Wound Healing Activity of Povidone-iodine with 10 Philippine Plants

Revilla, Ana Fe B.a, Guzman, Florenze Wendell F.b, Dizon, PatrizeC.b, Dominno, Donna Mae L.b, Elayda,
Grace Chelsea C.b, Fernandez, Kirstie Lynn E.b, Javier, JhazelNianneC.b,
Juan, Shylane Kate A.b, Mangawa, Marlon A.b
a
Faculty Research Promoter, School of Natural Sciences, Saint Louis University
b
Student Researchers, Pharmacy Department, School of Natural Sciences, Saint Louis University

ABSTRACT
Povidone-iodine (Betadine) is a popular drug in the country and has been providing the people
its optimum benefit throughout the years. With this, the study aimed to find out if the use of 10
Philippine plant extracts containing antibacterial property will help in the innovation of the
products efficacy and effectivity towards wound healing activity. The study intends to
determine which of the plants will have a synergistic effect with the PVP-I and will be ranked
according to the results in the observation of the change in length of wound and scab formation
on the test subjects. Thirty-six adult male Sprague-dawley rats (190 10g) with an open wound
of 20 10mm on the dorsal area were divided into three major groups: experimental group for
the PVP-I with the 10 plant extracts, positive control group for the PVP-I alone and negative
control group. Complete wound healing (full epithelialization) and scab formation was measured
using superficial observation of the wound area daily. After 4 weeks, all rats were not euthanized
rather they were sent back to the Department of Veterinary Medicine, Benguet State University
to be donated to other research groups. The results of the study showed that the combined
antiseptic activity of PVP-I with antibacterial activity of the 10 Philippine plants improved the
wound healing process. The guava extract showed the greatest result both in the superficial
wound healing observation and area of zone of inhibition compared to the others.

Key Words: Wound healing, Povidone-Iodine (PVP-I), Zone of inhibition, Antibacterial

Introduction
Wound healing is an innate response of the body to repair or restore injured tissues. The
presence of wounds makes the skin susceptible to microbial infections because broken skin
serves as a route of entry of pathogens. It is important to maintain the integrity of the skin since it
is part of the first line of defense of the body against its environment. The wound healing
process is divided into four phases: hemostasis, inflammation, granulation/proliferation,
remodeling/maturation.

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 Controversies have emerged from different studies regarding the use of povidone-iodine
(PVP-I) in wound healing. Most of the studies showed either an increase, reduction or no effect
on the healing rate. There is no definite evidence that suggests a positive or a negative effect on
wound healing as shown from the conflicting results between animal and human studies. The
studies only proved that 10% PVP-I can act as a bactericidal which will improve healing rate
because of the decrease in incidence of having a risk of infection. It did not prove on whether
PVP-I has a direct role in the process of wound healing (Drosou, Falabella&Kirsner, 2003).

Previous clinical studies have shown that povidone-iodine is a safe and effective
antiseptic for wound healing in the health care setting. Povidone-iodine (PVP-I) is an iodophor, a
complex of polyvinylpyrrolidine and elemental iodine. Povidone-iodine preparations were
introduced in the 1960s and it is now the most common iodophor in clinical use. It is available in
different formulations, including solution, cream, wound dressings, ointment, and spray (Sibbald,
2011).

PVP-I is popularly known in its commercial preparation as Betadine. Its active

ingredient, iodine, is a halogen that works as a strong oxidizing agent. Since many enzymes
contain sulfhydryl group (-SH) in their active site, it is susceptible to oxidation. The process
alters the sulfhydryl group into a disulfide bridge (-S-S). The chemical changes causes
inactivation of the pathogen. As the enzyme is oxidized, the iodine is reduced to iodide ion and
then removed. More PVP-I is converted to free iodine in order to maintain the equilibrium
(Shikashio, 1969; Simonsen, 1933; Stuart, 2009).

Studies on wound healing activity of povidone-iodine combined with other plant extracts
have not been conducted yet. Some studies only compared the antiseptic property of PVP-I
against other plants with antiseptic and antibacterial properties. The studies only showed the
degree of wound healing activity when compared with other plants. PVP-I is a well-known
product worldwide that is why the researchers wanted to do experiments and researched in order
to improve the products effect, to know if PVP-I with the plant extracts would cause a shorter
duration of wound healing as compared to PVP-I alone.

