EFFECTS OF GROWTH HORMONES TO THE ROOT DEVELOPMENT OF

(Mussaenda philippica A. Rich) THROUGH CUTTINGS UNDER VARIOUS

LEVEL OF CONCENTRATION

JHON MAR E. GATAO

A THESIS PROPOSAL SUBMITTED TO THE FACULTY OF THE COLLEGE OF

FORESTRY AND ENVIRONMENTAL SCIENCE CENTRAL MINDANAO
UNIVERSITY IN PARTIAL FULFILMENT OF THE REQUIREMENT FOR THE
DEGREE OF

BACHELOR OF SCIENCE IN FORESTRY

MAY 14, 2020

 CHAPTER 1

INTRODUCTION

BACKGROUND OF THE STUDY

The propagation of plants has been a fundamental occupation of mankind since

cultivation began hence, the essence of propagation is to increase the number of plants
and also to preserve its essential characteristics and the ornamental horticulture industry,
therefore, comprises all process relating to growth, production or use of plant materials
for aesthetic or functional value. Vegetative propagation proposes the prospects,
changes,
and opportunities to generate a dependable and sufficient supply of better-quality
planting stock locally, timely, and quickly. Various tropical tree species have been
propagated by cuttings (Tchoundjeu et al. 2002). In the vegetative propagation method
of rooting cuttings, very little work has been carried out on the ability of the tree species
to propagate vegetatively, but studies on tropical trees elsewhere indicate that many can
be propagated using this method (Henrique et al., 2006). The promotion of the
cultivation and conservation of Mussaenda philippica can easily be carried out if proper
vegetative propagation methods and growth media have been developed.

The plant Mussaenda philippica commonly known as "Queen of Philippines" is a

deciduous shrub widespread all over the world for its aesthetic nature for beautification,
fences, air purification, etc. the demand for its use as landscape plants and cut flowers is
undoubtedly on the increase. Adeduntan SA (2013). This plant cultivated mainly for its
long-lasting noticeable and colorful flag sepals form an important focal or spot plant in
the gardens whenever they are used singularly or in a group. They are used to create
an accent in the entrance, approach to a strategic garden structure or lawn background
as hedge and spot plants. Their beautiful colorful flowering twigs also make good cut
flower materials for vase or wreath making Ogbu JU, Ubani SI, Anaele MU (2008).
Mussaenda philippica is also used in traditional medicine for curing dysentery and
snake bites, a decoction of roots and leaves used for affections of the chest and lung
despite the
numerous economic uses of Mussaenda philippica, the population of the plant is
declining since it possesses a difficult to root morphological characteristic; only a small
percentage of the stem cutting propagated develops root Ogbu J (2011). Although the
plant can be traditionally propagated through stem cutting without any manipulation
which yields a very low percentage, this method can be improved when rooting
hormones are used to hasten root initiation. Rooting hormone hastens root initiation,
increase the number and percentage of cutting rooted as well as the quality of root
produced by cutting Leonardi C, Ruggeri A, Malta S (2001). But the technicalities
involved in the proper application, availability, and the impact it has on the environment
alongside the fact that they are expensive and not readily available to peasant
gardeners necessitate the need to find alternative means of stimulating stem cuttings to
increase the rooting percentage for better production. Many horticulturists prefer a plant
that is easily rooted after propagation, hence the difficulty in rooting is seen as an
undesirable trait Henry A, Olukemi Y (2013). This makes plant propagation one of the
most lucrative business opportunities among small and large scale ornamental growers.
To support the rapid multiplication of planting materials, propagation of M. philippica by
cuttings using rooting hormone with different level concentration can be a viable option.

