A customised target capture sequencing tool for molecular identification of Aloe vera and relatives.

Plant molecular identification studies have, until recently, been limited to the use of highly conserved markers from plastid and other organellar genomes, compromising resolution in highly diverse plant clades. Due to their higher evolutionary rates and reduced paralogy, low-copy nuclear genes overcome this limitation but are difficult to sequence with conventional methods and require high-quality input DNA. Aloe vera and its relatives in the Alooideae clade (Asphodelaceae, subfamily Asphodeloideae) are of economic interest for food and health products and have horticultural value. However, pressing conservation issues are increasing the need for a molecular identification tool to regulate the trade. With > 600 species and an origin of ± 15 million years ago, this predominantly African succulent plant clade is a diverse and taxonomically complex group for which low-copy nuclear genes would be desirable for accurate species discrimination. Unfortunately, with an average genome size of 16.76 pg, obtaining high coverage sequencing data for these genes would be prohibitively costly and computationally demanding. We used newly generated transcriptome data to design a customised RNA-bait panel targeting 189 low-copy nuclear genes in Alooideae. We demonstrate its efficacy in obtaining high-coverage sequence data for the target loci on Illumina sequencing platforms, including degraded DNA samples from museum specimens, with considerably improved phylogenetic resolution. This customised target capture sequencing protocol has the potential to confidently indicate phylogenetic relationships of Aloe vera and related species, as well as aid molecular identification applications.

Anti-tyrosinase activity of South African Aloe species and isolated compounds plicataloside and aloesin.

Tyrosinase is the key enzyme in the production of melanin. Tyrosinase inhibitors have gained interest in the cosmetics industry to prevent hyperpigmentation and skin-related disorders by inhibiting melanin production. It has been reported that several Aloe species exhibit anti-tyrosinase efficacy in vitro. In this study, the exudates of thirty-nine South African Aloe species were screened to identify species and compounds with anti-tyrosinase activity. Qualitative screening revealed that twenty-nine Aloe species exhibited tyrosinase inhibition activity with one to three active bands. Quantitative screening was performed for 29 species and expressed as IC50 values. Three species were further analysed and subsequently, aloesin and aloeresin A was isolated from A. ferox and plicataloside from A. plicatilis and A. chabaudii. Aloeresin A was determined to be a substrate of mushroom tyrosinase. Dose-response assays showed that aloesin (IC50 = 31.5 µM) and plicataloside (IC50 = 84.1 µM) exhibited moderate to weak activity. Molecular docking scores for plicataloside were considerably lower than for aloesin (P < 0.01), confirming its lower IC50. Several Aloe species may have potential for the management of hyperpigmentation or as a skin lightening agent. This is the first report showing that plicataloside, present in A. plicatilis and A. chabaudii, exhibits anti-tyrosinase activity.

Preparation and characterization of liposomes loaded with silver nanoparticles obtained by green synthesis

The objective of this work was to develop and characterize liposomes loaded with silver nanoparticles (LAgNPs) to show improvement in stability characteristics. AgNPs were prepared by the green synthesis method with Aloe vera gel extract and exposure to sunlight. Liposomes were prepared by the modified reverse phase method. Particle size, polydispersity index, zeta potential, as well as the scanning electron microscopy (SEM) morphological aspects of AgNPs and LAgNPs were evaluated. In addition, was used flame atomic absorption spectroscopy to determine the amount of AgNP that was encapsulated in liposomes. The AgNPs presented as amorphous and polydisperse structures, with a mean diameter of 278.46 nm and zeta potential of -18.3 mV. LAgNPs had a mean diameter between 321 and 373 nm, the polydispersity index close to 0.2 and a zeta potential around -40 mV, which indicates greater stability to the AgNPs. The images obtained by SEM show semicircular structures for AgNPs and well-defined spherical shape for LAgNPs. The percentage of encapsulation was between 51.81 to 58.83%. These results showed that LAgNPs were obtained with adequate physicochemical characteristics as a release system.