The 10 Philippine plants that were utilized in the study are: Dama de noche (Cestrum
nocturnum, Solanacea), Guava (Psidiumguajava, Myrtaceae), Garlic (Allium sativum, Liliaceae),
Tsaang-gubat (Carmona retusa, Boraginaceae), Sampa-sampalukan (Phyllanthusniruri,
Euphorbiaceae), Ginger (Zingiberofficinale, Zingiberaceae), Oregano (Origanum vulgare,
Lamiaceae), Kutsay (Allium tuberosum, Amaryllidacae), Lantana (Lantana camara,
Verbenaceae) and Lemon grass (Cymbopogoncitratus, Poaceae) because this plants are the most
common with antibacterial property which is also available within our vicinity.

Kirby-Bauer test was also used to validate the antibacterial properties of the plant
extracts. Many studies have shown that flavonoids and alkaloids are constituents mostly found in
plant extracts. These plant metabolites are one of the primary constituents which are responsible
for the plants antibacterial property since studies showed that the presence of these constituents
contributed to the reduction of microorganisms as shown by the zone of inhibition.

2
 The 10 plants that will be used in the research were studied by previous researchers and
they have found out the presence of certain different flavonoids and alkaloids unique to the
plants which showed antibacterial properties.

The study will determine whether each of the 10 plants that will be added to PVP-I can
have a synergistic effect when used together. Synergism has been defined as a phenomenon in
which two different compounds are combined to enhance their individual activity (Rani,
Dureja&Kumar, 2009) Also, the researchers would like to determine whether each of the plants
can contribute to wound healing by shortening the wound healing process.

Materials and Methods

Freshly harvested plants were weighed 50g each. And washed, dried, and cut into small
pieces. Each sample was soaked in 250ml of 95% ethanol for three days, filtered, evaporated and
concentrated using the water bath.

Plant extract discs were prepared using Whatman filter paper that was prepared using a
puncher. The discs were placed in a small petri dish covered with aluminum foil and autoclaved
at 121C for 15 minutes to blast the bugs and germs on things with steam long enough
to sterilize them. The extra pressure in an autoclave means that water boils at a temperature
higher than its normal boiling pointroughly 20C hotterso it holds and carries more heat and
kills microbes more effectively. A lengthy blast of high-pressure steam is much more effective at
penetrating and sterilizing things than a quick wipe in ordinary hot water (Woodford,
2016).Approximately, twelve discs were soaked in each plant extract for three days this is to
ensure that the antimicrobial constituent of the plant will be absorbed by the discs.

The antibacterial activity of the plant extracts were tested against four bacterial species:
Staphylococcus aureus, Bacillus cereus, Escherichia coli and Pseudomonas aeruginosa. Disc
diffusion method was used for assessment of the antibacterial activity through the use of
measurement of the zones of inhibition.

The plant extract discs were placed in the cultured media and positioned far from each
other. Three discs were assigned to a plant extract per specimen. The cultured media was
incubated at 37C for 24 hours. After incubation, the zones of inhibition of each plate were
observed and measured control and expressed in millimetres. PVP-I solution was used as
positive control.

A ratio of 1:1 is used in the preparation of topical solution for the application on test
animals. Ten ml of the plant extracts was mixed with 10ml of 10 % PVP-I (Betadine)
purchased commercially. Each topical solution was stored in an amber-colored bottle.

Thirty-six (36) healthy male Sprague -Dawley albino rats with an average weight of 180
to 200g were used in the study. All were fed with dog food diet (10g/100g/day), since it contains
a higher amount of protein. Feeding was done at the same time every day, which is 8AM. Water
was given ad libitum. Three rats were housed in one individual plastic cages and was assigned
for one plant sample. Food item was provided in food hoppers. A separate drinking bottle was
3
provided with purified water. Sterile sawdust was used as beddings and changed with clean ones
at an interval of 3 days. The animals were housed under normal conditions of light, room
temperature and humidity.

