Rapid in situ diversification rates in Rhamnaceae explain the parallel evolution of high diversity in temperate biomes from global to local scales.

The macroevolutionary processes that have shaped biodiversity across the temperate realm remain poorly understood and may have resulted from evolutionary dynamics related to diversification rates, dispersal rates, and colonization times, closely coupled with Cenozoic climate change. We integrated phylogenomic, environmental ordination, and macroevolutionary analyses for the cosmopolitan angiosperm family Rhamnaceae to disentangle the evolutionary processes that have contributed to high species diversity within and across temperate biomes. Our results show independent colonization of environmentally similar but geographically separated temperate regions mainly during the Oligocene, consistent with the global expansion of temperate biomes. High global, regional, and local temperate diversity was the result of high in situ diversification rates, rather than high immigration rates or accumulation time, except for Southern China, which was colonized much earlier than the other regions. The relatively common lineage dispersals out of temperate hotspots highlight strong source-sink dynamics across the cosmopolitan distribution of Rhamnaceae. The proliferation of temperate environments since the Oligocene may have provided the ecological opportunity for rapid in situ diversification of Rhamnaceae across the temperate realm. Our study illustrates the importance of high in situ diversification rates for the establishment of modern temperate biomes and biodiversity hotspots across spatial scales.

Bioassay-guided Fractionation of Anti-Leishmania amazonensis Extracts from Ampelozizyphus amazonicus by High-speed Countercurrent Chromatography.

Ampelozizyphus amazonicus Ducke, known as "saracura-mirá" in the Amazon rainforest, is valued for its traditional use in malaria prevention and treatment. The plant's roots and bark are employed for these purposes, while the wood is often overlooked. Given the global importance of leishmaniasis, research focused on A. amazonicus anti-Leishmania amazonensis potential. The ethyl acetate extract from the bark (EAEB) exhibited the most effective inhibition of intracellular amastigote growth with IC50 7.0 µg.mL-1 but showed high toxicity (CC50 9.0 µg.mL-1). The wood ethanol (EW) and ethyl acetate (EAEW) extracts demonstrated promising results against L. amazonensis with IC50 15.8 µg.mL-1 and 10.0 µg.mL-1, CC50 50.5 µg.mL-1 and 22.7 µg.mL-1, respectively. High-speed countercurrent chromatography combined with mass spectrometry  off-line detection was employed for the bioassay-guided fractionation of EAEB and EAEW using HEMWat as solvent system. These extracts showed to be rich in saponins and triterpenes, besides minor glycosylated flavonoids. HSCCC was effective in obtaining high purity fractions, leading to the identification of a di-glycosylated triterpene saponin from EAEB, and melaleucic acid from EAEW. These findings highlight A. amazonicus as a potential source for developing new therapeutic alternatives for leishmaniasis and HSCCC as a technique enabling better separation efficiency of complex saponin matrices.

Revisiting chloroplast genomic landscape and annotation towards comparative chloroplast genomes of Rhamnaceae.

BACKGROUND: Massive parallel sequencing technologies have enabled the elucidation of plant phylogenetic relationships from chloroplast genomes at a high pace. These include members of the family Rhamnaceae. The current Rhamnaceae phylogenetic tree is from 13 out of 24 Rhamnaceae chloroplast genomes, and only one chloroplast genome of the genus Ventilago is available. Hence, the phylogenetic relationships in Rhamnaceae remain incomplete, and more representative species are needed. RESULTS: The complete chloroplast genome of Ventilago harmandiana Pierre was outlined using a hybrid assembly of long- and short-read technologies. The accuracy and validity of the final genome were confirmed with PCR amplifications and investigation of coverage depth. Sanger sequencing was used to correct for differences in lengths and nucleotide bases between inverted repeats because of the homopolymers. The phylogenetic trees reconstructed using prevalent methods for phylogenetic inference were topologically similar. The clustering based on codon usage was congruent with the molecular phylogenetic tree. The groups of genera in each tribe were in accordance with tribal classification based on molecular markers. We resolved the phylogenetic relationships among six Hovenia species, three Rhamnus species, and two Ventilago species. Our reconstructed tree provides the most complete and reliable low-level taxonomy to date for the family Rhamnaceae. Similar to other higher plants, the RNA editing mostly resulted in converting serine to leucine. Besides, most genes were subjected to purifying selection. Annotation anomalies, including indel calling errors, unaligned open reading frames of the same gene, inconsistent prediction of intergenic regions, and misannotated genes, were identified in the published chloroplast genomes used in this study. These could be a result of the usual imperfections in computational tools, and/or existing errors in reference genomes. Importantly, these are points of concern with regards to utilizing published chloroplast genomes for comparative genomic analysis. CONCLUSIONS: In summary, we successfully demonstrated the use of comprehensive genomic data, including DNA and amino acid sequences, to build a reliable and high-resolution phylogenetic tree for the family Rhamnaceae. Additionally, our study indicates that the revision of genome annotation before comparative genomic analyses is necessary to prevent the propagation of errors and complications in downstream analysis and interpretation.