Ecological Distribution and Genetic Variations of Some Aloe Species in Taif, KSA.

BACKGROUND AND OBJECTIVES: Aloe is a medicinally and economically important genus. Many Aloes seem an endangered species because of over-collection, destruction of plants and destroyed of natural habitats. The objectives of current study was to survey, collect and identification of some Aloe species and to analyze genetic variations between the collected Aloe species. MATERIALS AND METHODS: Four Aloe species (A. armatissima, A. edentata, A. parvicoma and A. pseudorubroviolacea) and Agave americana (Asperagaceae) were used as plant materials for ecological and genetic studies. In RAPD and ISSR analysis 23 and 16 primers, respectively were screened. RESULTS: Ecological study showed that the 4 species are endemic: 2 are endangered (A. edentata and A. parvicoma) and the others are not-endangered (A. armatissima and A. pseudorubroviolacea), while A. americana was introduced as ornamental species. Concerning RAPD, a total of 134 reproducible bands of them 131 bands are polymorphic ~ 97.65% polymorphism were produced, which ranged from 9 bands (primer OPC-04) to 18 (primer OPA-03) bands, with an average 13.4 bands/ primer, ranging from ~300-2500 bp. According to ISSR, 113 reproducible bands were totally yielded with an average 12.6 bands/primer, from ~180-1500 bp, of which 107 poly-morphic bands number (PBN) ~94.96% polymorphism ranged from 10 bands (primer UBC-818 and primer UBC-819) to 14 (primer UBC-814) with an average of 11.9 PB/primer. CONCLUSION: The results revealed high genetic variations between 4 bands Aloe species and A. americana species, which will be in concern for improvement, breeding and conservation programs.

Comparative Analysis of Soluble Proteins in Four Medicinal Aloe Species by Two-Dimensional Electrophoresis and MALDI-TOF-MS.

Background: Aloe barbadensis Miller 1768, A. vera L. var. chinensis (Haw.) Berger 1908, A. ferox Miller 1768, and A. arborescens Miller 1768 are the most widely cultivated species of Aloe and are used in Asia along with 400 other Aloe species worldwide because of their potent and potential bioactivity. Objective: The objective was to analyze and compare the soluble proteins of four commonly used medicinal Aloe species. Methods: Aloe protein samples were obtained by TCA/acetone-saturated phenol-methanol/ammonium acetate combined extraction (phenol extraction), and then were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and two-dimensional gel electrophoresis. Finally, the differentially expressed proteins of four Aloe species were identified by matrix-assisted laser desorption ionization-time-of-flight-MS analysis. Results: The phenol extraction method was the most suitable method for the protein extraction of Aloe. Fifty differentially expressed proteins in four Aloe species were successfully identified and divided into eight functional categories. Furthermore, Malate dehydrogenase and ran-binding protein in A. barbadensis, cytoskeletal-related protein tubulin in A. vera var. chinensis and auxin-induced protein PCNT-115 in A. arborescens are closely related to their morphological characteristics. Conclusions: There are differences in the soluble proteins of the four Aloe species. Those proteins, related to the difference of their morphology of Aloe, might be used to identify different species. Highlights: Fifty differentially expressed proteins in four medicinal Aloe species were identified, and these proteins were classified into eight categories according to their biological functions. Four special proteins closely related to the morphological characteristics of Aloe were found and might be used to identify these four Aloe species.

Medicinal Mascarene Aloes: An audit of their phytotherapeutic potential.

