Phytochemical Profiling and Quality Control of Terminalia sericea Burch. ex DC. Using HPTLC Metabolomics.

Terminalia sericea is used throughout Africa for the treatment of a variety of conditions and has been identified as a potential commercial plant. The study was aimed at establishing a high-performance thin layer chromatography (HPTLC) chemical fingerprint for T. sericea root bark as a reference for quality control and exploring chemical variation within the species using HPTLC metabo3lomics. Forty-two root bark samples were collected from ten populations in South Africa and extracted with dichloromethane: methanol (1:1). An HPTLC method was optimized to resolve the major compounds from other sample components. Dichloromethane: ethyl acetate: methanol: formic acid (90:10:30:1) was used as the developing solvent and the plates were visualized using 10% sulfuric acid in methanol as derivatizing agent. The concentrations of three major bioactive compounds, sericic acid, sericoside and resveratrol-3-O-ß-rutinoside, in the extracts were determined using a validated ultra-performance liquid chromatography-photodiode array (UPLC-PDA) detection method. The rTLC software (written in the R-programming language) was used to select the most informative retardation factor (Rf) ranges from the images of the analysed sample extracts. Further chemometric models, including principal component analysis (PCA) and hierarchical cluster analysis (HCA), were constructed using the web-based high throughput metabolomic software. The rTLC chemometric models were compared with the models previously obtained from ultra-performance liquid chromatography coupled with mass spectrometry (UPLC-MS). A characteristic fingerprint containing clear bands for the three bioactive compounds was established. All three bioactive compounds were present in all the samples, although their corresponding band intensities varied. The intensities correlated with the UPLC-PDA results, in that samples containing a high concentration of a particular compound, displayed a more intense band. Chemometric analysis using HCA revealed two chemotypes, and the subsequent construction of a loadings plot indicated that sericic acid and sericoside were responsible for the chemotypic variation; with sericoside concentrated in Chemotype 1, while sericic acid was more abundant in Chemotype 2. A characteristic chemical fingerprint with clearly distinguishable features was established for T. sericea root bark that can be used for species authentication, and to select samples with high concentrations of a particular marker compound(s). Different chemotypes, potentially differing in their therapeutic potency towards a particular target, could be distinguished. The models revealed the three analytes as biomarkers, corresponding to results reported for UPLC-MS profiling and thereby indicating that HPTLC is a suitable technique for the quality control of T. sericea root bark.

History of the fragmentation of the African rain forest in the Dahomey Gap: insight from the demographic history of Terminalia superba.

Paleo-environmental reconstructions show that the distribution of tropical African rain forests was affected by Quaternary climate changes. They suggest that the Dahomey Gap (DG)-the savanna corridor that currently separates Upper Guinean (UG, West Africa) and Lower Guinean (LG, western Central Africa) rain forest blocks-was forested during the African Humid Holocene period (from at least 9 ka till 4.5 ka), and possibly during other interglacial periods, while an open vegetation developed in the DG under drier conditions, notably during glacial maxima. Nowadays, relics of semi-deciduous forests containing UG and LG forest species are still present within the DG. We used one of these species, the pioneer tree Terminalia superba (Combretaceae), to study past forest fragmentation in the DG and its impact on infraspecific biodiversity. A Bayesian clustering analysis of 299 individuals genotyped at 14 nuclear microsatellites revealed five parapatric genetic clusters (UG, DG, and three in LG) with low to moderate genetic differentiation (Fst from 0.02 to 0.24). Approximate Bayesian Computation analyses inferred a demographic bottleneck around the penultimate glacial period in all populations. They also supported an origin of the DG population by admixture of UG and LG populations around 54,000 (27,600-161,000) years BP, thus before the Last Glacial Maximum. These results contrast with those obtained on Distemonanthus benthamianus where the DG population seems to originate from the Humid Holocene period. We discuss these differences in light of the ecology of each species. Our results challenge the simplistic view linking population fragmentation/expansion with glacial/interglacial periods in African forest species.