Dispersal modes affect Rhamnaceae diversification rates in a differentiated manner.

The evolution of dispersal modes has been proposed to promote the diversification of angiosperms. However, little is known about the relative impact of different dispersal modes on plant diversification. We test the association between dispersal modes and diversification rates using Rhamnaceae, the cosmopolitan buckthorn family, as a model. We found that species with diplochory have the highest diversification rates followed by those with myrmecochory and ballistic dispersal, while lineages dispersed by vertebrates and wind have relatively low diversification rates. The difference in diversification rates may be closely linked to the difference in dispersal distance and ecological interactions implied by each dispersal mode. Species which disperse over larger geographical distances may have much higher speciation rates due to the increased chance of establishing isolated populations due to geological barriers or habitat fragmentation. However, long-distance dispersal may also increase the chance of extinction. By contrast, species with short-distance dispersal modes may have low speciation rates. Complex interactions with the surrounding environment may, however, impact diversification rates positively by increasing plant survival and reproductive success.

Solid-phase adsorption methodologies of naturally occurring anthraquinones: A review.

INTRODUCTION: Solid-phase extraction applied to plant matrices is nowadays a well-validated technique allowing to concentrate and purify different secondary metabolites. Several classes of phytochemicals have been selectively extracted by this methodology. During the last decade attention has been focused on biologically active anthraquinones from numerous sources like edible, healthy, and medicinal plants. OBJECTIVES: The aim of this review is to provide a detailed literature survey of the solid-phase adsorption methodologies for the extraction of natural anthraquinones reported so far and to discuss and propose future directions in this field of research. MATERIALS AND METHODS: Substructure search was performed in the SciFinder Scholar, PubMed, Medline, and Scopus databases. RESULTS: The first report about application of solid-phase adsorption for the purification of anthraquinones appeared in the literature in 2002. From this date, and in particular during recent years, the most notable examples included the use of chitin- and chitosan-based polymers, of molecularly imprinted polymers, of coated magnetic nanoparticles, of miniaturized matrix solid-phase dispersion, of functionalized resins, of differently structured lamellar solids, and finally of vortex-synchronized matrix solid-phase dispersion. CONCLUSIONS: The herein detailed solid-phase adsorption methodologies are powerful tools to selectively extract natural anthraquinones and/or provide anthraquinone-enriched phytopreparations. Nevertheless, many other important methods have been applied to synthetic anthraquinones (e.g., azo dyes). These could be conveniently employed also for natural anthranoids. Studies in this field are discussed in this review article.

Anti-SARS-CoV-2 Activity of Ampelozizyphus amazonicus (Saracura-Mirá): Focus on the Modulation of the Spike-ACE2 Interaction by Chemically Characterized Bark Extracts by LC-DAD-APCI-MS/MS.

Traditional medicine shows several treatment protocols for COVID-19 based on natural products, revealing its potential as a possible source of anti-SARS-CoV-2 agents. Ampelozizyphus amazonicus is popularly used in the Brazilian Amazon as a fortifier and tonic, and recently, it has been reported to relieve COVID-19 symptoms. This work aimed to investigate the antiviral potential of A. amazonicus, focusing on the inhibition of spike and ACE2 receptor interaction, a key step in successful infection. Although saponins are the major compounds of this plant and often reported as its active principles, a polyphenol-rich extract was the best inhibitor of the spike and ACE2 interaction. Chemical characterization of A. amazonicus bark extracts by LC-DAD-APCI-MS/MS before and after clean-up steps for polyphenol removal showed that the latter play an essential role in maintaining this activity. The effects of the extracts on viral replication were also assessed, and all samples (aqueous and ethanol extracts) demonstrated in vitro activity, inhibiting viral titers in the supernatant of Calu-3 cells after 24 hpi. By acting both in the SARS-CoV-2 cell entry process and its replication, A. amazonicus bark extracts stand out as a multitarget agent, highlighting the species as a promising candidate in the development of anti-SARS-CoV-2 drugs.