A phytochemical and biological investigation of the endemic Mascarene Aloes (Aloe spp.), including A. tormentorii (Marais) L.E.Newton & G.D.Rowley, A. purpurea Lam, A. macra Haw., A. lomatophylloides Balf.f and A. vera (synonym A. barbadensis Mill.), which are used in the traditional folk medicine of the Mascarene Islands, was initiated. Methanolic extracts of the Aloes under study were analysed using high resolution LC-UV-MS/MS and compounds belonging to the class of anthraquinones, anthrones, chromones and flavone C-glycosides were detected. The Mascarene Aloes could be distinguished from A. vera by the absence of 2″-O-feruloylaloesin and 7-O-methylaloeresin. GC-MS analysis of monosaccharides revealed the presence of arabinose, fucose, xylose, mannose and galactose in all the Mascarene Aloes and in A. vera. The crude extracts of all Aloes analysed displayed antimicrobial activity against Bacillus cereus, Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa. Only extracts of A. macra were active against P. aeruginosa and Klebsiella pneumoniae, while none of the Aloe extracts inhibited Propionibacterium acnes. A. macra displayed anti-tyrosinase activity, exhibiting 50% inhibition at 0.95mg/mL, and extracts of A. purpurea (Mauritius) and A. vera displayed activity in a wound healing-scratch assay. In vitro cytotoxicity screening of crude methanolic extracts of the Aloes, using the MTT (3-(4, 5-dimethylthiazolyl-2)-2, 5-diphenyltetrazolium bromide) showed that only A. purpurea (Réunion) elicited a modest toxic effect against HL60 cells, with a percentage toxicity of 8.2% (A. purpurea-Réunion) and none of the Aloe extracts elicited a toxic effect against MRC 5 fibroblast cells at a concentration of 0.1mg/mL. Mascarene Aloe species possess noteworthy pharmacological attributes associated with their rich phytochemical profiles.

Identification and discrimination of three common Aloe species by high performance liquid chromatography-tandem mass spectrometry coupled with multivariate analysis.

Aloe arborescens, Aloe barbadensis and Aloe ferox are the most widely cultivated and used among 500 aloe species due to their potent bioactivity. However, the difference of aloe species is neglected and labeled only one name Aloe in the market without specifying aloe species discrimination in general. Furthermore, differences in bioactivity and side effects from different aloe species have not been well investigated. This study develops an effective method for simultaneous qualitative and quantitative determination of three common aloe species using high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) and high performance liquid chromatography-diode array detection (HPLC-DAD). The extraction conditions were optimized by response surface methodology (RSM) based on methanol concentration, extraction time and solvent-to-material ratio. A partial least squares-discriminant analysis (PLS-DA) was used to identify the three aloe species. The developed HPLC-MS/MS method coupled with multivariate analysis can be applied to discriminate three aloe species successfully.

The uses of Kenyan aloes: an analysis of implications for names, distribution and conservation.

BACKGROUND: The genus Aloe is renowned for its medicinal and cosmetic properties and long history of use. Sixty-three Aloe species occur in Kenya, of which around 50 % are endemic. Several species of aloes are threatened with extinction and knowledge about their use is of major importance for sound conservation strategies. The main aims of this study were to assess the biocultural value of Aloe in Kenya by documenting local uses of aloes and evaluating how the vernacular names reflect the relative importance in different ethnic groups. METHODS: Ethnobotanical and ethnotaxonomical data were collected using field observations and semi-structured interviews. Information was collected by interviewing 63 respondents from nine different ethnic groups, representing different ages, gender and occupations. Statistical analyses were performed using R version 3.1.2. RESULTS: A total of 19 species of Aloe were found in the study area, of which 16 were used. On the generic level Aloe was easily distinguished. At species level, the local and scientific delimitation were almost identical for frequently used taxa. Aloe secundiflora, with 57 unique use records was the most important species. The two most frequently mentioned Aloe treatments, were malaria and poultry diseases. In our study area neither age nor gender had a significant influence on the level of knowledge of Aloe use. Finally, no correlation was found between extent of use and people's perception of decrease in local aloe populations. The aloes are highly appreciated and are therefore propagated and transported over large areas when people relocate. CONCLUSION: Biocultural value is reflected in the ethnotaxonomy of Aloe in Kenya. Different ethnic groups recognise their most-valued Aloe at the genus level as "the aloe" and add explanatory names for the other species, such as the "spotted aloe" and the "one-legged aloe". Widespread species of Aloe have the highest number of uses. There is no obvious correlation with high use and decrease in abundance of aloes locally, and we found no compelling evidence for local uses causing devastating damage to populations of the 19 species in use, whereas habitat loss and commercial harvesting appear to be of urgent concern for these important plants.

