Effects of Dimethyl Sulfoxide and Ethanol on Adventitious Root Growth of Panax quinquefolium and Ginsenosides / 中国实验方剂学杂志

Objective:To screen out the suitable nonpolar molecular cosolvent and concentration with adventitious root phenotype and ginsenoside content in the controlled experiment as the evaluation indexes， so as to lay a solid foundation for exploring the causes for good shape and high quality of <italic>Panax quinquefolium</italic>. Method:After being treated with different concentrations of dimethyl sulfoxide （DMSO） and ethanol， the adventitious roots were scanned using a panoramic scanner， and the resulting images were used for measuring the branch number and average diameter by WinRHIZO Pro 2016， Synbiosis ProtoCol 3 colony counter， Image J， and SmartRoot. The contents of ginsenosides Rg₁， Rb₁， and Re were determined by ultra-performance liquid chromatography tandem mass spectrometry （UPLC-MS/MS）. Result:Compared with the blank control， the 0.1% DMSO and 75% ethanol made the adventitious root phenotype and ginsenoside contents significantly changed. Specifically， the branch number and average diameter were significantly reduced. The ginsenoside Rg₁ in the adventitious roots decreased after 0.1% DMSO treatment， whereas the ginsenosides Rg₁ and Re increased after 75% ethanol treatment. The adventitious root phenotype and ginsenoside contents in the 0.1% DMSO treatment group were not significantly different from those in the control group. Conclusion:The 0.01% DMSO does not affect the adventitious root growth of <italic>P. quinquefolium </italic>and is insoluble in water， enabling it to be considered as a suitable nonpolar molecular cosolvent for future research on the genetic causes for the good shape and high quality of <italic>P. quinquefolium</italic>.

Morphology, germination, and geographic distribution of Pentaclethra macroloba (Fabaceae): a hyperdominant Amazonian tree / Morfología, germinación y distribución geográfica de Pentaclethra macroloba (Fabaceae): árbol amazónico hiperdominante

Introduction: Pentaclethra macroloba is a hyperdominant tree in the Amazon estuary of great socioeconomic importance for the region because the oil from its seeds is a powerful herbal medicine. Objective: We aimed to characterize the morphological structure, the morphological adaptations in response to the daily flooding of the Amazon estuary and the biogeographic area of P. macroloba. Methods: Detailed description of the external morphology, from germination to the adult tree, was obtained from individuals located in floodplain forest, Northeast Amazonia. The occupation area and the geographical extension of P. macroloba were determined from point of geographical coordinates of botanical samples deposited in the digital collections of Mobot and SpeciesLink. Results: Adult individuals have adapted structures in response to daily flooding, such as: adventitious roots to increase respiratory efficiency and lenticels in the trunk, serving as a connection point for oxygen transport between the root and the aerial part. Dried fruit favours the activation of the explosive dehiscence mechanism, allowing the seed to be expelled long distance. Deltoid shape of the seed allows water fluctuation and more efficient dispersal. Seedling is phanerocotylar hypogeal and with one pair of reserve cotyledons that provide the seedling an extra source of energy to escape the flood. Germination rate was 78 % and the speed was 0.2 seeds.day-1. Circular buffer method revealed the presence of 123 subpopulations of P. macroloba distributed in a radius of 5 699 943 km² across the Neotropical region. Conclusions: Much of the morphological structures of P. macroloba are adaptive and evolutionary responses to the periodically flooded environment of the Amazon estuary, showing that these environments select the trees, best adapted, to inhabit the flood. P. macroloba has a wide geographical area denotes the plasticity of adapting to different environments, which may justify its monodominance in some regions.

