COSTA, M. P. da

GAVILANES, M. L.

LAMEIRA, O. A.

PINTO, J. E. B. P.

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COMUNICAÇÃOTISSUE CULTURE PROP AGATION OF Cephaelis ipecacuanha A.Richard:EFFECT OF GROWTH REGULATORS ON PLANTLET ROOTFORMATIONOSMAR ALVES LAMEIRA 1MARLY PEDROSO DA COSTA2JOSÉ EDUARDO BRASIL PEREIRA PINT03MANUEL LOSADA GAVILANES4ABSTRACT - Cephaelis ipecacuanha ARichard indole-ô-butyric acid, gibberellic acid, and activated(Rubiaceae) was propagated "in vitro" at 28 ± 1°C charcoal, root formation was initiated. Conditions forunder 16h light and 8h darkness. Shoots were grown 100% efficiency of root forrnation were found. Plantletsfrom internodal segments on basic Gamborg medium to grew to about 30mm height within five weeks. Thea height of 15mrn within three weeks. By switching the plantlets were then transferred to soil, where theysegments to Murashige-Skoog medium containing established.INDEX TERMS: Cephaelis ipecacuanha, medicinal plant; "in vitro ", growth regulatorEFEITO DE REGULADORES DE CRESCIMENTO NO ENRAIZAMENTO"DE PLANTULAS DE Cephaelis ipecacuanha A. Richard EM CULTURA DETECIDORESUMO - Cephaelis ipecacuanha A Richard(Rubiaceae) foi propagada "in vitro" a 28 ± 1°C numfotoperiodo de 16 h de luz e 8 h de escuridão. Os brotosforam crescidos de segmento internodal no meio básicode Gamborg a uma altura de 15 mm dentro de 3semanas. Na transferência para o meio básico deMurashige-Skoog contendo ácido indol butirico, ácidogiberelico, e carvão ativado, o enraizamento foraminiciadas. Condições de 100% de enraizamento foiencontrado. Plântulas cresceram por volta de 30 mm dealtura dentro de 5 semanas. As plântulas foram entãotransferidas para o solo, onde se aclimataram.TERMOS PARA INDEXAÇÃO: Cephaelis ipecacuanha, planta medicinal, "in vitro ", regulador de crescimentoCephaelis ipecacuanha A Richard, ipecac, is animportant medicinal plant native to Brazil. The roots ofthis plant contain emetine and cephaline as activeprincipIes and have been used for years as expectorants,emetics, and amoebacides (Gupta, 1971).lpecac seeds require at least six monhts forgermination. To increase rooting and root growth,propagation methods using root cuttings have beenestablished, but these methods are not very efficient(Gattoni, 1959; Kalyanasundaram, 1969). Also,propagation methods using tissue culture have beendeveloped (Ikeda et a1.l988; Yoshimatsu; Shimomura1991; Lameira; Costa; Pinto, 1994). Our laboratory hasstarted investigations to establish methods with higherefficiencies for plantlet formation of ipecac "in vitro".This paper describes the effect of growth regulators onplantlet formation of ipecac in tissue culture"In vitro" stock cultures of Cephaelis ipecacuanhafrom micropropagation of internode segments were used as asource for explants. The cells proliferated on a liquidmedium containing the macro and micro nutrientsand vitamins of Garnborg; Miller; Ojima (1968),1. Eng.Agr., Dr. EMBRAP A - Centro de pesquisa Agroflorestal da Amazônia Oriental, Caixa Postal 48 - Belém,PA, 66095-100 - Brasil.2. Profa. do Departamento de Ciências Biológicas da UFP A - Belém, PA, 66095-100 - Brasil.3. Prof. do Departamento de Agricultura da UNlVERSIADE FEDERAL DE LAVRAS (UFLA) - Caixa Postal 37 -37.200-000 - Lavras - MG.4. Prof. do Departamento de Biologia da UFLA - Lavras - MG.391supplemented with 0.67 ~ benzylaminopurine and2% sucrose. The cultures were maintained in a growthchamber at 28 ± 1° C in 16/8 