Revealing the Impact of the Environment on Cistanche salsa: From Global Ecological Regionalization to Soil Microbial Community Characteristics.

To understand the regulatory relationship between the environment and Cistanche salsa, here we integrated the macro- and microdimension methods. From a macroperspective, the MaxEnt model indicated that countries along the Belt and Road Initiative, such as China, Egypt, and Libya, were particularly suitable for growth of C. salsa from ancient times (Last Glacial Maximum and mid-Holocene) to the future (2050 and 2070). The Jackknife test revealed that precipitation is an important ecological factor that affects C. salsa's distribution. From a microperspective, 16S rRNA amplicon sequencing data showed that the soil microbial communities of three ecotypes (desert-steppe, grassland, and gravel-desert) were significantly different (p < 0.001). Core microbiome analysis demonstrated that the bacterial genera Arthrobacter, Sphingomonas, and Bacillus were enriched core taxa of C. salsa. LEfSe and random forest were used to excavate the Gillisia (desert-steppe), Flavisolibacter (grassland), and Variibacter (gravel-desert) as biomarkers that can distinguish among microbial communities from the three ecotypes. The prediction profile showed that the metabolic function of the microbial community was enriched in metabolic pathways and environmental information processing. Correlation analyses revealed that the altitude, precipitation of the warmest quarter (bio18), mean diurnal range (bio2), and mean temperature of the warmest quarter (bio10) were important ecological factors that affect the composition of soil microbial communities. This work provided new insights into the regulatory relationship among the suitable distribution of C. salsa, soil microbial communities, and ecological drivers. Moreover, it deepened the understanding of the interaction between desert plants and ecological factors in arid environments.

Chemical Diversity and Prediction of Potential Cultivation Areas of Cistanche Herbs.

Owing to hostile growth environments and increasing related production, Cistanche plants have decreased in number. The aim of the present study was to evaluate the quality of and to predict potential suitable regions for two official species and two nonofficial species (C. salsa and C. sinensis) by high-performance liquid chromatography and the MaxEnt model. The results indicated that 2'-acetylacteoside was present only in C. deserticola. The compound can be used as a potential chemical marker to discriminate C. deserticola from the three other Cistanche plants. Anthocyanin A and carotenoid F were the common constituents of the two official species only and can thus be used as chemical markers to differentiate between official and nonofficial species. The prediction results of a potentially suitable distribution indicated that C. sinensis has much wider regions for potential distribution than the other species. Finally, the echinacoside content in C. deserticola was significantly different between the two suitable potential distributions, and the contents of samples from Inner Mongolia were significantly higher than those from Gansu Province. This is the first application of the combination of the contents of chemical components and the results of MaxEnt models for the quality assessment of herbal medicine. Our results may provide a reference for the sustainable utilization of endangered Cistanche species.

Ecological modeling of Cistanche deserticola Y.C. Ma in Alxa, China.

Cistanche deserticola Y.C. Ma has long been used for medical purposes in China. It mainly grows in the Chinese provinces of Inner Mongolia, Ningxia, Gansu, and Xinjiang, and the species in the Alxa region of northwest China, have the most distinct qualities. To explain the geoherbalism quality and geographical distribution of C. deserticola, we sampled 65 wild plants in Alxa, determined their echinacoside and acteoside content, and assessed the relationship between the ecological environment and quality of C. deserticola through maximum entropy modeling and geographic information system. We identified the areas suitable for the growth of high-quality C. deserticola species. The regionalization analysis of growth suitability showed that the most influential ecological factors for the growth of C. deserticola are soil type, annual sunshine duration, altitude, temperature seasonality (standard deviation ×100), vegetation type, sunshine duration in the growing season, mean precipitation in August and mean temperature in July. The most suitable areas for growing C. deserticola are southeast of Ejin Banner, central Alxa Right Banner, and north of Alxa Left Banner. The regionalization analysis of quality suitability showeds that the most influential ecological factors for glycosides in C. deserticola are sunshine duration in June, average precipitation in May, and average temperature in March, and the best-quality C. deserticola grows in Dalaihob Town, Ejin Banner. Upon inspection, the result of the experiment reached a high accuracy of 0.994, which indicates that these results are consistent with the actual distribution of C. deserticola in Alxa. The results of this study may serve as a scientific basis for site selection of artificial planting bases for C. deserticola.