Isolation and identification of novel selective antitumor constituents, sidrin and sidroside, from Zizyphus spina-christi.

Background: The leaves of Zizyphus spina-christi (L.) Willd contain several compounds exhibiting different pharmacologic activities. However, studies on the cytotoxic activity of these compounds are limited. Objectives: We aimed to investigate and isolate cytotoxic compounds with selective antitumor effects from the leaves of Z. spina-christi using bioassay-guided fractionation of methanol extract. Methods: Powdered, dried leaves were subjected to methanol extraction and fractionated using n-hexane, chloroform, ethyl acetate, and n-butanol. Fractions with positive cytotoxicity against HeLa and THP-1 cell lines were further fractionated and eluted using various concentrations of organic solvents. Active compounds were isolated using different chromatographic methods and their chemical structures were determined using extensive spectroscopic methods, such as 1D NMR (1H NMR, 13C NMR, and DEPT), 2D NMR (COSY, HMBC, and HMQC), HRFAB-MS, and IR. Furthermore, the cytotoxic effects of the isolated compounds were evaluated against 62 tumor cell lines (including HeLa and THP-1) in addition to normal bone marrow cells. Results: The chloroform and aqueous methanol fractions of the leaves showed cytotoxic activity. Two compounds were successfully isolated and named "sidrin" (13-ß-hydroxy-lup-20(30)-ene-2,3-ß-epoxy-28-carboxylate) and "sidroside" (3-O-ß-D-glucopyranosyl-(1-3)-α-L-arabinopyranosyl-jujubogenin-20-O-α-L-rhamnopyranoside). Sidrin exhibited cytotoxic activity against the human leukemia (Hl-60, RPMI-8226), lung cancer (A549, EKVX), breast cancer (BT-549, MDA-MB-231/ATCC), colon cancer (KM12), melanoma (M14, SK-MEL-5), and central nervous system (CNS) cancer (SF-295) cell lines, and selectivity was observed against the Hl-60, EKVX, BT-549, KM12, and SF-295 cell lines. In addition, sidrin was more active than sidroside and doxorubicin against the Hl-60 and EKVX cell lines. In contrast, sidrin had a similar effect to doxorubicin against the BT-549 and renal cancer (UO-31) cell lines. Sidroside was more selective against the leukemia (CCRF-CEM, MOLT-4), lung cancer (HOP-92, NCI-H322M), breast cancer (MDA-MB-468), melanoma (LOX IMVI), CNS cancer (SNB-19), ovarian cancer (OVCAR-8), renal cancer (UO-31, RXF 393), and prostate cancer (PC-3) cell lines. Both compounds exhibited similar activity against the breast cancer (MDA-MB-231, T-47D), colon cancer (HCC-2998, HCT-116), ovarian cancer (OVCAR-3), renal cancer (UO-31, 786-0, and SN 12C) cell lines. Normal bone marrow cells were unaffected at the same concentrations of sidrin and sidroside applied to tumor cells. Conclusions: These results suggest tumor-selective cytotoxicity of sidrin and sidroside.

Cyclopeptide Alkaloids from Discaria chacaye (Rhamnaceae) as Result of Symbiosis with Frankia (Actinomycetales).

Cyclopeptide alkaloids with different biological activities are present in plants of the family Rhamnaceae. Plants of this family grow in a symbiotic relationship with aerobic Gram-positive actinomycetes belonging to the genus Frankia. This goal of this research was a study of the comparative profile of alkaloids present in Discaria chacaye and to establish a connection between the presence or absence of Frankia sp. and the alkaloids. In addition, insecticidal activities of the alkaloidal extract were examined. A total of 24 alkaloids were identified, of which 12 have a benzylisoquinoline skeleton, 9 were cyclopeptides, 2 isoquinolines, and 1 aporphine. The presence of cyclopeptide alkaloids is associated with Frankia nodules in the plant root. The alkaloid extracts showed insecticidal activity with mortality dose-dependence and LD50 values between 44 to 71 µg/mL.

Historical biogeography of Pomaderris (Rhamnaceae): Continental vicariance in Australia and repeated independent dispersals to New Zealand.