Growing media is another important factor in propagation studies because rooting

performance depends on the type of medium used Ingram DL, Henley RW, Yeager TH
(1993). In the production of nursery plants, the selection and preparation of the medium
are extremely important in terms of plant growth and quality. A good rooting media used
alongside a rooting hormone will therefore hasten root initiation. Hence, the experiment
aimed at evaluating the root formation of Mussaenda philipica in response to the type of
hormone used and growth media that will enhance the successful establishment of the
plant in the nursery
STATEMENT OF THE PROBLEM

Tree ornamental species in our country is in a shortage of seedling because of

irregular flowering, seeds do not germinate, and the number of seedlings in the nursery
will decrease sharply in the year between mast flowerings. To overcome the problem,
the propagation protocol of Mussaenda philippica through cuttings is deemed important.
Stem cuttings with the use of growth media and rooting hormone will be utilized under
various levels of concentration as well as the rooting performance will be evaluated.
Specifically, this study will answer the following questions:

1. What are the appropriate rooting hormone and growth media suitable for Mussaenda
philippica cuttings in terms of various parameters?

2. Which levels of concentration on rooting hormone suitable for Mussaenda philippica

cuttings?

3. What are the treatment combinations showing the highest rooting performance among
the interaction of rooting hormones (IBA and NAA) and concentration levels (0, 500,
1000, 1500 ppm) to the cuttings of M.philippica?

OBJECTIVES OF THE STUDY

This study aims to evaluate the effect of rooting performance of Mussaenda philippica
through cuttings treated with different levels of concentration with Indole-3-Butyric Acid
(IBA) and Naphthalene Acetic Acid (NAA) rooting hormones.

Generally this study aims:

1. To determine the appropriate rooting hormone suitable for Mussaenda philippica

cuttings in terms of root length, number of roots, and rooting percentage;

2. To identify the levels of concentration on rooting hormone suitable for Mussaenda

philippica cuttings in terms of root length, number of roots, and rooting percentage; and
3. To determine the treatment combinations showing the highest rooting performance
among the interactions of rooting hormones (IBA and NAA) and concentration levels (0,
1000, 2000, 3000 ppm) to the cuttings of M. philippica

SIGNIFICANCE OF THE STUDY

This study will focus on the rooting performance of Mussaenda philippica through
cuttings treated with two rooting hormones under the various level of concentrations.
The study will provide information on the proper technique in the vegetative propagation
of this species. Specifically, the result of this study helps the manager of the nursery to
produce quality planting material of Mussaenda philippica species. Moreover, the data
generated from this study will serve as a future reference for students and researchers
in terms of utilizing rooting hormones for the rooting performance of Mussaenda
philippica species. Farmers and other farming enthusiasts can use the result of this
study as a guide for improving their technique in vegetative propagation for their farming
activities.

SCOPE AND LIMITATION OF THE STUDY

The study will focus only on one species; Mussaenda philippica found Mt. Kalayo
Musuan Peak. To evaluate the rooting performance of M. philippica treated only with two
rooting hormones; namely the Indole-3-butyric Acid (IBA) and Naphthalene Acetic Acid
(NAA) and only four levels of concentration (0, 500, 1000, 1500 ppm) and the untreated
will served as the control of the study. The duration of this study will start in January
2021 to March 2021 to terminate the specimen. The parameters used in the study are
limited only to the measurement of the root length, the number of roots, and rooting
percentage.
CHAPTER II

REVIEW OF RELATED LITERATURE

Description of Mussaenda philippica

Mussaenda philippica is a large shrub or small tree that is native to the Philippines and
frequently cultivated as an ornamental in tropical regions. It can be found growing in
parks, botanical gardens, private and public gardens, and along roadsides and
highways. Although plants in cultivation usually do not produce fruits, they can spread
vegetative by stem fragments and cuttings. M. philippica is a species of environmental
concern because it can grow up to 8 m tall and has a wide-spreading crown that may
shade out other vegetation including native plant species. Within its native distribution
range in the Philippines, M. philippica flowers throughout the year and fruits from
January to February, April to June, and August to December (Alejandro et al., 2016).
When growing in cultivation, M. philippica produces flowers throughout the year, but
rarely produces fruits (Fosberg et al., 1993; Alejandro et al., 2016). In Puerto Rico, it
has been recorded flowering from April to November (Hume, 1951). In Nigeria, it
produces flowers almost throughout the year (Ibironke and Victor, 2016). Mussaenda
philippica grows best in humid habitats with full sunlight but can tolerate some partial
shade. It prefers well drained clay, loamy or sandy soils with pH in the range 5.0-6.0. It
has moderate tolerance to salt spray (Alejandro et al., 2016).