The rats were randomly divided into three major groups: the experimental group treated
with the plant extracts with PVP-I, the control group treated with PVP-I, and the negative control
group, which was left untreated. The first group was further subdivided into ten groups to
represent the ten plants. Each group consisted of three rats which were treated with the
designated plant extract with PVP-I treatment. The results of the three groups were compared
with each other.
The rats were anesthetized with 20-40 mg/kg of Zolazepam/Tiletamine (Zoletil), a
general anesthetic, before incision. The drug was administered via the intraperitoneal route. The
hair on the dorsal area was removed with surgical scissor. An open wound of approximately 10
to 30 mm long up to the level of subcutaneous adipose tissue with the use of a surgical blade was
made. The operation was performed with the assistance of a registered veterinarian under clean
conditions. After the incision, the wounds of each rat was treated with an appropriate amount of
the topical formulation on the wound area. Topical application was repeated and continued until
the wound is completely healed. Application of treatment was done once a day at a constant time
(8:00 PM). The wound area was measured using a ruler every day until healing is completed.
The measurements were recorded in millimeters.
After 4 weeks, all rats were not euthanized but were sent back to the Department of
Veterinary Medicine, Benguet State University to be donated to other research groups.

Results and Discussion

Table 1a. Mean Changes in Wound Length on Rats Treated with Povidone-Iodine
And Plant Extracts at 3-day Intervals

TREATMENT Mean Changes in Wound Length

After Day 3 After Day 6 After Day 9 After Day 12
Guava -6.67 -13.67 0.00 -1.33
Ginger -3.33 -3.33 -3.67 -7.00
Lantana -4.23 -5.93 -1.67 -1.69
Garlic -4.67 -2.33 -3.67 -2.33
Lemon Grass -2.67 -1.67 -1.67 -5.00
Sampa-sampalukan -1.00 -6.00 -4.67 -4.00
Kutsay -2.67 -3.00 -3.00 -1.67
Oregano -1.83 -3.83 -2.00 -4.00
TsaangGubat -1.67 -1.00 -5.00 -1.67
Dama de Noche -3.67 0.00 -1.67 1.67
*Povidone Iodine -0.67 -4.33 -4.67 -2.67
*Positive control

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 Table 1a shows the change in length of the wound of rats when treated with PVP-I and
the plant extracts. It shows the difference in the wound size taken every 3 days.

Based on the table, after day 3 guava showed the greatest difference in wound length
followed by garlic and lantana. The least difference was shown by sampa-sampalukan.PVP-I
alone ranked last since it had the lowest difference in wound length. After day 6, guava still
showed the greatest difference in wound length followed by sampa-sampalukan and lantana. The
least difference was shown by tsaang gubat, the rank of PVP-I alone was next to lantana in terms
of difference in wound length. After day 9, tsaang-gubat showed the greatest difference in wound
length followed by sampa-sampalukan, ginger and garlic. The least difference was shown by
guava, PVP-I alone exhibited the same difference in wound length with sampa-sampalukan.
After day 12, ginger showed the greatest difference in wound length followed by lemon grass
and sampa-sampalukan with oregano. The least difference was shown by kutsay, dama de noche
and tsaang-gubat. PVP-I alone ranked next to sampa-sampalukan and oregano in terms of
difference in wound length.

To test for the significance of the wound healing activities of the povidone-iodine with
plants during the different time intervals, ANOVA was used.