AIM: Gondwanan biogeographic patterns include a combination of old vicariance events following the breakup of the supercontinent, and more recent long-distance dispersals across the southern landmasses. Floristic relationships between Australia and New Zealand have mostly been attributed to recent dispersal events rather than vicariance. We assessed the biogeographic history of Pomaderris (Rhamnaceae), which occurs in both Australia and New Zealand, by constructing a time-calibrated molecular phylogeny to infer (1) phylogenetic relationships and (2) the relative contributions of vicariance and dispersal events in the biogeographic history of the genus. LOCATION: Australia and New Zealand. METHODS: Using hybrid capture and high throughput sequencing, we generated nuclear and plastid data sets to estimate phylogenetic relationships and fossil calibrated divergence time estimates for Pomaderris. BioGeoBEARS and biogeographical stochastic mapping (BSM) were used to assess the ancestral area of the genus and the relative contributions of vicariance vs dispersal, and the directionality of dispersal events. RESULTS: Our analyses indicate that Pomaderris originated in the Oligocene and had a widespread Australian distribution. Vicariance of western and eastern Australian clades coincides with the uplift of the Nullarbor Plain c. 14 Ma, followed by subsequent in-situ and within-biome diversification with little exchange across regions. A rapid radiation of southeastern Australian taxa beginning c. 10 Ma was the source for at least six independent long-distance dispersal events to New Zealand during the Pliocene-Pleistocene. MAIN CONCLUSIONS: Our study demonstrates the importance of dispersal in explaining not only the current cross-Tasman distributions of Pomaderris, but for the New Zealand flora more broadly. The pattern of multiple independent long-distance dispersal events for Pomaderris, without significant radiation within New Zealand, is congruent with other lowland plant groups, suggesting that this biome has a different evolutionary history compared with the younger alpine flora of New Zealand, which exhibits extensive radiations often following single long distance dispersal events.

Squandering water in drylands: the water-use strategy of the phreatophyte Ziziphus lotus in a groundwater-dependent ecosystem.

PREMISE: Water is the most limiting factor in dryland ecosystems, and plants are adapted to cope with this constraint. Particularly vulnerable are phreatophytic plants from groundwater-dependent ecosystems (GDEs) in regions that have to face water regime alterations due to the impacts of climate and land-use changes. METHODS: We investigated two aspects related to the water-use strategy of a keystone species that dominates one of the few terrestrial GDEs in European drylands (Ziziphus lotus): where it obtains water and how it regulates its use. We (1) evaluated plants' water sources and use patterns using a multiple-isotope approach (δ2 H, δ18 O, and Δ13 C); (2) assessed the regulation of plant water potential by characterizing the species on an isohydric-anisohydric continuum; and (3) evaluated plants' response to increasing water stress along a depth-to-groundwater (DTGW) gradient by measuring foliar gas exchange and nutrient concentrations. RESULTS: Ziziphus lotus behaves as a facultative or partial phreatophyte with extreme anisohydric stomatal regulation. However, as DTGW increased, Z. lotus (1) reduced the use of groundwater, (2) reduced total water uptake, and (3) limited transpiration water loss while increasing water-use efficiency. We also found a physiological threshold at 14 m depth to groundwater, which could indicate maximum rooting length beyond which optimal plant function could not be sustained. CONCLUSIONS: Species such as Z. lotus survive by squandering water in drylands because of a substantial groundwater uptake. However, the identification of DTGW thresholds indicates that drawdowns in groundwater level would jeopardize the functioning of the GDE.

A multiple-trait analysis of ecohydrological acclimatisation in a dryland phreatophytic shrub.

Water is the main limiting factor for groundwater-dependent ecosystems (GDEs) in drylands. Predicted climate change (precipitation reductions and temperature increases) and anthropogenic activities such as groundwater drawdown jeopardise the functioning of these ecosystems, presenting new challenges for their management. We developed a trait-based analysis to examine the spatiotemporal variability in the ecophysiology of Ziziphus lotus, a long-lived phreatophyte that dominates one of the few terrestrial GDEs of semiarid regions in Europe. We assessed morpho-functional traits and stem water potential along a naturally occurring gradient of depth-to-groundwater (DTGW, 2-25 m) in a coastal aquifer, and throughout the species-growing season. Increasing DTGW and salinity negatively affected photosynthetic and transpiration rates, increasing plant water stress (lower predawn and midday water potential), and positively affected Huber value (sapwood cross-sectional area per leaf area), reducing leaf area and likely, plant hydraulic demand. However, the species showed greater salt-tolerance at shallow depths. Despite groundwater characteristics, higher atmospheric evaporative demand in the study area, which occurred in summer, fostered higher transpiration rates and water stress, and promoted carbon assimilation and water loss more intensively at shallow water tables. This multiple-trait analysis allowed us to identify plant ecophysiological thresholds related to the increase in salinity, but mostly in DTGW (13 m), and in the evaporative demand during the growing season. These findings highlight the existence of tipping points in the functioning of a long-lived phreatophyte in drylands and can contribute to the sustainable management of GDEs in southern Europe, paving the way for further studies on phreatophytic species.