Distribution pattern of arthropods and their ecological interactions on the leaf surfaces of Terminalia argentea saplings.

Terminalia argentea tree, native to Brazil, is widely used in landscaping, recovering degraded areas, its wood, coal production, and the bark or leaf extracts has medicinal use. Despite of its importance, the arthropod fauna associated to this plant and its interspecific relationships still needs further studies. The objectives of this study were to evaluate the arthropods, their ecological indices and the distribution in the leaf faces on T. argentea saplings. The numbers of phytophagous insects (e.g., Cephalocoema sp.), pollinators (e.g., Tetragonisca angustula), and natural enemies (e.g., Oxyopidae), and their ecological indices (e.g., species richness), were higher on the adaxial leaf faces on T. argentea saplings. Aggregated distribution of phytophagous insects (e.g., Aphis spiraecola), pollinators (e.g., Trigona spinipes), and natural enemies (e.g., Camponotus sp.) on T. argentea saplings was observed. Abundance, diversity, and species richness of natural enemies correlated, positively, with those of phytophagous and pollinators insects. Predators and tending ants followed their prey and sucking insects, respectively. Tending ants protected sucking insects against predators, and reduced chewing insects. The high number of Cephalocoema sp. on T. argentea saplings is a problem, because this insect can feed on leaves of this plant, but its preference for the adaxial leaf face favors its control. The aggregation behavior of arthropods on T. argentea saplings favors the control of potential pests of this plant. There seems to be competition between tending ants for space and food resources on T. argentea saplings.

Nutritional evaluation of outer fleshy coat of Terminalia catappa fruit in two varieties.

The nutritional potential of outer (50-55%) fleshy edible cover of Terminalia catappa fruit was selected for investigation in search of a new source of nutrients. Two different varieties, red and yellow in their ripe and unripe stages, were analyzed for proximate and mineral composition along with phytochemicals. In the results (per 100 g sample), the red variety has shown to be a rich source of protein (1.95 g vs. 1.65 g) while the yellow variety has shown a high content of carbohydrate and ash (12.03 g vs. 6.14 g and 1.21 g vs. 0.70 g). Of the phytochemicals, ß-carotene and vitamin C were found to be present in high amount in the red variety (2,090 µg vs. 754 µg and 138.6 mg vs. 105.4 mg), wherein the former increased while the latter decreased with ripening of the fruit. The results of the study show that the edible outer cover of tropical almond can contribute significantly to the nutrient intake.

Variation in chemical composition and antimalarial activities of two samples of Terminalia albida collected from separate sites in Guinea.

BACKGROUND: The disparity of harvesting locations can influence the chemical composition of a plant species, which could affect its quality and bioactivity. Terminalia albida is widely used in traditional Guinean medicine whose activity against malaria has been validated in vitro and in murine models. The present work investigated the antimalarial properties and chemical composition of two samples of T. albida collected from different locations in Guinea. METHOD: T. albida samples were collected in different locations in Guinea, in Dubréka prefecture (West maritime Guinea) and in Kankan prefecture (eastern Guinea). The identity of the samples was confirmed by molecular analysis. In vitro antiplasmodial activity of the two extracts was determined against the chloroquine resistant strain PfK1. In vivo, extracts (100 mg/kg) were tested in two experimental murine models, respectively infected with P. chabaudi chabaudi and P. berghei ANKA. The chemical composition of the two samples was assessed by ultra-high-performance liquid chromatography coupled to high resolution mass spectrometry. RESULTS: In vitro, the Dubréka sample (TaD) was more active with an IC50 of 1.5 µg/mL versus 8.5 µg/mL for the extract from Kankan (TaK). In vivo, the antiparasitic effect of TaD was substantial with 56% of parasite inhibition at Day 10 post-infection in P. chabaudi infection and 61% at Day 8 in P. berghei model, compared to 14 and 19% inhibition respectively for the treatment with TaK. In addition, treatment with TaD further improved the survival of P. berghei infected-mice by 50% at Day 20, while the mortality rate of mice treated with Tak was similar to the untreated group. The LC/MS analysis of the two extracts identified 38 compounds, 15 of which were common to both samples while 9 and 14 other compounds were unique to TaD and TaK respectively. CONCLUSION: This study highlights the variability in the chemical composition of the species T. albida when collected in different geographical locations. These chemical disparities were associated with variable antimalarial effects. From a public health perspective, these results underline the importance of defining chemical fingerprints related to botanical species identification and to biological activity, for the plants most commonly used in traditional medicine.