In traditional medicine, leaves of guava have been reported to promote the healing of
open wounds when dressed with it. This prohealing activity of leaves on open wounds could be
attributed to their antimicrobial activity (Mudliar, Patil&Torgal et. al. 2008).Comparative study
of hydroalcoholic and methanolic extract of Psidium guajava leaves was done to investigate
wound healing activity. The results showed that the hydroalcoholic extract was more effective in
comparison with the methanolic extract. From the results, it was concluded that Psidium guajava
leaves extract when formulated in gel showed significant improvement in wound contraction and
hence this is a promising candidate for wound healing activity (Kasar, N.S. 2016).Comparative
evaluation of wound healing effects of Ocimum gratissimum, Vernonia amygdaline, and
Zingiber officinalis extracts were investigated using incisional wound model in rats. Results
showed that animals treated with V. amygdaline have greater wound healing than O. gratissimum
and Z. officinalis. However, it showed no significant difference (p<0.05) when analyzed
statistically (Eyo, J.E., et al 2014).According to a study, aqueous extracts of garlic bulbs in
combination with honey significantly accelerated wound healing compared with honey alone.
This means that garlic has beneficial effects for accelerating the wound healing process (Sidik,
Mahmod&Salmah 2006). Lemon grass accelerated wound closure on diabetic and non-diabetic
rats which indicates that it can improve wound healing process (Umasankar, Balwin& Manley
Backyavathy, 2013).Ethanolic extract of dama de noche contains the phytoconstituents that
promote natural healing process and it could be effectively used as a wound healing agent. It
efficiently stimulates the wound strength and increases the rate of epithelialization, tensile
strength, and collagen viability around the wound area (Hemant, Amit &Rajnish et. al., 2016).
Sampa-sampalukan could significantly enhance the rate of wound healing. Its extracts possess a
broad spectrum of biological activities. It plays a significant role in the wound healing process
5
and protect tissues from oxidative damage. Wound healing mechanisms may be contributed to
stimulate the production of antioxidants in wound site and provides a favorable environment for
tissue healing and wound healing effects may be due to up-regulation of human collagen I
expression and an increase in tensile strength of the wounds (Khaled, Mahmood, &Fahmi, 2012).
Lantana camara leaf extract has shown to contain flavonoids (Pan et al., 1993) and terpenes
(Begum et al., 2003), which may mediate the wound healing process. L. camara prevented lipid
peroxidation that may suggest that it may be acting as antioxidant (Saini et al., 2007).
Antioxidants have been reported to play a significant role in the wound healing process. It
appears that antioxidant may be an important contributory factor in the wound healing property
(Shukla et al., 1999). Studies have shown the topical application of compounds with antioxidant
properties on patients or animals to significantly improve wound healing and protect tissues from
oxidative damage (Martin, 1996). L. camara leaf possesses antitumor, anti-viral, antibacterial
and anti-inflammatory activities and thus the wound healing activity could be attributed to these
properties (Xu et al., 1996). Wound healing properties of L. camara could also be due to
antimicrobial activity (Deena and Thoppil, 2000). L. camara extract exhibited significant
antimicrobial activity and properties that support folkloric use in the treatment of some diseases
as broad spectrum antimicrobial agents. This probably explains the use of these plants by the
indigenous people against a number of infections since earlier generation (Kumar et al., 2006).
Origanum vulgare helped in treating postsurgical wounds and had equivalent scar appearance
compared to petrolatum, this is because of its effect in decreasing bacterial contamination and
subsequent infection. In Chinese medicine, Allium tuberosum leaves and bulbs are used in bites,
cuts and wounds. It helped in fast healing of bites, cuts and wounds obtained from insect,
animals and other sources.

Table 1b. Test of significance of the wound healing activity of Povidone-Iodine enhanced
by the plant extracts in different time interval using ANOVA.

Time Interval Test Statistics F-value p-value

After day 3 3.539 0.006*
After day 6 9.626 0.000*
After day 9 1.526 0.196
After day 12 2.053 0.077
*significant @ p-value < 0.05

The table shows the test of significance of the wound healing activity of Povidone-iodine
enhanced by the plant extracts at different time interval using F-test or ANOVA. Based from the
results, it shows that after 3 and 6 days of observation, the f-value of 3.539 and 9.626 have p-
values of 0.006 and 0.000respectively, indicating that the difference in wound healing of the
plant extract enhanced by the plant extracts is significant. After 9 and 12 days of observation the
f-value of 1.526 and 2.053 have p-values of 0.196 and 0.077 respectively, indicating that the

6
difference in wound healing enhanced by the plant extract is not significant. This means that
when povidone-iodine and the plant extracts were used to treat the wounds of the rats, there was
a noticeable shortening of the wounds when compared with each other and when the treatment
was compared to povidone-iodine alone.

To determine where the significant difference actually transpired, post hoc analysis was
conducted using Tukey procedure.

Table 1c. Post hoc analysis after 3 days of observation using Tukey Procedure

Subset for alpha = 0.05

Extract N 1 2
Guava 3 -6.67
Garlic 3 -4.67 -4.67
Lantana 3 -4.23 -4.23
Dama de noche 3 -3.67 -3.67
Ginger 3 -3.33 -3.33
Lemon grass 3 -2.67 -2.67
Kutsay 3 -2.67 -2.67
Oregano 3 -1.83
Tsaang gubat 3 -1.67
sampa-sampalukan 3 -1.00

Table 1c illustrates the post hoc analysis on the wound healing activity of the different
extract after 3 days. The table consists of 2 clusters. For the first cluster, guava, garlic, lantana,
dama de noche, ginger, lemon grass and kutsay have the same effectivity, hence, are not
significantly different with each other. This means that the wound healing activity of these plants
when used together with povidone-iodine have similar action in shortening the length of the
wounds. For the second cluster, garlic, lantana, dama de noche, ginger, lemon grass, kutsay,
oregano, tsaang gubat and sampa-sampalukan exhibited the same effectivity and are not
significantly different with each other. However, comparatively guava is significantly different
with oregano, tsaang gubat and sampa- sampalukan.