Mass spectrometry as a tool for the dereplication of saponins from Ampelozizyphus amazonicus Ducke bark and wood.

INTRODUCTION: Mass spectrometry in natural products research has been used as a first step to identify possible chemical structures and to guide subsequent efforts to isolate novel compounds. Preparations of Ampelozizyphus amazonicus Ducke (AA) are known for their high content of saponins, especially dammarane-type triterpenoid aglycones. In the Amazon, where it is known as "saracura-mirá", roots and bark are widely used for the treatment and prevention of malaria, while the wood is discarded. The extract prepared from the wood is also saponin-rich, but its exact chemical composition has not been described. OBJECTIVE: This study provides information on the chemical profiling and tentative structural identification of the major compounds (saponins) present in aqueous and ethanol extracts of bark and wood of AA by mass spectrometry. METHODS: The strategy used to identify compounds present in all samples was ultra-high-performance liquid chromatography with an ultraviolet detector coupled to tandem mass spectrometry (UHPLC-UV-MS/MS) for the analysis of fragmentation patterns through product ion scan using MZmine 2 software. Also, direct sample injection and electrospray ionisation combined with high-resolution mass spectrometry (DI-ESI-HRMS) measurements were performed. RESULTS: The extracts showed chemical similarity, and 95 saponins were tentatively identified in AA wood and bark, including 73 which are described for the first time as tentative structures for this plant species. CONCLUSION: This research describes a useful method for the fast and simultaneous tentative identification of major saponins in AA, contributing to the study of the chemical properties of this genus and family. Furthermore, it demonstrates the importance of the qualitative dereplication process, allowing a straightforward way to propose the tentative identification of compounds.

Comprehensive mass spectrometry-guided phenotyping of plant specialized metabolites reveals metabolic diversity in the cosmopolitan plant family Rhamnaceae.

Plants produce a myriad of specialized metabolites to overcome their sessile habit and combat biotic as well as abiotic stresses. Evolution has shaped the diversity of specialized metabolites, which then drives many other aspects of plant biodiversity. However, until recently, large-scale studies investigating the diversity of specialized metabolites in an evolutionary context have been limited by the impossibility of identifying chemical structures of hundreds to thousands of compounds in a time-feasible manner. Here we introduce a workflow for large-scale, semi-automated annotation of specialized metabolites and apply it to over 1000 metabolites of the cosmopolitan plant family Rhamnaceae. We enhance the putative annotation coverage dramatically, from 2.5% based on spectral library matches alone to 42.6% of total MS/MS molecular features, extending annotations from well-known plant compound classes into dark plant metabolomics. To gain insights into substructural diversity within this plant family, we also extract patterns of co-occurring fragments and neutral losses, so-called Mass2Motifs, from the dataset; for example, only the Ziziphoid clade developed the triterpenoid biosynthetic pathway, whereas the Rhamnoid clade predominantly developed diversity in flavonoid glycosides, including 7-O-methyltransferase activity. Our workflow provides the foundations for the automated, high-throughput chemical identification of massive metabolite spaces, and we expect it to revolutionize our understanding of plant chemoevolutionary mechanisms.

The assembly of the Cape flora is consistent with an edaphic rather than climatic filter.

