Types of cuttings and treatment with indolebutyric acid for propagation of Lippia alba (linalool chemotype)

ARTÍCULO ORIGINAL

 

Types of cuttings and treatment with indolebutyric acid for propagation of Lippia alba (linalool chemotype)

 

Tipos de estacas y tratamiento con ácido indolbutírico para la propagación de Lippia alba quimiotipo linalool

 

 

Erik Nunes Gomes
Leandro Marcolino Vieira
Maíra Maciel Tomazzoli
Cíntia de Moraes Fagundes
Rafaela Cristina Brunetti Machado
Katia Christina Zuffellato-Ribas
Cícero Deschamps

Departamento de Fitotecnia e Fitossanitarismo, Setor de Ciências Agrárias, Universidade Federal do Paraná, Brasil.

 

 

ABSTRACT

Introduction: Lippia alba (Mill.) NE Br. Ex P. Wilson, commonly known as bushy lippia, is a plant native to Brazil used to treat respiratory conditions as well as gastrointestinal and hepatic disorders.
Objective: Evaluate the survival, rooting and budding of different types of cuttings of L. alba (linalool chemotype) using indolebutyric acid (IBA).
Method: L. alba branches with leaves were collected in September 2016 in the city of Pinhais, Paraná, Brazil. Four types of cuttings were evaluated: apical herbaceous and thin, medium-thickness and thick semi-woody (0.25±0.06; 0.60±0.1; 1.2±0.2 cm in diameter, respectively) taken from the central section of the branch and without leaves. The bases of the different types of cuttings were submerged in three treatment substances: IBA (2 000 mg/l-1), hydroalcoholic solution (50 % v/v) and distilled water. The cuttings were planted in 120 cm3 plastic tubes filled with commercial substrate Tropstrato HT® and were kept under intermittent mist. At 30 days an evaluation was performed of the rates of survival, rooting and budding, the average number and length of roots, and the dry mass of roots and buds.
Results: No interaction was observed between the types of cuttings and the treatments applied. The four types of cuttings exhibited high survival and rooting percentages (averages above 86.7 %) and did not differ between themselves. Treatment with IBA fostered rooting with the highest percentages (98.1 %). Dry root mass was greater in apical herbaceous and thick semi-woody cuttings.
Conclusions: For propagation of L. alba (linalool chemotype), use may be made of apical herbaceous and thin, medium-thickness or thick semi-woody cuttings, which exhibit rooting percentages above 85 %. Treatment of the cuttings with IBA at 2 000 mg/l-1 improves the rooting percentage.

Key words: auxin, bushy lippia, ethanol, medicinal plant, rooting.

RESUMEN

Introducción: Lippia alba (Mill.) NE Br. Ex P. Wilson, conocida como hierba cidera, es una planta oriunda de Brasil utilizada para tratar trastornos gastrointestinales y hepáticos, y enfermedades respiratorias.
Objetivo: Evaluar la supervivencia, el enraizamiento y el brote de diferentes tipos de estacas de L. alba quimiotipo linalool con la aplicación de ácido indolbutírico (IBA).
Método: Se recolectaron ramas con hojas de L. alba en septiembre de 2016 en la ciudad de Pinhais, Paraná, Brasil. Se evaluaron cuatro tipos de estacas:esquejes herbáceos apicales y estacas semileñosas finas, medianas y gruesas, (0,25±0,06; 0,60±0,1; 1,2±0,2 cm de diámetro, respectivamente) sin hojas, tomadas del centro de la rama. Se sumergieron las bases de los diferentes tipos de estacas en 3 sustancias de tratamientos: IBA (2 000 mg×L-1), solución hidroalcohólica (50 % v/v) y agua destilada. Las estacas se plantaron en tubos de plástico de 120 cm³ llenos de sustrato comercial Tropstrato HT® y se mantuvieron bajo niebla intermitente. Después de 30 d se evaluaron las tasas de supervivencia,enraizamiento y brote, el número y la longitud promedio de las raíces y la masa seca de las raíces y de los brotes.
Resultados: No se comprobóinteracción entre los tipos de estacas y los tratamientos aplicados. Los cuatro tipos de estacas presentaron elevados por cientos de supervivencia y enraizamiento (promedios superiores a 86,7 %) y no se diferenciaron entre sí. El tratamiento con IBA favoreció el enraizamiento con los mayores porcentajes (98,1 %). La masa seca de las raíces fue mayor en las estacas herbáceas apicales y semileñosas gruesas.
Conclusiones: Para la propagación de L. Alba quimiotipo linalool se pueden utilizar estacas herbáceas apicales y semileñosas finas, medias y gruesas que muestran porcentajes de enraizamiento superiores a 85 %. El tratamiento de las estacas con IBA a 2 000 mg×L-1 es capaz de incrementar el porcentaje de enraizamiento.