h light/dark cycles with alight intensity of 25 umol.m'ê.s" irradiance.The culture medium used for shoot elongation ofthe rosette-type was B5 mineral salts and vitamins(Gamborg; Miller; Ojima, 1968) supplemented with0.27, 0.54 or 0.82 ~ gibberellic acid. The mediumwas adjusted to pH 5.8 prior to autoclaving at 108 kpaat 121°C for 15 mino Culture tubes contained 10 ml ofthe medium.After five weeks, the elongated shoots over 10mm in lenght were separated individually and weretransferred to 100 ml Erlenmeyer flasks containing30 ml of Murashige and Skoog (1962) mineral saltsand vitamin medium supplemented with 1.48, 4.92, or14.76 ~ indole-3-butyric acid, 0.82 ~ gibberellicacid, and 0.1 or 0.3 % activated charcoal. All mediacontained 3 % (w/v) sucrose were solidified with 0.7 %(w/v) agar. After two weeks on this medi um, forrooting the elongated shoots were transferred to half-strength Murashige-Skoog medium without growthregulators and maintained as described above. Datawere submitted to analysis of variance and means werecompared by Duncan's multiple range test at O. 1 %level. Each experiment was repeated threetimes with 10 replications per treatment and fiveshoots per repetition.For shoot elongation all treatments were efficientHowever, the medium containing 0.82 ~ gibberellic acidwas the most effective. Within three weeks of inoculation,the shoots grew to 15 mm height and showed twoleaf1ets. Crozier (1981) showed that shoot elongationwas best with adequate levels of gibberelin (1,0mg/l). In our study, the levels of gibberelin testedwere within the concentrations used for shootelongation by others ( Paleg, 1965; Mertz, 1966).Occasionally, a low levei of gibberelin may be usefulsince it will permit some growth extension (Kriokorian,1982).The synergistic effect of indole-3-butyric acid,gibberelin, and activated charcoal in inducing plantletformation is shown in Table 1. Mertz (1966) reportedthat root elongation is stimulated when the levels ofauxin and gibberelin are balanced. After five weeks ofculturing, our results revealed that the best conditionsfor plantlet formation were 4.92 ~ indole-3-butyricacid, 0.82 ~ gibberellin, and 0.1 % activatedcharcoal. Plantlets showed 100% root formation andgrew to a height of about 30 mm. All media containing4.92 ~ indole-3-butyric acid were more effective inroot formation than media containing 1.48 or 14.76 ~indole-3-butyric acid, although the plantlet height wasnot affectedby this hormone. It is possible that the rootcells are more sensitive to auxin than the stem cells.The inhibition of root formation by high concentrationsof auxin was observed in intact or in cut roots (Evans,1976).The medium containing only activated charcoalwas the least effective for plantlet formation. Thepresence of activated charcoal can have inhibitory orstimulatory effects on growth "in vitro" (Fridborg et aI.1978). For plantlet formation, the inhibitory effect wasonly observed in media containing this adsorbent alone.TABLE 1 - Synergistic effect of IBA(indole -3-butyric acid), GA(gibberelin) and activated charcoal (AC) onplantlet formation of ipecac after five weeks in culture.mA GA AC Rooting Average number of Average lenght Average height(~M) (~M) (%) (%) roots/segments root (mm) plantlet (mm)1.48 0.82 0.3 20 1.2 c 5.6 a 25 b4.92 0.82 0.3 80 3.2 b 5.8 a 20 c4.92 0.82 60 2.1 c 4.8 ab 25 b4.92 0.82 0.1 100 15.5 a 7.4 a 30 a14.76 0.82 0.1 50 1.6 c 4.2 ab 30 a14.76 0.82 0.3 30 1.5 c 2.0 