The Cape flora is compositionally biased, being dominated by a few fynbos clades (such as Iridaceae, Ericaceae, Proteaceae and Restionaceae) that make up major part of the distinct heathland vegetation in the Cape Floristic Region. Uncertainty exists concerning what excluded the subtropical to tropical palm-dominated woodland/forest vegetation that was the dominant component in the CFR in the Paleocene and allowed the fynbos clades, which are largely derived from outside Africa, to establish and radiate. Two filters have been proposed. The first postulates that the establishment of the Mediterranean climate driven by the late Miocene initiation of the cold-water Benguela Upwelling System (BUS) eliminated the African lineages and allowed the establishment and radiation of sclerophyllous plant clades ("the Mediterranean climate model", MCM). Alternatively, the "oligotrophic soils model" (OSM) postulates that the oligotrophic soils, gradually exhumed by post-Gondwanan Late Cretaceous - early Cenozoic erosion, acted as a filter excluding the African lineages. In this study, we re-calibrate the fynbos clade Phylica (Rhamnaceae), the genus initially used to test the MCM, using new fossil data to test if the crown age precedes the Late Miocene. Our results indicate that we cannot significantly reject a crown age of Phylica consistent with the MCM. We compare the MCM and OSM model for the Cape fynbos flora by compiling the crown ages of 22 fynbos clades. We show that crown ages are not clustered in time around the initiation of the BUS but, are dispersed throughout the Cenozoic. This suggests that oligotrophic soils, rather than summer drought, acted as a filter. Consequently, we argue that the fynbos clades radiated separately in expanding edaphically controlled heathland patches in the Cape mountains as sandstone exhumation after the Gondwanan break-up progressed.

Physico-Chemical and Phytochemical Characterization of Moroccan Wild Jujube "Zizyphus lotus (L.)" Fruit Crude Extract and Fractions.

Wild jujube "Ziziphus lotus (L.) Desf." belongs to the Rhamnaceae family and is a traditionally herbaceous medicinal plant. It is very common in arid and semi-arid regions and is currently used for its antidiabetic, sedative, analgesic, anti-inflammatory and hypoglycemic activities. The aim of the present work was to characterize the physico-chemical properties and the phytochemical profile of wild jujube sample collected from the Guercif region, in order to determine the polyphenolic compounds and the antioxidant ability Analyses were carried out directly after the harvest for the determination of pH, refractive index, total soluble solid (°Brix), dry matter, sugar/acidity, total sugars, reducing sugars, as well as lipid and protein content. Results showed that the investigated fruit is acidic (pH 4.9 ± 0.23) and rich in sugars (80.2 g/100 g ± 3.81). The GC-MS analysis of the fruit revealed a number of volatile compounds, as many as 97, belonging to different chemical classes. The HPLC-DAD-ESI/MS analysis showed the presence of a total of 20 polyphenolic compounds in both EtOAc and MeOH-water extracts. Among them, p-Hydroxybenzoic acid was the most abundant in the EtOAc extract (185.68 µg/100 mg ± 0.5) whereas Quercetin 3-O-rhamnoside-7-O-glucoside was found in higher amounts in the MeOH-water extract (25.40 µg/100 mg ± 0.5). These components have medical interest, notably for human nutrition, as well as health benefits and therapeutic effects. Therefore, Moroccan jujube "Zizyphus lotus (L.)" fruit may have potential industrial applications for food formulations.

Evolution of leaf structure and drought tolerance in species of Californian Ceanothus.

PREMISE OF THE STUDY: Studies across diverse species have established theory for the contribution of leaf traits to plant drought tolerance. For example, species in more arid climates tend to have smaller leaves of higher vein density, higher leaf mass per area, and more negative osmotic potential at turgor loss point (πTLP ). However, few studies have tested these associations for species within a given lineage that have diversified across an aridity gradient. METHODS: We analyzed the anatomy and physiology of 10 Ceanothus (Rhamnaceae) species grown in a common garden for variation between and within "wet" and "dry" subgenera (Ceanothus and Cerastes, respectively) and analyzed a database for 35 species for leaf size and leaf mass per area (LMA). We used a phylogenetic generalized least squares approach to test hypothesized relationships among traits, and of traits with climatic aridity in the native range. We also tested for allometric relationships among anatomical traits. KEY RESULTS: Leaf form, anatomy, and drought tolerance varied strongly among species within and between subgenera. Cerastes species had specialized anatomy including hypodermis and encrypted stomata that may confer superior water storage and retention. The osmotic potentials at turgor loss point (πTLP ) and full turgor (πo ) showed evolutionary correlations with the aridity index (AI) and precipitation of the 10 species' native distributions, and LMA with potential evapotranspiration for the 35 species in the larger database. We found an allometric correlation between upper and lower epidermal cell wall thicknesses, but other anatomical traits diversified independently. CONCLUSIONS: Leaf traits and drought tolerance evolved within and across lineages of Ceanothus consistently with climatic distributions. The πTLP has signal to indicate the evolution of drought tolerance within small clades.