Propagation of Mussaenda philippica

Mussaendas is known for its low pollen fertility, poor fruit production, and limited seed
production (Rosario, 1998: Steentoft, 1988). Mussaenda species are mostly propagated
by stem cuttings taken from hardwoods, semi-hard woods, and sometimes softwoods
and rooted in a rooting media under optimum nursery conditions, however, some
species/cultivars of potential aesthetic value are difficult to propagate by stem cuttings.
(Chadha and Choudhury, 2007; Ogbu, 2011). Phytohormones, especially various
auxins such as Indole acetic acid (IAA), Indole butyric acid (IBA), Naphthalene acetic
acid (NAA), and 2,4- Dichloropheoxy acetic acid (2,4-D) have been reported to promote
rooting in cuttings of most of the plant species (Hartmann et al., 2007). Menon
et al., (2013) reported that NAA has great sprouting and rooting potential in stem
cuttings of Bougainvillea. It was found that naphthalene acetic acid has a great
influence on adventitious root development and associated physiological changes in
stem cutting of Hemarthria compressa (Yan et al., 2014). Recently, the use of
Mussaenda species has been increased in the garden landscape in Bangladesh.
However, challenges arise for bud sprouting and root formation in the stem cuttings of
Mussaenda. For quality root production concentrations of phytohormone, chemicals are
of utmost importance, otherwise, it will lead to an undesirable effect. Therefore, the
present study was undertaken to find out the effect of phytohormone naphthalene acetic
acid on sprouting and rooting of stem cuttings in Mussaenda species.

Vegetative propagation by cutting

Vegetative propagation proposes the prospects, changes, and opportunities to

generate a dependable and sufficient supply of better-quality planting stock locally,
timely, and quickly. Various tropical tree species have been propagated by cuttings
(Tchoundjeu et al. 2002). For the production of high-quality timber and faster tree
growth, it is essential to start a vegetative propagation program by selecting elite or
genotypes from which the shoot cutting is to be taken (Gehlot et al., 2014). Vegetative
propagation has the advantage that it captures non-additive gene effects, but it should
be noted that vegetative propagation only replicates existing genetic traits within the
newly formed plantlet and does not in itself to improve the genetic quality of the being
propagated (Leakey, 2014).

Vegetative propagation is considered ideal in the rapid multiplication of a species under

threat while trying to maintain certain desired characteristics (Hartmann et al. 2002;
Tchoundjeu et al. 2004). However, the rooting ability of tree species through stem
cuttings is markedly affected by several factors such as cutting position from the shoot,
age of the donor plant, and IBA treatment (Hartmann et al. 2002; Husen and Pal 2006).
Variations in rooting ability for cuttings taken from different positions from the shoot are
often observed (Al-Salem and Karam 2001; Agbo and Obi 2007). Different cutting
positions from the shoot influence the overall quality, rooting ability, and subsequent
growth habit of rooted stem cuttings (Tchoundjeu and Leakey 1996; Husen and Pal
2003). The effect of the age of donor plants on the rooting ability of stem cuttings has
been reported in
many trees (Stenvall et al. 2004; Bhardwaj and Mishra 2005; Husen and Pal 2007a).
Variation in rooting ability of stem cuttings taken from different ages of donor plants has
been described that could be due to decrease in the content on endogenous auxins or
decreased sensitivity of aging tissues to rooting promoters and/or accumulation of
inhibitory substances which inhibit rooting (Husen and Pal 2006). It is well-established
fact that growth regulators such as IBA play an important role in controlling the rooting
ability of stem cuttings (Tchoundjeu et al. 2002; Gateable and Pastor 2006; Husen and
Pal 2007a).

