Cultivo in vitro de raíces pilosas del arbusto Phyllanthus acuminatus (Phyllanthaceae) / In vitro culture of hairy roots of the shrub, Phyllanthus acuminatus (Phyllanthaceae)

Introducción: En condiciones naturales, las raíces del arbusto, Phyllanthus acuminatus, producen bajas concentraciones de metabolitos secundarios de interés medicinal. Esto abre una oportunidad para el cultivo in vitro, para aumentar la concentración de metabolitos. Objetivo: Determinar las condiciones óptimas de cultivo líquido para raíces pilosas de P. acuminatus. Métodos: Se utilizó la evaluación del crecimiento de la biomasa según porcentaje de inóculo inicial (0.50 y 0.10 %), porcentaje de nutrientes de los medios (100, 50 y 25 %) y tasa de agitación (90, 100 y 110 min-1) (N= 15 repeticiones). Resultados: Las mejores condiciones de cultivo líquido fueron: 0.10 % de inóculo inicial, nutrientes al 25 % y 90 min-1 para la tasa de agitación. Hay diferencias entre las raíces pilosas y las raíces no transformadas. Conclusiones: es factible producir raíces pilosas de P. acuminatus a gran escala, aplicando e implementando las condiciones evaluadas de porcentaje de inóculo, nutrientes en el medio y tasas de agitación utilizadas en este estudio.

Introduction: Under natural conditions, the roots of the shrub, Phyllanthus acuminatus, produce low concentrations of secondary metabolites of medicinal interest. This opens an opportunity for in vitro culture, to increase metabolite concentration. Objective: To determine the optimal liquid culture conditions for hairy roots of P. acuminatus. Methods: We used biomass growth evaluation according to initial inoculum percentage (0.50 and 0.10 %), percentage of medium nutrients (100, 50 and 25 %) and agitation rate (90, 100 and 110 min-1) (N=15 replications). Results: The best liquid culture conditions were: 0.10 % of initial inoculum, nutrients at 25 % and 90 min-1 for the agitation rate. There are differences among hairy roots and non-transformed roots. Conclusions: It is feasible to produce P. acuminatus hairy roots at a large scale, applying and implementing the evaluated conditions of inoculum percentage, nutrients in the medium and agitation rates.

Densidade de plantio e idade de colheita de quebra-pedra [Phyllanthus amarus (Schumach. & Thonning) genótipo Unicamp-CPQBA 14] na produtividade de filantina / Density of planting and age of Phillanthus amarus genotype UNICAMP/CPQBA-14 on the productivity of phyllanthin

Avaliou-se o efeito da densidade de plantio e da idade de Phyllanthus amarus CPQBA-14, sobre o teor e produtividade de filantina. O delineamento experimental foi em blocos ao acaso, em esquema fatorial 4 x 6, com cinco repetições, envolvendo 4 tratamentos de densidade de plantio (400.000 pl ha -1; 200.000 pl ha -1; 100.000 pl ha -1 e 66.667,68 pl ha -1) com 6 colheitas (30, 45, 60, 75, 90 e 105 dias após o transplante (DAT)). Os dados foram submetidos a Análise de Variâncias e Regressão (P > 0.001). Para o teor de Filantina, foram observados efeitos significativos e independentes dos tratamentos de idade da planta na colheita e densidade de plantio. O teor de filantina aumentou linearmente com a idade da planta, atingindo o máximo aos 105 DAT, com teor de 11,52 g Kg-1. O adensamento populacional estimado que proporcionou máximo teor de filantina (8,66 g kg-1) foi de 299.860 pl ha-1. A produtividade de filantina apresentou interação significativa entre densidade de plantio e idade da planta na colheita. Observou-se que ao longo do crescimento da planta, os tratamentos com 200 e 400.000 pl ha -1 apresentaram as maiores produtividades de filantina (39,3 e 37,8 kg ha-1) aos 97,12 e 95,17 DAT, respectivamente. Conclui-se que o espaçamento apropriado para o melhor arranjo espacial de cultivo é de 299.860 pl ha-1 para as produtividades de 3.974,19 Kg (massa de matéria seca foliar) x 0,00866 kg (teor de filantina) = 34,416 kg ha-1 de filantina, com colheita programada aos 97 DAT.

The effect of planting density and age of Phyllanthus amarus- genotype CPQBA-14, on the content and productivity of phyllanthin were evaluated. The experimental design was randomized blocks in factorial scheme 4 x 6, with five replicates, consisting of 4 treatments of planting density (400.000 pl ha-1; 200.000 pl ha-1; 100.000 pl ha-1 and 66667.68 pl ha-1) with six ages of plants (30, 45, 60, 75, 90 and 105 days after transplanting (DAT)). Data were subjected to analysis of variance and regression (P>0.005). For phyllanthin content independent and significant effects of harvest and planting density were observed. The phyllanthin contents increased with plant age, peaking at 105 DAT, with a content of 11.52 g kg-1. The estimated population density that provided the high phyllanthin contents (8.66 g kg-1) was at 299.860 pl ha-1. The productivity of phyllanthin showed significant interaction between planting density and plants ages. It was observed during the plant growth, the treatments with 200 and 400.000 pl ha -1 had the highest yield of phyllanthin (39.3 and 37.8 kg ha-1) with 95.17 and 97.12 DAT, respectively. We conclude that the proper spacing for best spatial arrangement of the cultivation is 299.860 pl ha-1 for the yield of 3974.19 kg (dry weight leaf) x 0.00866 kg (phyllanthin contents) = 34.416 kg ha-1 of phyllanthin with harvest scheduled at 97 DAT.

High frequency in vitro propagation of Phyllanthus amarus Schum. & Thom. by shoot tip culture.

With the aim of micropropagation of Phyllanthus amarus, an important medicinal herb, shoot tips were cultured in Murashige and Skoog's medium supplemented with kinetin/ BAP singly or in combination with IAA. Growth regulators at lower range (0.1-1.0 mg L(-1)) stimulated direct regeneration of shoots. Kinetin was superior to BAP and kinetin-IAA combination was more suitable than kinetin alone. About 15 shoots were yielded per explant after 30 days of culture in the medium containing kinetin and IAA both at 0.1mg L(-1). The cluster of proliferated shoots elongated and rooted simultaneously under the same treatment following another subculture, thus shortening the total time schedule of micropropagation. Shoot tips of regenerated shoots were continuously used to regenerate new shoots with periodic transfer to fresh medium resulting in a steady supply of normal, healthy plants without any deviation in the production rate during a continuous one year culture. Micropropagated plants were successfully established in soil with high survivality (80%).