b 25 b0.3 10 0.9 c 2.0 b 7dValues within each row followed by the same letter are not significantly different (P = 0.01), by Duncan'sMultiple Range Test.Ciênc. e Agrotec., Lavras, v.21, n.3, p.390-392, jul./set., 1997392However, the combmation of activated cnarcoal wnngrowth regulators is stimulatory and synergistic.Plantlets removed from culture tuhes andmaintained seven to nine days in shade conditions, thentransferred to natural conditions, survived and grewwell (100%). Thus, the method descrihe here is usefulfor the fast and efficient propagation of ipecac.ACKNOWLEDGMENTWe thank Dr. Herrmann M. Klaus (PurdueUniversity - USA) and Dr" Cherry Kitto (DelawareUniversity - USA) for their corrections of the languagestyle and critical comments on the manuscript.REFERENCESCROZIER, A. Aspects of metabolism and physiology ofgibberellins. Advance Botany Research, NewYork, v.9, n.I, p.33-149, Feb.,I981.EVANS, M.L. A new sensitive root auxanometer:Preliminary studies of the interaction of auxin andacid pH in the regulation of intact root elongation.Plant Physiology, Washington, v.58, n.4, p.599-601, Oct.,I976.FRIDBORG,G.; PEDERSON,M.; LANDSTROM,L.;AND ERIKSSON, T. The effect of activatedcharcoal on tissue cultures: Absorptionof metabolitesinhibiting morphogenesis. Physiology Plantarum,Copenhagen,v.43, n.I, p.104-I06, May,I978.GAMBORG,O.L.; MILLER, R.A. AND OJIMA,K.Nutrient requirements of suspension cultures ofsoybean root cells. Experiment Cell Research,London, v.50, n.I, p.I51-158, Feb., 1968.GATONNI, L.A. La raicilla a ipecacuanha. In:BOLETIM do Ministério da Agricultura comércio eIndústria Panamá, 1959, p.I-32.GUPTA, R. lpecac a promissmg subsiãlary crop fornorth-eastern plantation regions. Indian Fanning,Nova Delhi, v.21, n.4, p.19-2I, July, 1971.IKEDA, K.; TESHIMA, T.; AOYAMA,M. ANDSHIMONURA,K. Clonal propagation of Cephaelisipecacuanha. Plant Cell Report, New York, v.7,n.I, p.288-291, Feb., 1988.KALYANASUNDARAM, S. Effect of boron andindole-butyric acid on rooting of ipecac rootcuttings. Madras Agricultura Journal, Coimbatore,v.56, n.6, p.812-820, June, 1969.KRIOKORIAN, A Cloninghigher plants from asepticallycultured tissues and cells. Biology Review, NewYork, v.57, n.I, p.151-218, Feb., 1982.LAMElRA, O.A; COSTA, M.P. da.; PINTO, JE.B.P.The efficiency of shoot and plantlet formation ofCephae/is ipecacuanha after three subcultures "invitro ". Ciência Rural, Santa Maria, v.24, n.3,p.523-526, set./dez. 1994.MERTZ, D. Hormonal control of root growth. PlantCell Physiology, Osaka, v,7, n. 1, p.125-135, Feb.,1966.MURASHIGE, T. AND SKOOG, F. A revised mediumfor rapid growth and bioassay with tobacco tissuecultures. Physiology Planta rum, Copenhagen,v.15, n.2, p.473-497, April, 1962.PALEG, L.G. Physiological effects of gibherellins.Annual Review Plant Physiology, Paio Alto, v.18,p.291-322, 1965.YOSHIMATSU,K. AND SHIMOMURA, K. Efficientshoot formation on internodal segmets and alkaloidformation in the regeneratesof Cephae/is ipecacuanhaA. Richard. Plant Cell Report, New York, v.9, n.5,p.567-570, Sept., 1991.Ciênc. e Agrotec., Lavras, v.21, n.3, p.390-392,jul./set., 1997

Tissue culture propagation of Cephaelis ipecacuanha A. Richard: effect of growth regulators on plantlet root formation.

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Tissue culture propagation of Cephaelis ipecacuanha A. Richard: effect of growth regulators on plantlet root formation.

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