Palabras clave: auxina; hierba cidera; etanol; planta medicinal; enraizamiento.

 

 

INTRODUCTION

Lippia alba (Mill.) NE Br. Ex P. Wilson (Verbenaceae), popularly known in Brazil as erva-cidreira, is a Brazilian native plant widely used by folk medicine to treat diseases related to gastrointestinal disorders, respiratory diseases and liver problems.1 It is a perennial bush or sub-bush very branched, measuring between 1.5 m and 2.0 m in height, with branches whitish, arched and brittle. The flowers are grouped in capituliform inflorescences with short axis, and with blue to violet color.2

The species can be used as a medicinal product, and also in the agrochemical industries due to its proven antifungal, antibacterial, antiviral, insecticidal, repellent and nematicidaleffects.3,4 In addition, it is a promising plant for the use in the pharmaceutical industry as a fixing agent for fragrances.1 These characteristics and bioactive properties are directly related to the composition of the essential oils present in the species.5

As for the essential oil, L. alba is a species characterized by great phytochemical diversity, mainly in the group of terpenes.6 In Brazil, concerning the major compounds found in the essential oil, the main chemotypes of L. alba are the chemotype I (citral, b-myrcene and limonene), chemotype II (citral and limonene) and chemotype III (carvone and limonene).2,7 Januzzi et al.,6 however, draw attention to the existence of other chemotypes such as citral and linalool.

Linalool, 3,7-Dimethyl-1,6-octadien-3-ol, is a monoterpene with antibacterial, anxiolytic and antifungal properties.8 This metabolite is widely used in the industrial synthesis of various vitamins and chemicals of fragrances, cosmetics, household cleaning products and fine perfumery due to its pleasant aroma.9

In the agronomic evaluation of nine genotypes of L. alba in southern Brazil, materials with high yields of essential oil were selected. Special attention was drawn to the accession "Brasília 2" which produced 5.45 liters per hectare of essential oil with 92.1 % of linalool.10 Given the potential for exploitation of this material as a source of linalool, studies are required to define the best practices for its cultivation and propagation.

Vegetative propagation, when used in medicinal plants, avoids abrupt changes in the content of active principles, maintaining the quality of the final product. Among the vegetative propagation methods, the propagation through stem cuttings is characterized by multiplying and preserving the varietal characteristics of the stock plants quickly and with greater economic viability. However, the technical aspects of rooting stem cuttings such as substrates, containers and plant regulators should be evaluated for each genotype.11

Although the species presents generally high endogenous potential for rooting, L. alba chemotypes differ in the rhizogenic potential of stem cuttings.12 In apical and basal stem cuttings of the myrcene-citral and citral-limonene chemotypes, the application of plant regulators was considered unnecessary, whereas for the carvone-limonene chemotype the application of 250 mg L-1 of indolebutyric acid (IBA) in apical cuttings was able to stimulate rooting.13-15 In the evaluation of different types of cuttings and substrates, the myrcene-citralchemotype showed higher dry mass of roots and shoots compared to citral-limonene and carvone-limonene chemotypes.7

Although the studies with the vegetative propagation of L. alba are relatively abundant,2,7,11-18 researches evaluating the best conditions for the rooting and formation of linalool chemotype seedlings are scarce.

Given the above, the aim of the present study was to evaluate the survival, rooting and sprouting of different types of L. alba linalool chemotype stem cuttings with the application of the plant growth regulator IBA.

 

METHODS

The plant material was collected in September 2016 from 10 stock plants cultivated at the medicinal, aromatic and condiment plants collection in the Center of Experimental Stations of Cangüiri, Federal University of Parana. The station is located in the city of Pinhais, Brazil, under the following coordinates: 25°23'30 "S and 49°07'30" W, with an altitude of 930 m. The climate of the region is temperate humid and classified as Cfb in the Köppen climatic classification system.19

The genotype "Brasília 2" was used in the experiment.A voucher specimen of the plant material was deposited in the Curitiba Botanical Museum under the code 375880. Branches with leaves of this genotype were moistened and conditioned in polypropylene bags for transportation to the Agrarian Sciences Sector of the Federal University of the Paraná, Curitiba, Brazil, where the experiment was conducted.