Evolutionary history and leaf succulence as explanations for medicinal use in aloes and the global popularity of Aloe vera.

BACKGROUND: Aloe vera supports a substantial global trade yet its wild origins, and explanations for its popularity over 500 related Aloe species in one of the world's largest succulent groups, have remained uncertain. We developed an explicit phylogenetic framework to explore links between the rich traditions of medicinal use and leaf succulence in aloes. RESULTS: The phylogenetic hypothesis clarifies the origins of Aloe vera to the Arabian Peninsula at the northernmost limits of the range for aloes. The genus Aloe originated in southern Africa ~16 million years ago and underwent two major radiations driven by different speciation processes, giving rise to the extraordinary diversity known today. Large, succulent leaves typical of medicinal aloes arose during the most recent diversification ~10 million years ago and are strongly correlated to the phylogeny and to the likelihood of a species being used for medicine. A significant, albeit weak, phylogenetic signal is evident in the medicinal uses of aloes, suggesting that the properties for which they are valued do not occur randomly across the branches of the phylogenetic tree. CONCLUSIONS: Phylogenetic investigation of plant use and leaf succulence among aloes has yielded new explanations for the extraordinary market dominance of Aloe vera. The industry preference for Aloe vera appears to be due to its proximity to important historic trade routes, and early introduction to trade and cultivation. Well-developed succulent leaf mesophyll tissue, an adaptive feature that likely contributed to the ecological success of the genus Aloe, is the main predictor for medicinal use among Aloe species, whereas evolutionary loss of succulence tends to be associated with losses of medicinal use. Phylogenetic analyses of plant use offer potential to understand patterns in the value of global plant diversity.

Biologia reprodutiva e ausência de frutificação de Aloe saponaria (Aiton) Haw. (Xanthorrhoeaceae) fora do local de origem / Reproductive biology and absence of fruiting of Aloe saponaria (Aiton) Haw. (Xanthorrhoeaceae) outside its place of origin

RESUMO O gênero Aloe, originário principalmente da África, tem atualmente uma ampla distribuição no mundo. No entanto, são poucas as regiões que têm realizado estudos quanto ao sistema reprodutivo. O objetivo do presente trabalho foi analisar as características e o comportamento reprodutivo de Aloe saponaria em Florianópolis, Santa Catarina. Foram conduzidos estudos sobre sua morfologia e biologia floral, visitantes florais e sistema reprodutivo. Esta espécie apresentou uma inflorescência por planta, com um comprimento de 105 ± 0,1 cm e 267 ± 92,7 flores. A razão pólen/óvulo sugere que a espécie é xenogâmica. O volume e concentração de sólidos solúveis totais do néctar potencial foi 16,6 ± 6,3 μL e 22 ± 2,4 °Brix respectivamente. O néctar instantâneo não apresentou diferenças significativas nos períodos avaliados (9:00h e 15:00h) e o estigma permaneceu receptivo até o segundo dia após a antese. Foram coletados 110 insetos visitantes florais, dos quais 61,8% foram indivíduos de Trigona spinipes. Entretanto, nos testes de polinização não foi observada frutificação efetiva, indicando que a propagação vegetativa é o principal tipo de reprodução usado nessa população. Isto pode estar relacionado a um mecanismo de autoincompatibilidade esporofítica, a anormalidades cromossômicas durante a formação do pólen, as condições climáticas, e a escassa variabilidade genética no local de estudo.