Introducción: Pentaclethra macroloba es un árbol hiperdominante en el estuario del Amazonas, de gran importancia socioeconómica para la región, pues el aceite de sus semillas es un poderoso medicamento natural. Objetivo: Nuestro objetivo fue caracterizar la estructura morfológica, las adaptaciones morfológicas en respuesta a las inundaciones diarias del estuario del Amazonas y el área biogeográfica de P. macroloba. Métodos: Una descripción detallada de la morfología externa, desde la germinación hasta el árbol adulto, se obtuvo de individuos ubicados en el bosque periódicamente inundado, al noreste de la Amazonia. El área de ocupación y la extensión geográfica de P. macroloba se determinaron a partir del punto de coordenadas geográficas de muestras botánicas depositadas en las colecciones digitales de Mobot y SpeciesLink. Resultados: Los individuos adultos tienen estructuras adaptadas en respuesta a las inundaciones diarias, tales como: raíces adventicias para aumentar la eficiencia respiratoria y lenticelas en el tronco, que sirven como punto de conexión para el transporte de oxígeno entre la raíz y la parte aérea. Los frutos secos favorecen la activación del mecanismo de dehiscencia explosiva, permitiendo que la semilla sea expulsada a larga distancia. La forma deltoidea de la semilla permite la fluctuación en el agua y una dispersión más eficiente. La plántula es hipogel fanerocotiledones y con un par de cotiledones de reserva que proporcionan a la plántula una fuente extra de energía para escapar de la inundación. La tasa de germinación fue del 78 % y la velocidad fue de 0.2 semillas día-1. El método Circular buffer reveló la presencia de 123 subpoblaciones de P. macroloba distribuidas en un radio de 5 699 943 km² a lo largo de la región neotropical. Conclusiones: Gran parte de las estructuras morfológicas de P. macroloba son respuestas adaptativas y evolutivas al ambiente periódicamente inundado del estuario del Amazonas, lo que demuestra que estos ambientes actúan como filtro ambiental seleccionado las especies mejor adaptadas al medio. Su amplia área geográfica denota la plasticidad de adaptarse a diferentes ambientes, lo que puede justificar su monodominancia en algunas regiones.

Advances in Bioculture Technology of Bupleuri Radix and Biosynthesis Pathways of Saikosaponins / 中国实验方剂学杂志

Synthetic biology is an emerging discipline that analyzes the biosynthesis pathways of active constituents in traditional Chinese medicine and explores genes involved in biosynthesis. Bupleuri Radix is one of the most commonly used Chinese medicinal materials with remarkable medicinal value, its index component is saikosaponins, which has significant anti-inflammatory, anti-viral and anti-tumor activities. However, the current wild resources of Bupleuri Radix have been destroyed, and there were some problems in the process of artificial cultivation. The application of biological culture technology and synthetic biology can expand the sources of saikosaponins and protect resources of Bupleuri Radix. The culture conditions of different plants can be followed without a fixed pattern, and the biosynthetic pathways of different medicinal active ingredients are also inconsistent. At present, there is no review report on the culture technology of Bupleuri Radix and the research on the biosynthesis pathway of saikosaponins. This paper introduces the research progress of biological culture techniques, such as callus culture, adventitious root culture, hairy root culture and suspension cell culture used in synthetic biology, and the biosynthesis pathway of saikosaponins and its key enzyme functional genes. It is suggested to optimize the biological culture technology of Bupleuri Radix by referring to the tissue culture technology of other traditional root medicinal materials, so as to provide a reference for the in-depth study on the biosynthesis pathway and metabolic regulation of saikosaponins.

Microbial transformation of ginsenosides extracted from Panax ginseng adventitious roots in an airlift bioreactor

Background: Ginsenoside is the most important secondary metabolite in ginseng. Natural sources of wild ginseng have been overexploited. Although root culture can reduce the length of the growth cycle of ginseng, the number of species of ginsenosides is reduced and their contents are lower in the adventitious roots of ginseng than in the roots of ginseng cultivated in the field. Results: In this study, 147 strains of ß-glucosidase-producing microorganisms were isolated from soil. Of these, strain K35 showed excellent activity for converting major ginsenosides into rare ginsenosides, and a NCBI BLAST of its 16S rDNA gene sequence showed that it was most closely related to Penicillium sp. (HQ608083.1). Strain K35 was used to ferment the adventitious root extract, and the fermentation products were analyzed by high-performance liquid chromatography. The results showed that the content of the rare ginsenoside CK was 0.253 mg mL-1 under the optimal converting conditions of 9 d of fermentation at pH 7.0 in LL medium, which was significantly higher than that in the adventitious roots of ginseng. Conclusion: These findings may not only solve the problem of low productivity of metabolite in ginseng root culture but may also result in the development of a new valuable method of manufacturing ginsenoside CK.

