S. ningpoensis can tolerate are 50 mmol/L and 150 mmol/L respectively.