In vitro and in vivo antiplasmodial activity and cytotoxicity of water extracts of Phyllanthus emblica, Terminalia chebula, and Terminalia bellerica.

OBJECTIVE: To evaluate the in vitro and in vivo antiplasmodial activity and the cytotoxicity of Phyllanthus emblica Linn, Terminalia chebula Retz, and Terminalia bellerica (Gaertn) Roxb extracts. MATERIAL AND METHOD: Standard phytochemical screening tests were used to detect metabolites in the plant extract. The water extracts of medicinal plants were tested for their antiplasmodial activity in vitro by assessing their ability to inhibit the uptake of [3H] hypoxanthine into the Plasmodium falciparum K1 multidrug-resistant strain. Cytotoxicity of all extracts was determined on Vero cell line. The in vivo antiplasmodial activity in Plasmodium berghei infected mice was evaluated by the standard 4-day suppressive test. RESULTS: Phytochemical screening of the water extracts of three plants revealed the presence of flavonoids, hydrolysable tannins, saponin and terpenes. All plant extracts showed antimalarial activity (IC50 values ranging from 14.33 +/- 0.25-15.41 +/- 0.61 microg/ml). The water extract of Terminalia bellerica (Gaertn) Roxb had the highest in vitro antiplasmodial activity followed by Phyllanthus emblica Linn. and Terminalia chebula Retz. The cytotoxic activity was exhibited by all plant extracts on Vero cells with IC50 values of 157.86 to 238.70 mg/ml. All of the plant extracts showed selectivity with the selectivity index (SI) ranged from 11 to 17. A standard 4-day suppressive test on P. berghei infected mice was used to evaluate the in vivo antiplasmodial activity of the extracts at 250 mg/kg/day. The results revealed that in vivo antiplasmodial activity with good suppression activity ranged from 53.40% to 69.46%. CONCLUSION: All of the plant extracts exhibited interesting in vitro and in vivo antiplasmodial activity with good selectivity.

The combination of ITS2 and psbA-trnH region is powerful DNA barcode markers for authentication of medicinal Terminalia plants from Thailand.

The dried fruits of Terminalia plant (Combretaceae) called "Samo" have been used as herbal medicine in Thai traditional medicine. Four "Samo" crude drugs, namely, Samo thai, Samo thed, Samo dee-ngu, and Samo phiphek, are used as the main ingredients in Triphala and Trisamo recipes. Their commercial products are available in processed and powdered form, but are difficult to authenticate by conventional methods. In this study, we aimed to discriminate species of genus Terminalia for the identification of their crude drugs by a DNA barcoding technique. A total of 208 closely related nucleotide sequences were obtained from nine Terminalia species collected from Thailand and the DDBJ/EMBL/GenBank database. An effective DNA barcode marker was selected from six DNA loci (matK, rbcL, psbA-trnH, ITS, ITS1, and ITS2) and their two-locus combination. All sequences were analyzed by three major methods: (1) BLAST search; (2) the genetic divergence method using Kimura 2-parameter (K2P) distance matrices; and (3) tree topology analysis based on the neighbor-joining method. Comparison of the six candidate DNA loci indicated that ITS identified Terminalia with 100% accuracy at the species and genus levels in the BLAST1 method. ITS2 showed the highest K2P variability. The data from the single markers and the two-locus combinations revealed that only the two-locus combinations, namely, the combinations of rbcL, ITS, ITS1, and ITS2 with psbA-trnH, clearly discriminated all the species. From the results of DNA sequence analysis and the three methods, ITS2 is recommended for the identification of Terminalia species to supplement psbA-trnH.