ABSTRACT The Aloe genus, originating mainly from Africa, currently has a wide distribution in the world. However, in few regions studies about the reproductive system have been carried on. The aim e of this study was to analyze the characteristics and reproductive performance of the Aloesaponaria in Florianópolis, Santa Catarina. The morphology, floral biology, flower visitors and the reproductive system were determined. The plants presented an inflorescence per plant, with 105 ± 0,1 cm in length and 267 ± 92.7 flowers. The pollen/ovule ratio suggested that the species is xenogamic. The volume and concentration of total soluble solids in the potential nectar were 16.6 ± 6.3 μL and 22 ± 2.4°Brix, respectively. The instant nectar showed no significant differences between the evaluated periods (9:00h and 15:00h) and the stigma remained receptive until the second day the after anthesis. 110 insects were collected, from which 61.8% were from theTrigona spinipesspecies. However, in the pollination tests the fruit set was not observation, indicating that vegetative propagation is the main type of reproduction used by this population. This may be related to a mechanism of sporophytic self-incompatibility, to chromosomal abnormalities during the formation of pollen, to weather conditions, and to the low genetic variability at the study site.

Cloning and sequence analysis of an actin gene in aloe.

Aloe (Aloe spp), containing abundant polysaccharides and numerous bioactive ingredients, has remarkable medical, ornamental, calleidic, and edible values. In the present study, the total RNA was extracted from aloe leaf tissue. The isolated high-quality RNA was further used to clone actin gene by using reverse transcription-polymerase chain reaction (RT-PCR). The result of sequence analysis for the amplified fragment revealed that the cloned actin gene was 1012 bp in length (GenBank accession No. KC751541.1) and contained a 924-bp coding region and encoded a protein consisting of 307 amino acids. Homologous alignment showed that it shared over 80 and 96% identity with the nucleotide and amino acid sequences of actin from other plants, respectively. In addition, the cloned gene was used for phylogenetic analyses based on the deduced amino acid sequences, and the results suggested that the actin gene is highly conserved in evolution. The findings of this study will be useful for investigating the expression patterns of other genes in Aloe.

A comparative study of three Aloe species used to treat skin diseases in South African rural communities.

OBJECTIVES: Aloe species have been noted to be a miracle cure used by indigenous people of southern Africa. Geographically, each of the three Aloe species-Aloe arborescens, Aloe excelsa, and Aloe ferox-has a specific habitat. Although some species overlap in geographical regions, the species most abundant in a region is most often utilized by indigenous people. All three species display similar curative properties, aiding in wound healing, cures against other skin ailments, and some systemic conditions. RESULTS: All three Aloe species indicated high inhibitory activity against all gram-positive bacteria under investigation. The ethanol extract was most effective and inhibited all gram-positive bacteria and two gram-negative bacteria (i.e., Proteus vulgaris and Escherichia coli). CONCLUSIONS: All fungal species under investigation were successfully inhibited by both the boiled water as well as the ethanol extract, substantiating the traditional usage of this species.

Assessment of genetic stability and instability of tissue culture-propagated plantlets of Aloe vera L. by RAPD and ISSR markers.

Efficient plantlet regeneration with and without intermediate callus phase was achieved for a selected genotype of Aloe vera L. which is sweet in test and used as a vegetable and source of food. Random amplified polymorphic DNA (RAPD) and inter simple sequence repeats (ISSR) marker assays were employed to evaluate genetic stability of plantlets and validate the most reliable method for true-to-type propagation of sweet aloe, among two regeneration systems developed so far. Despite phenotypic similarities in plantlets produced through both regeneration systems, the differences in genomic constituents of plantlets produced through intermediate callus phase using soft base of inflorescence have been effectively distinguished by RAPD and ISSR markers. No polymorphism was observed in regenerants produced following direct regeneration of axillary buds, whereas 80% and 73.3% of polymorphism were observed in RAPD and ISSR, respectively, in the regenerants produced indirectly from base of the inflorescence axis via an intermediate callus phase. Overall, 86.6% of variations were observed in the plantlets produced via an intermediate callus phase. The occurrence of genetic polymorphism is associated with choice of explants and method used for plantlet regeneration. This confirms that clonal propagation of sweet aloe using axillary shoot buds can be used for commercial exploitation of the selected genotype where a high degree of fidelity is an essential prerequisite. On the other hand, a high degree of variations were observed in plantlets obtained through indirect regeneration and thus cannot be used for the mass multiplication of the genotype; however, it can be used for crop improvement through induction of somaclonal variations and genetic manipulations.