Rooting media use in cutting

An appropriate rooting medium generally has to have an optimal volume of gas-filled

pore space and oxygen diffusion rate adequate for the needs of respiration (Fonteno and
Nelson, 1990). According to Caron et al. (2000), media physical properties should not be
constrained to just measurements of air-filled porosity, water holding capacity, and bulk
density, but also gas exchange characteristics. The highest number of shoots and length
of shoots recorded from milled pine bark may also be due to easy translocation of water
and mineral nutrients to the above-ground parts of the cuttings, leading to their rapid
growth and multiplication. Compared to soil and sand, pine bark is loose in texture hence
allowing for more aeration and water flow. Aeration and water holding capacity of the
media are often negatively correlated and therefore a balance between these must be
achieved to ensure optimal rooting (Ofori et al., 1996). The significant-high rooting in
milled pine bark may therefore be attributed to better aeration and water drainage
because high aeration and porosity are responsible for promoting root development
(Olabunde and Fawusi, 2003; Puri and Thompson, 2003). Therefore the type of rooting
media used can have a major influence on the rooting capacity of cuttings. A study of
rooting performance in Vitellaria paradoxa by Yeboah and Amoah (2009) showed that
high aeration in rooting media is responsible for promoting metabolic activities and
enhancing root initiation. Stem cuttings propagated in sand produced the least
percentage and number of roots per rooted stem cutting. This result was interesting
given that high rooting percentages to have been obtained from Cordia alliodora stem
cuttings propagated in sand media (Leakey et al., 1990). The sand was also identified
as the best
rooting media for Gongronema latifilia stem cuttings by Agbo and Omaliko (2006).
However, there is variation in response to different rooting media that has been reported
for many tree species. For example, in Southwest Cameroon and Ghana, the highest
rooting percentages of Irvingia gabonensis (Shiembo et al., 1996), Milicia excelsa (Ofori
et al., 1996), and V. paradoxa (Yeboah and Amoah, 2009) have been reported for stem
cuttings propagated in sawdust. The differences in the rooting ability of various species
propagated in different rooting media could be explained by their xeromorphic or
hydromorphic status (Loach, 1992) and the effects of this status on the water relations of
the cuttings (Mensen et al., 1997).

The importance of rooting medium for the rooting of leafy stem cuttings is widely
recognized (Leakey et al., 1990; Tchoundjeu and Leakey, 2001). Results of the present
experiment showed that the rooting ability of P. johimbe leafy stem cuttings is not very
sensitive to different rooting media used, although the water-retentive sawdust was the
best medium. The lack of any pronounced effect of rooting medium variation on the final
rooting percentage is, however, unusual. In Prunus africana (Tchoundjeu et al., 2002),
Dacryodes edulis (Mialoundama et al., 2002), and Ricinodendron heudelotii (Shiembo et
al., 1997) high rooting percentages were recorded for cuttings in sawdust than in other
rooting media tested. In contrast, Vochysia hondurensis (Leakey et al., 1990) and
Cordia alliodora (Mese´n et al., 1997) are rooted better in gravel and sand.
Rooting hormone use in cutting