Safety assessment of powdered Cistanche deserticola Y. C. Ma by a 90-day feeding test in Sprague-Dawley rats.

Cistanche deserticola (C. deserticola), a holoparasitic plant widely distributed in arid or semi-arid areas in Eurasia and North Africa, has been used as an important tonic in traditional Eastern medicine for centuries. However, little information on the systemic toxicity and safety evaluation of it is available. The purpose of this study was to investigate the potential toxicity of powdered C. deserticola as a novel food ingredient by use of a subchronic toxicity study in Sprague-Dawley (SD) rats. A total of 80 male and female rats were fed with diets containing 8, 4, 2 and 0% (control) powdered C. deserticola for 90 days. A toxicological assessment was performed including mortality, body and organ weight, food consumption, blood biochemistry, hematology, gross necropsy and histopathological examinations. There were no signs of toxicity and treatment-related changes in rats treated with powdered C. deserticola. The no-observed-adverse-effect level (NOAEL) of powdered C. deserticola was 7.8 g kg-1 body weight for males and 8.0 g kg-1 body weight for females of rats under the experimental conditions of this study.

[Experiment on pruning of Cistanche deserticola inoculated in artificial Haloxylon ammodendron forest].

At present, the objective of cutting and pruning Cistanche deserticola is to harvest in successive years and enhance the harvesting yield and quality of C. deserticola in the process of the artificial cultivating C. deserticola. An experiment was conducted focusing on cutting and pruning C. deserticola in artificial forests of Haloxylon ammodendron drip-irrigated with saline water at the hinter-land of the Taklimakan desert, according to different growth stages and lengths. The results were following: (1) The effect of cutting on C. deserticola was similar to that of pruning, which resulted in three kinds of morphological types, not related to the bloom and size of C. deserticola. (2) The growth forms were diversified after pruning. Among them, there had sprouting new body, died or maintaining life with no sprouting, mildewed on its surface layer, etc. However, some of new bodies were sprouting from the lower part of the old body. The death rate of bloomed C. deserticola was higher than that of the underground, and the death rate of the 40 cm in stubble height for C. deserticola was higher than those with the stubble height of 20 cm and 5 cm. (3) Most of the diameter of living C. deserticola after pruning was increasing, but some of them changed little. (4) The mildew and rot of C. deserticola and the broken of the roots of the H. ammodendron and the fallen of the point of the inoculated when it was dug, which would cause the death of the C. deserticola. On the other, the yield-increasing effect and the economic benefit of the techniques of the pruning of Cistanche would need further research and evaluate. Therefore, the application of this technique needs to be cautious.

[Lignans from stems of Cistanche deserticola cultured in Tarim desert].

In order to clarify the chemical constituents of Cistanche deserticola cultured in Tarim desert, a systematically phytochemical investigation was carried out. The chemical constituents were isolated by column chromatography, such as silica gel, Sephadex LH- 20, MCI gel, ODS and semi-preparative HPLC, and their structures were determined on the basis of MS, NMR spectroscopic analysis and/or comparison with literature data. Eleven lignans were isolated from the 85% ethanol extract of the stems of C. deserticola cultured in Tarim desert. Their structures were identified as (+)-syringaresinol-4'-O-ß-D-glucopyranoside (1), (+)-isoeucommin A (2), eucommin A (3), (+)-pinoresinol monomethylether ß-D-glucoside (4), lariciresinol 4'-O-ß-D-glucopyranoside (5), lariciresinol 4-O-ß-D-glucopyranoside (6), conicaoside (7), dehydrodiconiferyl alcohol 4-O-ß-D-glucopyranoside (8), dehydrodiconiferyl alcohol γ'-O-ß-D-glucoside (9), citrusin A (10), and alaschanioside A (11). Compounds 1, 3-7, 10 and 11 were isolated from this genus for the first time, and compounds 2, 8 and 9 were obtained from this species for the first time.