[Identification of Aloe species by random amplified polymorphic DNA (RAPD) analysis].

Juice and integument of leaves of 3 Aloe species, Aloe vera, A. ferox and A. africana, are not allowed to be used as food according to the Pharmaceutical Affairs Law in Japan. On the other hand, whole leaves of A. arborescens can be used as food. The present study was designed to distinguish Aloe species by random amplified polymorphic DNA (RAPD) analysis. DNA was isolated from fresh and dried leaves of the 4 Aloe species. Five out of 32 different 10-mer primers examined were useful for analysis. By comparison of the characteristic bands of PCR products on agarose gel, it was possible to distinguish the 4 species. Thus, the botanical species of Aloe in commercial food products can be identified by RAPD analysis.

Status of certain additional over-the-counter drug category II and III active ingredients. Final rule.

The Food and Drug Administration (FDA) is issuing a final rule stating that the stimulant laxative ingredients aloe (including aloe extract and aloe flower extract) and cascara sagrada (including casanthranol, cascara fluidextract aromatic, cascara sagrada bark, cascara sagrada extract, and cascara sagrada fluidextract) in over-the- counter (OTC) drug products are not generally recognized as safe and effective or are misbranded. This final rule is part of FDA's ongoing OTC drug product review.

Efecto agudo del aloe vera linneo y ciclanthera pedata shrad en ratas diabéticas / Acute hypoglycemic effect of aloe vera linneo and cyclanthera pedata shrad on diabetic rats

Se compara el efecto hipoglicemiante y la toxicidad del gel gel Aloe vera linneo y del extracto de Ciclanthera pedata schrad en ratas hiperglicémicas y ratones normales utilizando un estudio randomizado, cohorte de inicio de tipo longitudinal, con seguimiento de 48 horas. Se utilizaron 36 ratas albinas "Wistar". Se dividió en dos grupos de 18 ratas c/u A y B, que a su vez se subdividieron en 3 grupos de 6 ratas c/u. Al grupo A se le administró 125 mg/Kg de peso de aloxano y al grupo B 100 mg/Kg de peso via subcutanea. Posteriormente se dio a los grupos A1 y B1 aloe 10 mg/Kg, A2 y B2 caihua 10 mg/Kg; por último A3 y B3 reciben aloe (4 mg.Kg) y Caihua (6 mg/Kg) juntos. Todos los extractos se dieron por vía intraperitoneal (IP). Además, se utilizó 24 ratones blancos normales para la prueba de Dosis Letal 50 (DL50) agudo, recibieron via IP la dosis de 1, 10, 100 y 1000 mg/Kg de peso de aloe, caihua y ambos. Para la DL50 no se reportó ninguna muerte dentro de las 72 horas siguientes. En los animales del grupo A no se observó efecto hipoglicemiante de los extractos. En los 3 subgrupos B se observó una disminución de los niveles de glucosa con relación a la glicemia basal; la estadística fue estadística significativa en los subgrupos B1 y B3 (p<0,05). El grupo B3 mostró efecto a partir de la 3ra hora, mientras los otros grupos lo conseguian a la 5ta o 7ma hora. Se demuestra la carencia de toxicidad del gel del Aloe vera linneo y el extracto de Ciclanthera pedata schrad a dosis aguda; también sus efectos hipoglicemiantes con mejor resultado al administrarlos juntos.