Before making the stem cuttings, the plant material was subjected to treatment with sodium hypochlorite at 0.5 % (v/v) for 15 minutes, followed by washing in running water for 5 minutes. Four types of stem cuttings were made: apical herbaceous cuttings and three types of semi-hardwood leafless cuttings from the median region of the branch. The apical cuttings were made with 7 cm length and with two leaves at the apex. The median semi-hardwood cuttings were made with 12 cm length and three different diameters: thin, medium and thick stem cuttings (0.25±0.06; 0.60±01; 1.2±0.2 cm diameter, respectively). All the cuttings were prepared with a bevel cut at the base and a straight cut at the apex.

The different types of stem cuttings had their bases immersed for 10 seconds in the following treatments: IBA (Vetec®) at the concentration of 2 000 mg× L-1 diluted in hydroalcoholic (ethanol) solution (50 % v/v), only hydroalcoholic solution (50 % v/v) and distilled water.

After the treatments the stem cuttings were planted in plastic tubes of 120 cm³ filled with Tropstrato HT® commercial substrate. According to information from the manufacturer, this substrate is composed by pine bark, peat, expanded vermiculite and enriched with macro and micronutrients. It has a pH of 5.8 and density in dry basis of 200 kg×m-³. The cuttings were kept in a greenhouse under intermittent mist (5 seconds every 30 minutes).

The experimental design was completely randomized in a 4x3 factorial scheme (4 types of stem cuttings and 3 treatments on the cuttings base), with 4 replicates and 10 cuttings as experimental unit.

After 30 days of planting, the variables of survival, rooting and sprouting percentages, number and average length of roots and dry mass of roots and sprouts were analyzed. The data were analyzed for the variances homogeneity by the Bartlett test and, since homogeneity was verified for all the variables, they were submitted to variance analysis (ANOVA). When they presented significant differences by the F test, the means were compared by the Tukey's test at 5 % probability. The statistical software Assistat ®20 was used to perform the analysis.

 

RESULTS

The variance analysis revealed no interaction between the types of cuttings and the treatments carried out at the base of the propagules, demonstrating that they are independent factors. Analyzing the factors alone, the application of IBA was only significant for the rooting percentage variable. The type of cutting influenced the number and dry mass of roots, sprouting percentage and the sprouts dry mass (Table 1).

The four types of cuttings presented high survival and rooting percentages (averages greater than 86.7 %) and did not differ among themselves. As for treatment at the cuttings base, the hydroalcoholic solution with 2000 mg×L-1 of IBA promoted higher rooting percentages (98.1 %) compared to the other treatments (Table 2).

The average number of roots in apical cuttings was superior in relation to the thin and medium cuttings, not differing from the thick ones. The roots dry mass was higher in apical herbaceous and thick semi-hardwoodcuttings (Table 3).

The sprouting percentage was lower in the thinsemi-hardwood cuttings in comparison with the others, which did not differ among themselves. Regarding the sprouts dry mass, the thick semi-hardwood cuttings had the highest values with an average of 0.31 g. In the Figure it is possible to observe the greater root development in herbaceous apical cuttings and greater development of the new shoots in thick semi-hardwood cuttings.

 

DISCUSSION

As previously described, L. alba can be considered a species easily propagated by stem cuttings, reaching more than 85 % rooting without IBA application. Several studies have also reported high percentages of survival and rooting in different types of stem cuttings without IBA use.2,11,12,18

Importantly, Lima et al.11 have related no differences withIBA application for this species, since the control treatment showed 100% of stem cuttings rooted. On the other hand, in the present study, IBA application increased rooting in cuttings of the linalool chemotype. A similar result was found by Albuquerque et al.13 for the L. alba citral-limonene chemotype, in which IBA treatments showed better results for rooting than the control treatment.

The exogenous application of auxins usually increases the percentage, speed, quality and uniformity of rooting in stem cuttings, through the activation ofthe vascular cambium cells, the stimulation of cell division and adventitious roots formation.21 This justifies the superiority of the IBA in relation to the treatments with water and hydroalcoholic solution in the present experiment.