[Ecology suitability study of Cistanche deserticola in Alxa].

The distribution information of Cistanche deserticola was collected by interview investigation and field survey, and 55 related environmental factors were collected, the habitat suitability study was conducted based on geographic information system (GIS) and Maximum entropy model. The AUCs of ROC curve were both above 0.9, indicating that the predictive results with the maxent model were highly precise. The results showed that 14 major environmental factors have obvious influence on ecology suitability distributions of C. deserticola, including vegetation type et al, the suitable distribution areas are mainly concentrated in the central of Alxa Youqi, the north of Alxa Zouqi and the south-east of Ejin Banner, including Tamusu towns, Alateng towns et al, The zoning results basically coincide with the genuine producing areas, and further afford new suitable distribution areas, which can provide reference for the siting of introduction and cultivation of C. deserticola.

Effects of feeding tyrosine and phenylalanine on the accumulation of phenylethanoid glycosides to Cistanche deserticola cell suspension culture.

AIM: To investigate the effects of feeding phenylalanine (Phe) and tyrosine (Tyr) on the accumulation of total phenolic compounds and four phenylethanoid glycosides (PeGs) to a cell suspension culture of the parasitic plant Cistanche deserticola. METHOD: A cell suspension culture of C. deserticola was established and precursors of different concentrations were fed. In each group, the cell was sampled at the 24(th) day after inoculation. The content of total phenolic compounds and four PeGs compounds were determined using the Folin-Ciocalteu method and an HPLC method, respectively. RESULTS: In the Phe fed cells, the maximum PeGs yield was achieved when Phe was fed at 1.5 mmol·L(-1) and the yield reached 1.13 times the control cell concentration. In the Tyr fed cells, the maximum yield of PeGs was 1.60 times of control when 0.75 mmol·L(-1) Tyr was fed to the cells. Furthermore, it was found that the salidroside yield was 4.01 times of control group when 5 mmol·L(-1) Tyr was fed. CONCLUSION: Tyr is a better precursor for PeGs accumulation compared with Phe, and the rate limiting enzymes might be involved in the Tyr branch.

[Chemical constituents from stems of Cistanche deserticola cultured in Tarim desert].

In order to clarify the chemical constituents of Cistanche deserticola cultured in Tarim desert, a systematically phytochemical investigation was carried out. The chemical constituents were isolated by column chromatography, over silica gel, Sephadex LH-20, MCI gel, ODS and semi-preparative HPLC, and their structures were determined on the basis of MS, NMR spectroscopic data analysis, physicochemical properties and/or comparison with literature data Seventeen compounds were isolated from the 85% ethanol extract of the stems of C. deserticola cultured in Tarim desert. Their structures were identified as salsaside B (1), syringin (2), demethyl syrinyin (3), coniferin (4), (2E,6E)-3,7-dimethyl-8-hydroxyoctadien-1-O-beta-D-glucoside (5), (+)-syringaresinol (6), 2S,3S, 4S-trihydroxypentanoic acid (7), panaxytriol (8), beta-sitosterol-3-O-beta-D-xylopyranoside (9), androsin (10), 3-methyl-but-2-en-1-yl-beta-D-glucopyranoside (11), benzyl-glucopyranoside (12), 4-hydroxybenzyl-beta-D-glucoside (13), nicotinamide (14), p-hydroxybenzoic acid (15), 4-hydroxy-benzeneethanol (16), and galactitol (17). Compounds 3, 6-13 were isolated from this genus for the first time, and compounds 1, 4 and 5 were obtained from this species for the first time.

[Analysis of volatile compounds of inflorescence by GC-MS from Cistanche deserticola].