Identification of WRKY Transcription Factors Related to Saikosaponin Biosynthesis in Adventitious Roots of Bupleurum chinense / 中草药·英文版

Objective To identify the genes of WRKY transcription factors (TFs) from roots of Bupleurum chinense and genes that potentially regulate saikosaponin (SS) biosynthesis. Methods Firstly, the subfamily cluster analysis was mainly based on Arabidopsis thaliana WRKYs for 27 putative WRKY TFs selected from previous transcriptome sequencing data. Secondly, qPCR was used to screen such genes of WRKY TFs that could be induced by NaCl and PEG6000 in adventitious roots of B. chinense. Meanwhile, saikosaponins (SSs) in treated adventitious roots were determined by HPLC. The roots were collected at 0, 2, 4, 8, 12, 24, 48, and 72 h after treatments, and 120 h only for PEG. Finally, the tissue-specific expression was analyzed on screened genes by qPCR. Results The 27 genes were grouped into three categories: There were nine in Group I, 15 in Group II, and two in Group III. Four genes of WRKYTFs, BCWRKY6, BCWRKY16, BCWRKY32, and BCWRKY35 were obviously induced by NaCl in adventitious roots of B. chinense, while only BCWRKY32 was induced by PEG. The content of SSs increased at different levels in NaCl and PEG6000 treatment. Three genes including BCWRKY6, BCWRKY32, and BCWRKY35, expressed most in roots, were similar to the accumulation pattern of SS. Conclusion The three WRKY genes, BCWRKY6, BCWRKY32, and BCWRKY35, may be involved in the biosynthesis of SS.

Enraizamento de estacas de Sebastiania schottiana Müll. Arg. / Rooting of Sebastiania schottiana Müll. Arg. cuttings

O pouco domínio dos processos envolvidos no enraizamento de estacas de diversas espécies florestais tem sido um fator limitador para a produção de mudas via propagação vegetativa, impedindo possíveis ganhos econômicos e ecológicos nas atividades de reflorestamento e recuperação de áreas degradadas. O objetivo deste estudo foi verificar o potencial de enraizamento de estacas de Sebastiania schottiana, submetidas a diferentes doses e tempos de imersão em produto comercial Orgasol® FTS (composto de aminoácidos e micronutrientes). Estacas basais, medianas e apicais tiveram suas bases imersas em Orgasol® FTS nas doses de 0; 5; 10 e 15mL L-1, durante 1 minuto. O delineamento experimental foi realizado em blocos ao acaso, com três repetições de 20 estacas por parcela. Aos 75 dias, avaliou-se a porcentagem de estacas enraizadas, o comprimento e diâmetro da raiz mais longa. Em um segundo experimento, estacas basais e medianas foram tratadas em 15mL L-1 do produto, durante 0, 15, 30 e 45 minutos. O delineamento experimental foi realizado em blocos ao acaso, com três repetições e 20 estacas por parcela. Aos 150 dias, avaliou-se a porcentagem de estacas enraizadas, massa seca das raízes e o comprimento e diâmetro da raiz mais longa. No primeiro experimento, o aumento da dose do produto promoveu um maior potencial de enraizamento em todos os tipos de estacas, com maior porcentagem de estacas enraizadas no tratamento com 15mL L-1 do produto. No segundo experimento, os maiores potenciais de enraizamento foram observados em estacas basais e medianas tratadas com 15mL L-1 do produto, por 45 minutos. O uso do produto comercial Orgasol® FTS favoreceu a indução de raízes adventícias em estacas de Sebastiania schottiana.

The aim of this study was to verify the rooting potential of Sebastiania schottiana cuttings treated with different doses and immersion times in Orgasol® FTS (composed with amino acids and micronutrients). In a first trial, basal, median and apical cuttings had their bases immersed in Orgasol® FTS at levels of 0, 5, 10 and 15mL L-1, for 1 minute. The experimental design was in random blocks, with three replicates and 20 cuttings per plot. At 75 days, the rooted cuttings percentage, the length and diameter of the longest root were evaluated. In a second trial, basal and median cuttings were treated for 0, 15, 30 and 45 minutes in 15mL L-1 of Orgasol® FTS. The experimental design was in random blocks, with three replicates of 20 cuttings per plot. At 150 day, the rooted cuttings percentage, dry roots mass and the length and diameter of the longest root were evaluated. In the first trial, higher levels of product was beneficial to rooting, it was observed the largets rooted cuttings percentage in the treatment with 15mL L-1 of product. In second trial, the higher rooting potential was observed in the basal and median cuttings treated by 45 minutes in 15mL L-1 of product. The use of orgasol® FTS promotes adventitious roots in Sebastiania schottiana cuttings.