The use of the hydroalcoholic solution treatment was based on reports that ethanol and other alcohols are carbon sources with capacity to supply the carbohydrate demands of the vegetative propagules, and through this, it can stimulate the adventitious rooting of stem cuttings.22-24 The dilution of plant regulators in alcoholic solution is usual and, consequently, it is difficult to define if the stimulus to the adventitious rhizogenesisis due to the regulator, ethanol or to a synergistic effect. For stem cuttings of L. alba linalool chemotype studied in the present experiment, the ethanol did not increase the rooting when compared to the treatment with distilled water, confirming that the stimulatory effects are attributed to IBA.

Although the types of cuttings did not affect rooting and survival percentages, differences were observed in root vigor, as a function of root number and dry mass. Similar results with L. alba were found by Biasi and Costa,12 the authors also evaluated different types of cuttings and found great ease in roots formation, with differences only in the vigor of the root system.

The apical cuttings superiority in relation to the thin and medium semi-hardwoodones, with respect to the number and dry mass of roots, can be attributed to the fact that the apical propagules have an herbaceous consistency, the presence of leaves and by the higher rates of endogenous auxins in stem apices.

Similarly to that observed in this experiment, for the propagation of the medicinal species Piper hispidum Sw., the proximity of the auxin producing sites, the presence of leaves and lower lignification of the tissues in apical stem cuttings promoted a greater vigor of the root system in relation to median ones.25 Also in that experiment, similarly to the present study, the types of P. hispidum stem cuttings and different substrates did not interact significantly and had to be analyzed independently.25

The presence of leaves in stem cuttings contribute to the rhizogenesis, both by the synthesis of carbohydrates through photosynthesis, and by the auxins and rooting cofactors supply.26 The presence of leaves on L. alba cuttings is reported by Biasi and Costa 12 and Coelho et al.18 as a favorable condition for better root system development.

This study did not present statistical differences between herbaceous apical and thick semi-hardwood cuttings for number and dry mass of roots. This can be explained by the greater amount of reserves present in the thick semi-hardwood cuttings, compensating the presence of leaves and the proximity of the auxin producing sites in apical ones. For roots dry mass, the superiority of thick semi-hardwood cuttings, in the same way, can be justified by the greater quantity of reserves.

The diameter of the cuttings can influence rooting, since thick cuttings have a higher content of carbohydrates, starch and other nutrients.21,27 In a previous study performed with L. alba cuttings of different diameters, the cuttings with 1-1.2 cm showed superior rooting when compared to those with smaller diameter. According to the authors, this happened due to the greater availability of reserves in thicker cuttings.2 Duarte et al.16 also observed better performance of thick and medium cuttings in relation to the thin ones for the rooting of this species.

The availability of reserves according to the different diameters may also explain the lower sprouting percentages in thin semi-hardwood cuttings and the superiority in the sprouts dry mass of thick semi-hardwood cuttings. The emission of sprouts is an important characteristic for good quality seedlings formation, once, after the depletion of cuttings reserves, the presence of new leaves is fundamental for plant nutrition.28

Marchese et al.2 also verified higher vigor of sprouts in L. alba cuttings with larger diameter. In a study that evaluated different lengths of cuttings, the higher content of reserves in longer propagules was also a fundamental characteristic for the highest percentage and dry mass of shoots in the species.12 In the species Lippia gracilis Schauer29 and Lippia sidoides Cham.30 direct relationships between the cuttings carbohydrates availability and sprouts emission and vigor were also observed.

The stem cuttings of Lipia alba linalool chemotype presented high endogenous rooting potential. However, the treatment of the cuttings with IBA at 2 000 mgx l-1 is able to promote increases in the rooting percentage. The isolated hydroalcoholic solution does not affect rooting in this chemotype.

Herbaceous apical and thin, medium and thick semi-hardwood cuttings can be used for the propagation of the L. alba linalool chemotype, with percentages above 85 % of rooting. Apical and thick semi-hardwood cuttings showed greater vigor of the root system.


Acknowledgements

The authors would like to acknowledge the Brazilian Federal Agency for Support and Evaluation of Graduate Education - CAPES by sponsorship of the research and scholarships granted.


Conflicto de intereses

Los autores plantean que no tienen conflicto de intereses.

 

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Recibido: 28/08/2017
Aprobado: 05/06/2018

 

 

Erik Nunes Gomes. Universidade Federal do Paraná, Brasil. Correo electrónico: e93gomes@gmail.com



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