In vitro anti-Neisseria gonorrhoeae activity of Terminalia macroptera leaves.

We used the agar dilution method to evaluate the antibacterial effect of Terminalia macroptera leaf (Tml) extract against nine reference and clinical Neisseria gonorrhoeae strains, including penicillin- and tetracycline-resistant and -susceptible strains. Tml possesses anti-N. gonorrhoeae activity against all of the strains and the minimum inhibitory concentrations (MIC) were between 100 and 200 microg ml(-1). We then used a liquid-liquid partition method to divide the Tml extract into five fractions and determined the anti-N. gonorrhoeae activity of each of the fractions. All of the fractions showed antibacterial activity. The most active one was identified as the diethyl ether fraction and had MIC values of between 25 and 50 microg ml(-1) against all of the strains.

[Analysis of tannins in Fructus Chebulae and its confusion varieties by HPCE].

AIM: To analyze the hydrolyzable tannins-chebulinic acid (I) and chebulagic acid(II) in Fructus Chebulae and its confusion varieties by using high performance capillary electrophoresis (HPCE) method. METHODS: Using a capillary (375 microns OD x 50 microns ID; 81.5 cm x 61.5 cm) and a power supply set at 24 kV, with phosphate-borate buffer containing 20 mmol.L-1 Na2HPO4-60 mmol.L-1 boric acid and a UV detector at 280 nm, sample solution was loaded in decompression mode at the positive end of the capillary, the loading time was 5 s. RESULTS: The linear ranges of I and II were 0.0842-0.842 and 0.842 and 0.0940-0.940 mg.mL-1 respectively, the correlation coefficient were 0.9966 and 0.9957, the average recoveries were 95.6% (RSD = 4.0%, n = 5) and 95.0% (RSD = 4.4%, n = 5), the RSDs (n = 5) of measurement precision test were 2.2% and 1.7%, the RSDs (n = 6) of reproduction test were 5.4% and 4.0% respectively. The contents of I and II were obviously interrelated with the variety and characteristics of Fructus Chebulae, the contents of I and II in the confusion varieties of Fructus Chebulae were very low. CONCLUSION: It is suitable to use I and II as the criterion in quality evaluation of Fructus Chebulae, and the HPCE method is effective for quality evaluation of the crude Fructus Chebulae.

[Assay of three hydrolyzable tannins in Fructus Chebulae from different habitats by RP-HPLC].

Three hydrolyzable tannins chebulinic acid (I), chebulagic acid(II) and 1,3, 6-tri-O-galloyl-beta-D-glucose (III) in Fructus Chebulae from different habitats were determined by RP-HPLC method. The contents of I and II were obviously interrelated with the variety and characteristics of Fructus Chebulae. It's suitable to use I and II as indexes in quality evaluation of the crude drug of Fructus Chebulae.

Instrumental neutron activation analysis of Emblica officinalis, Terminalia belerica and Terminalia chebula for trace element efficacy and safety.

To assess the efficacy and safety, essential (Cl, Co, Cr, Fe, K, Mn, Na, Se, Zn), toxic (As, Br, Hg, Sb,) and other elements (Ba, Ce, Cs, Eu, Rb, Sc) were determined in Emblica officinalis (EO), Terminalia belerica (TB) and Terminalia chebula (TC) using instrumental neutron activation analysis. These herbs contain K as a major element, while Co, Cr and Na in EO, Fe, K and Mn in TB and Cl and Zn in TC are the highest.