Rhamnellosides A and B, ω-Phenylpentaene Fatty Acid Amide Diglycosides from the Fruits of Rhamnella franguloides.

Two new ω-phenylpentaene fatty acid amide diglycosides, rhamnellosides A (1) and B (2), were isolated from the fruits of Rhamnella franguloides (Rhamnaceae). These compounds were prioritized using LC-MS/MS molecular networking dereplication based on our previous discovery of 2-acetoxy-ω-phenylpentaene fatty acid triglycosides berchemiosides A-C from a phylogenetically related species, Berchemia berchemiifolia. The structures of the isolated compounds were elucidated by spectroscopic analyses in combination with chemical derivatization. The pentaene groups of 1 and 2 were found to have (6E, 8E, 10Z, 12Z, 14E)-geometry, which is the same as that found in berchemioside A.

Complete chloroplast genome sequence of a major economic species, Ziziphus jujuba (Rhamnaceae).

Ziziphus jujuba is an important woody plant with high economic and medicinal value. Here, we analyzed and characterized the complete chloroplast (cp) genome of Z. jujuba, the first member of the Rhamnaceae family for which the chloroplast genome sequence has been reported. We also built a web browser for navigating the cp genome of Z. jujuba ( http://bio.njfu.edu.cn/gb2/gbrowse/Ziziphus_jujuba_cp/ ). Sequence analysis showed that this cp genome is 161,466 bp long and has a typical quadripartite structure of large (LSC, 89,120 bp) and small (SSC, 19,348 bp) single-copy regions separated by a pair of inverted repeats (IRs, 26,499 bp). The sequence contained 112 unique genes, including 78 protein-coding genes, 30 transfer RNAs, and four ribosomal RNAs. The genome structure, gene order, GC content, and codon usage are similar to other typical angiosperm cp genomes. A total of 38 tandem repeats, two forward repeats, and three palindromic repeats were detected in the Z. jujuba cp genome. Simple sequence repeat (SSR) analysis revealed that most SSRs were AT-rich. The homopolymer regions in the cp genome of Z. jujuba were verified and manually corrected by Sanger sequencing. One-third of mononucleotide repeats were found to be erroneously sequenced by the 454 pyrosequencing, which resulted in sequences of 1-4 bases shorter than that by the Sanger sequencing. Analyzing the cp genome of Z. jujuba revealed that the IR contraction and expansion events resulted in ycf1 and rps19 pseudogenes. A phylogenetic analysis based on 64 protein-coding genes showed that Z. jujuba was closely related to members of the Elaeagnaceae family, which will be helpful for phylogenetic studies of other Rosales species. The complete cp genome sequence of Z. jujuba will facilitate population, phylogenetic, and cp genetic engineering studies of this economic plant.

Two new C-glucosyl flavonoids from Ziziphus jujube and their anti-inflammatory activity.

Two new C-glucosyl flavonoids 6''-O-feruloylspinosin and 6''-O-feruloyl-6'''-p-hydroxybenzoylspinosin, together with five known compounds, were isolated from the seeds of Ziziphus jujube (Rhamnaceae family). Their structures were elucidated on the basis of chemical and spectroscopic evidences. Compounds 1-7 showed moderate inhibitory effects against COX-1 and COX-2 enzymes.

Extract of Ziziphus jujuba Fruit (Jujube) Stimulates Expression of Enzymes Responsible for Heme Recycle via Anti-oxidant Response Element in Cultured Murine Macrophages.

Jujube, the fruit of Ziziphus jujuba Mill., is a functional food and commonly used as a health supplement worldwide. To study the beneficial role of jujube in heme iron recycling during erythrophagocytosis, the expression of heme oxygenase-1 (HO-1), biliverdin reductase A and B, and ferroportin were determined in jujube-treated cultured RAW 264.7 macrophages. Application of a chemically standardized jujube water extract in cultured RAW 264.7 cells for 24 h stimulated the expressions of HO-1, biliverdin reductase A, biliverdin reductase B, and ferroportin in a concentration-dependent manner, having the maximal responses from twofolds to threefolds. A plasmid containing anti-oxidant response element, a regulator for HO-1 transcription, was transfected into RAW 264.7 cells. Application of jujube water extract onto the transfected macrophages stimulated the anti-oxidant response element-mediated transcriptional activity by twofolds. These results supported the potential capacity of jujube by regulating expressions of heme iron recycling genes in cultured macrophages.