OBJECTIVE: To study the volatile compounds from inflorescence of Cistanche deserticola and provide basis for its utilization and seed breeding. METHODS: The volatile compounds were collected by dynamic headspace adsorption and analyzed by gas chromatography-mass spectrometry (GC-MS). RESULTS: Forty volatile components were identified in inflorescence of Cistanche deserticola from squaring period to full-bloom period. The main components in buds of Cistanche deserticola were hydrocarbons and green leaf volatiles in squaring period. Some components were characteristic in buds and disappeared or decreased in flowers. The relative contents of some components gradually increased with the buds blooming. And some components only emerged in flowers of Cistanche deserticola. The higher content of esters and aromatics were found in flowers, which were significantly increased in comparison with the volatile compounds from buds. CONCLUSION: The volatile compounds from inflorescence of Cistanche deserticola were complex, consisting of various compositions and significantly different with buds blooming.

[Preliminary study of reproductive allocation in Cistanche deserticola].

OBJECTIVE: Discuss the trade-off between vegetative growth and reproductive growth. METHOD: Vegetative modules and reproductive modules by large sampling were separated and quantitative charecters of organs and pattern of reproductive allocation were analysed of reproductive allocation were analysed. RESULT: The phenotype plasticity of quantitative charecters were significant. Reproductive biomass was increasing at different stages. The mass of reproductive investment increased significantly with increasing individual size, the opposite of reproductive allocation. There were all significant positive correlations between the seeds weight and height, total biomass, vegetative biomass, reproductive biomass, fruit numbers, fruit weight. CONCLUSION: The significance of phenotype plasticity is the cause of multiple elements. In the process of transferring from vegetative growth to reproductive growth, there was a trade-off between organs. Under individual growth and reproductive strategy, the variation of biomass was allocated coordinately.

[Mineral elements content of C. deserticola at different growth stages].

ICP - AES technology was used to determine the mineral elements content of Cistanche deserticola Ma (C. deserticola). The results showed that: (1) At succulent stem stages, the content of K was the highest in 5 macroelements, it can reach to 9.45 mg x g(-1), and the proportion for K : Na : P : Ca : Mg was 12 : 3.4 : 1.6 : 1.4 : 1. Among 5 microelements, the content of Fe was the highest and can reach to 97.31 microg x g(-1), and the proportion for Fe : Cu : Mn: Zn : Cu : B was 25 : 3.7 : 3.5 : 1.2 : 1. (2) At reproductive growth stages, the contents of Ca, Mg, Fe, Zn, Cu, Mn and B at unearthed stage were significantly higher than that at the underearth stage, especially for the content of Fe and Mn, they can respectively reach to 697. 55 and 38.75 microg x g(-1) at capsule formative stage. The contents of Fe and Mn at capsule formative stage were almost 7.2 and 8.3 times than that at succulent stem stage. (3) At reproductive growth stages, the aerial part will exclude Na and accumulate P, K, Ca, Mg, Fe, Mn, Zn, Cu and B. This result of the study will be a scientific basis for evaluating the quality of C. deserticola.

Application of molecular markers in predicting production quality of cultivated Cistanche deserticola.

We developed a set of molecular markers in Cistanche deserticola Y. C. MA to evaluate the production quality of cultivated C. deserticola individuals. This application utilizes the inter-simple-sequence repeat (ISSR) polymerase chain reaction (PCR) and random amplified polymorphic DNA (RAPD) PCR as molecular markers to determine the echinacoside content in cultivated C. deserticola individuals. The unweighted pair-group method using arithmetic average clustering (UPGMA) confirmed that the combined ISSR and RAPD data could categorize all C. deserticola individuals into three groups according to their respective echinacoside content. The stepwise multiple regression analysis (SMRA) revealed six potential markers associated with echinacoside accumulation in C. deserticola and produced 18 echinacoside-marker prediction models, four of which were successfully used to predict the quality of C. deserticola from Neimenggu. Both clustering and SMRA showed a correlation between the echinacoside content and molecular markers in cultivated C. deserticola. The relative average deviation of prediction (RADP) of the prediction models could be improved by simplifying and adjusting the model. It was found that the RADP value could reach 2.6% after adjustment and the simplified prediction models could successfully predict the quality of cultivated C. deserticola individuals.