Auxin is a group of hormones that regulates plant growth, particularly by stimulating cell
elongation in the stem and inhibiting it in roots. The induction of root in the cuttings
depends on the presence of endogenous auxin inside the plant tissues and its
synergistic effect with exogenous ones leads to the synthesis of ribonucleic acid and as
a result induces root primordial (Hartman et al, 2002). The Indole acetic Acid (IAA) is
the naturally occurring auxin found in plants. Some of the processes regulated by IAA
include the formation of the embryo in development, induction of cell division, stem and
coleoptiles elongation, apical dominance, induction of rooting, vascular tissue
differentiation, fruit development, and tropic movement such as bending toward the light.
Auxin is well known to play a significant role in stimulating adventitious roots from the
cutting of tree species (Poupard et al., 1994; Tchoundjeu et al., 2004). It has been
confirmed that auxin is required for adventitious root formation on the stem and the
division of the first root initial is dependent on the exogenous and endogenous level of
auxins (Ludwig-Muller 2000; Kochhar et al., 2005). The synthetic form of auxin is the
indole-3-Butyric Acid (IBA) and Naphthalene Acetic Acid (NAA). Larsen and Guse (1997)
and Kester et al. (1990) reported that the most reliable rooting hormone is indolebutyric
acid (IBA) although others such as naphthalene acetic acid (NAA) can also be used.
Although there are reports that it may also be toxic to young/ succulent cuttings of
certain species, IBA is still probably the best hormone for general use because of being
non-toxic to plants over a wide range of concentration levels (Kester et al., 1990) Like
many other species, rooting ability of D. melanoxylon was sensitive to the IBA
application. Cuttings treated with IBA produced more roots as well as longer roots than
untreated cuttings. Similar results were reported by Ofori et al. (1996) in which the mean
number of roots per cutting was higher when the cuttings of Milicia excelsa were treated
with IBA than untreated cuttings. The stimulatory effects of IBA in rooting of stem
cuttings of several other woody plant species have been reported by other workers who
have revealed that IBA has an important role in the development of adventitious roots,
increasing rooting percentage, improving quality of roots and uniformity in rooting of the
cuttings (Negash 2002; Tchoundjeu et al. 2002; Teklehaimanot et al. 2004; Husen and
Pal 2007b; Opuni Frimpong et al. 2008). Root formation process in cuttings is intensified
by IBA which influences polysaccharide hydrolysis resulting in increased content of
physiologically active sugar needed to provide energy for meristematic tissues and later
for root primordial and roots formed as reported in Tectona grandis (Husen and Pal
2007a) and Dalbergia sissoo (Husen 2008).

Levels of concentration of rooting hormones

The effects of growth regulators (IAA, NAA, IBA at the rate of 2000, 4000, or 6000 mg/l)
on softwood cuttings of C. lanceolatus were investigated and the highest rooting
percentage was achieved by the application of 6000 mg/l IBA (Balakrishna and
Bhattacharjee, 1991). However, in a Persian citation, IBA at the rate of 4000 mg/l was
found to generate the highest rooting response in semi-hardwood cuttings of C. viminalis
(Zarinbal et al., 2005). Application of IBA at the rate of 6000 mg/l was found to decrease
the rooting response of bottlebrush cuttings. The rooting percentage, root length, and
production of new shoots also were significantly decreased in cuttings treats and with a
high concentration of IBA. The negative effect of high hormonal dose and its toxic effect
was already reported in the propagation of Phalsa (Grewia subinaequalis) through
cutting (Ajaykumar, 2007). However, in the case of Callistemon lanceolatus, which is
difficult to root species of bottlebrush, the highest rooting percentage (80%) was
attained in 6000 mg/l IBA (Balakrishna and Bhattacharjee, 1991). Auxin causes cell
enlargement and elongation of the root cells. The high concentrations of auxin may
harm root elongation (Hartman et al., 2002). The auxin application may increase the
number of roots per each individual cutting (Hussein, 2008; Nair et al, 2008). There are
some data on the enhancement of the respiratory rate and presence of high levels of
stored amino acids in the base of the treated cuttings, 24 hours after external
application of IBA (Shokri, 2012).
CHAPTER III

METHODOLOGY

DURATION AND LOCATION OF THE STUDY

The study will be conducted with a duration of three months starting from June to
August 2021 at the Clonal Forest Nursery of the College of Forestry and Environmental
Science, Central Mindanao University, Musuan, Maramag, Bukidnon. The geographical
coordinates of the site 125 2' 47.38E and 7 51'44.452N.
 Figu

re 1. Location of Study Area

EXPERIMENTAL DESIGN AND TREATMENT

The experiment will be laid out in a Randomized Complete Block Design (RCBD) with 2
x 4 x 10 factorial experiment structure with four (4) replications. There will be a total of
three hundred twenty (320) cuttings of Mussaenda philippica will be used. A total of
eight (8) treatment combinations are established and distributed randomly in each Block.
The treatment of the study will be as follows:

Factor A. Treatment of hormones

H1 – Indole-3-Butyric Acid (IBA)

H2 – Naphthalene Acetic Acid (NAA)

Factor B. Concentration level of (IBA) and (NAA)

R0 – control (0 ppm)

R1 – 500 ppm

R2 – 1000 ppm

R3 – 1500 ppm
Preparation of the Rooting Bed

The rooting bed must be cleaned, two weeks before the implementation of the study.
The study will be conducted at Clonal Forest Nursery in the College of Forestry and
Environmental Science. The type of soil medium in the rooting bed is river sand and
topsoil. The rooting bed will be sterilized using the drenching method and spraying
fungicide to avoid infestation of cuttings.

Collection and Preparation of the Cuttings

The cuttings of Mussaenda philippica will be collected from selected plus trees in Central
Mindanao University (CMU) Campus Musuan, Maramag, Bukidnon. The cuttings will be
collected in the morning hours and kept moist/ cool by placing them in cool boxes (Agbo
and Obi, 2007). It lessens the transpiration losses as well as the cutting stress. In
selecting shoot cuttings, the young/ succulent shoot are prepared and trees will be
chosen from an orthotropic oriented shoot using a pruning shear. According to Kester et
al. (1990),
the cuttings will be dipped in water up to a length of 1 cm to avoid water loss and hence
preventing wilting. The one or two-nodal cuttings with at least one mature leaf trimmed
into one-half from its original size without damaging or pulling on the bark will be used.
The cuttings will be sterilized by soaking into a fungicide solution for at least 15 minutes
to 1 hour to prevent fungus infection. Wounds will then be made by scrapping/ removing
a small portion of the base (0.5 to 0.1 cm) to make a full surface contact of the cutting to
prepare rooting hormone for a faster response of cuttings.

Preparation of Rooting Hormone

The study prepared two rooting hormones (Indole-3- Butyric Acid (IBA) and
Naphthalene Acetic Acid (NAA)). The different levels of concentration include 0, 500,
1000, 1500 ppm where 0 is the control. By making the hormone solution, 500 ppm
resulted by mixing 0.05 g powder hormone and one tablespoon 70% alcohol to dissolve
the hormone, then the hormone will be mixed in a 1000 mL of distilled water. 2000 ppm
has 0.20 g and 3000 ppm have 0. 30 g the same process is done to 300 ppm. Each level
of concentration hormone will be prepared in a separate container.
Treatment of Cuttings with Rooting Hormones

The M. philippica cuttings will be cut in a slant direction at the base below the nodal
which will be placed in a cool gallon filled with water. For each cutting, 1 inch of the
basal end will be dipped into the different level concentration except for the control
treatment.

Planting the Cuttings into the Rooting Bed

Right after soaking the treated cuttings, holes will be made into the rooting bed with a
distance of 1 to 1.2 inches from each other. The cuttings that were treated with a
different level concentration of IBA and NAA will be immediately planted according to the
design of the study.

Care and Maintenance

The watering schedule and sanitation will be observed to ensure that the cuttings are
free from fungus infection.

Gathering of Data

The data will be gathered about the expected number of days set in the study upon the
emergence of the root. Measuring the root percentage, root number, and root length.

Root number – will be obtained by counting the root that developed the base of the
cuttings. It is done by terminating the study.

Root length – will be obtained by measuring the primary root and the longest root in
centimeters with the ruler. It is done by terminating the study.

Rooting percentage – will be obtained by dividing the number of cuttings that root, the
total number of cutting plant, and multiplying the result by 100. It is done by terminating
the study.

DATA ANALYSIS

The data will be analyzed using the Analysis of Variance (ANOVA) through the
Statistical Tool for Agricultural Research (STAR) software. To determine the difference
whether it is significant to the treatment using the Duncan's Multiple Range Test.
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