[Rapid and undamaged determination of C. deserticola by Fourier-transform infrared spectroscopy and clustering analysis].

Cistanche deserticola, an endemic species in China, has been one of the grade II national key conservation rare and endangered plants. The spectra of cultivated and wild C. deserticola samples were determined by Fourier transform infrared (FTIR) spectrometry. Based on the fingerprint infrared spectrum from 450 to 2 000 cm(-1), C. deserticola samples were rapidly classfied and closely studied by using the method of clustering analysis. Results showed that although there were tiny differences between the spectra of different origin, including the wild and cultivated C. deserticola samples, these samples could be successfully classified by soft independent modeling of class analogy (SIMCA). Recognition rate and rejection rate of different C. deserticola samples were up to 90%. When testing with the blind sample which the authors picked out from the chosen samples, the accuracy of clustering reaches up to 95%. On the whole, combined with clustering analysis, FTIR provides a effective way to evaluate the origin of the Chinese medicines rapidly and undamagedly.

[Effect of different plant growth regulators on transplanting survival ratio of Haloxylon ammodendron seedling and parasitic ratio of Cistanche deserticola].

OBJECTIVE: To study the effect of different plant growth regulators on the transplanting survival ratio of Haloxylon ammodendron seedlings and parasitic ratio of Cistanche deserticola. METHODS: Different combinations of PDJ and rooting powder were used for rooting soaking with Haloxylon ammodendron seedlings, then the survival ratio of Haloxylon ammodendron and parasitic conditions of Cistanche deserticola were statistical analyzed by SAS software. RESULTS: PDJ had promotable effect on the inoculating survival ratio of Haloxylon ammodendron and the parasitic ratio, parasitic number and size of Cistanche deserticola, but had no significant effect on the transplanting survival ratio. High concentration of PDJ (1 x 10(-5) mol/L) and rooting powder treatment improved the inoculation survival ratio, and low concentration of PDJ (1 x 10(6) mol/L) treatment improved the parasitic ratio of Cistanche deserticola, but PDJ with rooting powder treatment had inhibitive effect on the parasitism of Cistanche deserticola. Principal component analysis showed that the transplanting survival ratio, inoculating survival ratio, parasitic ratio and the parasited number of Cistanche deserticola could be taken to evaluate the effect of different plant growth regulators, and the comprehensive effect was the best with PDJ treatment. CONCLUSION: Rooting soaking with PDJ can be used to improve the survival ratio of Haloxylon ammodendron seedling and parasitic ratio of Cistanche deserticola.

[Suitability evaluation of Cistanche tubulosa based on TCMGIS-I].

OBJECTIVE: To analyze the appropriate producing area of Cistanche tubulosa by mean of TCMGIS-I. METHOD: The TCMGIS-I (Suitability evaluation geographic information system of traditional Chinese medicine producing area) was used to analyze the appropriate producing area of C. tubulosa basing on the optimum ecological factors of Sorth Sinkiang municipality which is the traditional producing area of C. tubulosa. RESULT: The suitable producing areas of C. tubulosa include 56 counties with 190952.1 km2 total areas in Sinkiang municipality, Neimongd municipality and Gansu province; Sinkiang municipality, Gansu and Qinghai provinces have the second-suitable producing areas of C. tubulosa, with 46 counties and 41217.37 km2 total areas; Sinkiang municipality, Inner Mongolia municipality, Gansu province and Ningxia municipality have the third-suitable producing areas of C. tubulosa with 56 counties and 421145.2 km2 total areas. CONCLUSION: It is important to analyze the appropriate producing area of C. tubulosa by means of TCMGIS-I. The results are useful for development of C. tubulosa transplanting and cultivation.

[Studies on chemical constituents of cultivated Cistanche salsa].

OBJECTIVE: To study the chemical constituents of cultivated Cistanche salsa. METHODS: Compounds were isolated and purified on several chromatography, and then were identified by physico-chemical properties and structurally elucidated by spectral analysis. RESULTS: Seven compounds were isolated and identified as beta-sitosterol (I), daucosterol (II), beta-sitosteryl glucoside 3'-O-heptadecoicate (III), 8-hydroxygeraniol 1-beta-D-glucopyranoside (IV), 2-methanol-5-hydroxy-pyridine (V), betaine (VI), galactitol (VII). CONCLUSION: The chemical constituents of artificial cultivated Cistanche salsa are studied for the first time. Among them, compound III and IV are isolated from the plant for the first time, compound V is isolated from this genus for the first time.

[Study on conditions of seed germination of Cistanche].

OBJECTIVE: To study the effect of fluridone concentration, stimulating period, temperature and salt on the seed germination of three species of Cistanche. METHOD: The seeds were cultured in Petri dish, and the germination percentage was counted. RESULT: The highest germination percentage was observed in Cistanche tubulosa, C. deserticola, C. sala seeds pre-treated by 0.1 mg x L(-1) fluridone for 24-29 h. The optimal temperature for the seeds germination of three species of Cistanche was at 20-30 degrees C, and the seeds did not germinate at sub-or supraoptimal temperatures (5 and 35 degrees C). The salt tolerance of C. sala seeds was strong, and the critical value of NaCl concentration was 0.04 mol x L(-1). By contrast, C. tubulosa and C. deserticola seeds were more sensitive to the salt stress, the critical value of NaCl concentration was 0.02 mol x L(-1). CONCLUSION: The optimal germination condition and the method of testing germination percentage of three species of Cistanche seeds are as follow: the seeds are pre-treated by 0.1 mg x L(-1) fluridone for 24 h and then cultured at 20-30 degrees C in salt solution which concentration is lower than 0.02 mol x L(-1).

[Study on reproduction biology of Cistanche deserticola--seedling emergence and flowering].

OBJECTIVE: To study the reproduction characteristic of Cistanche deserticola. METHOD: The reproduction characteristic and growing dynamic in Ningxia plantation were investigated, the contents of boschnaloside and echinacoside were determined during reproduction period. RESULT: The germination period of C. deserticola was in the first ten days of the month, the duration from the germination to flowering lasted about one month. The contents of boschnaloside and echinacoside decreased during reproduction period. CONCLUSION: The reproduction period of C. deserticolais lasts about two months, germination period of C. deserticola is the suitable harvest time.

[Suitability evaluation of Cistanche desertiola based on TCMGIS-I].

OBJECTIVE: To analyze the appropriate producing area of Cistanche desertiola. METHOD: The TCMGIS-I (suitability evaluation geographic information system of traditional Chinese medicine producing area) was used to analyze the appropriate producing area of C. desertiola basing on the optimum ecological factors of Aalashan, Inner-Mongolia autonomous region and North Xinjiang au-tonomous region, the traditional producing areas of C. desertiola. RESULT: The results showed that the suitable producing areas of C. desertiola included 159 counties of 5 autonomous regions and provinces, i.e. Xinjiang, Inner Mongolia, Ningxia autonomous regions, and Gansu as well as Qinghai provinces. The total area was 675 354.9 km2 and distributed mainly in Aalashan of Inner Mongolia, eastern part of North Xinjiang, northern part of Gansu and in the middle of Ningxia. CONCLUSION: The suitable producing areas based on TCMGIS-I covered all areas registered in the 3rd national investigation of Chinese traditional medicine resource. And the results were also corresponding to the traditional producing area of C. desertiola recorded in ancient literature and the successful cultivation areas nowadays. It suggested the rationality and reliability of TCMGIS-I.