7
 Table 1d. Post hoc analysis after 6 days of observation using Tukey Procedure

Subset for alpha = 0.05

Extract N 1 2
guava 3 -13.67
sampa-sampalukan 3 -6.00
lantana 3 -5.93
oregano 3 -3.83
ginger 3 -3.33
kutsay 3 -3.00
garlic 3 -2.33
lemon grass 3 -1.67
tsaang gubat 3 -1.00
dama de noche 3 0.00

Table 1d shows the post hoc analysis on the wound healing activity after 6 days of
observation. On the first cluster, only guava showed a significant difference with the rest of the
plants. On the second cluster, sampa-sampalukan, lantana, oregano, ginger, kutsay, garlic, lemon
grass, tsaang-gubat and dama de noche have the same effectivity and are not significantly
different with each other.

The day where the most significant change occurred was after 6 days of observation. This
shows that the earlier phase of the wound healing process have the better results of wound
reduction compared to the later phase.

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Table 2. Comparative Analysis of the wound healing activity of Povidone-Iodine enhanced
by the plant extracts as shown by the mean zone of inhibition on various test organisms

TREATMENT Mean Zone of Inhibition (in mm)

Staph aureus Bacillus cereus E. coli P. Aeruginosa
Guava 13.00 10.67 14.67 18.00
Ginger 8.00 10.00 0.00 0.00
Lantana 6.00 8.00 0.00 0.00
Garlic 12.00 9.33 0.00 0.00
Lemon Grass 6.00 9.33 8.00 0.00
Sampa-sampalukan 14.67 8.00 14.00 14.00
Kutsay 10.00 10.00 0.00 8.00
Oregano 7.67 0.00 0.00 8.67
Tsaang Gubat 9.33 8.00 0.00 2.67
Dama de Noche 8.67 9.33 0.00 8.00
Povidone Iodine 8.67 8.33 0.00 0.00

Table 2 shows the mean zone of inhibition on various test microorganisms. Based on the
table under the observation on Staphylococcus aureus, sampa-sampalukan showed the greatest
zone of inhibition followed by guava and garlic. The plant with the least zone of inhibition was
shown by lemon grass and lantana. For Bacillus cereus, guava showed the greatest zone of
inhibition followed by kutsay, ginger, garlic lemon grass and dama de noche. The plant with the
least zone of inhibition was shown by lantana, sampa-sampalukan and tsaang-gubat while the
plant without a zone of inhibition was oregano. For Escherichia coli, guava showed the greatest
zone of inhibition followed by sampa-sampalukan and lemon grass while the plants without a
zone of inhibition were ginger, lantana, garlic, kutsay, oregano, tsaang gubat, dama de noche.
For Pseudomonas aeruginosa, guava showed the greatest zone of inhibition followed by sampa-
sampalukan and oregano. The plant with the least zone of inhibition was tsaang-gubat while the
plants without a zone of inhibition were ginger, lantana, garlic and lemon grass.

Under the observation on Staphylococcus aureus and Bacillus aureus, PVP-I showed an
average zone of inhibition. It did not show any zone of inhibition on E. coli and P. aeruginosa.

The active constituent that are responsible for the antibacterial properties of these plants
are: For Dama De Noche are alcohols like phenylethyl alcohol, benzyl alcohol, 1-nonadecanol
phenols like eugenol, fatty acids like hexadecanoic acid, esters like methyl monoterpenoid like
anthranilate, hydrocarbons like nonadecene, aldehyde and ketones like tetradecanal and
citronellal (Reza, Rahman & Kang, 2009). Garlic commonly found to have antimicrobial
properties contains indole glycoside like Allicin (Cutler & Wilson,2004). Chemicals isolated
from guava leaves like quercetin, guaijaverin, several flavonoids and galactose-specific lecithins
have shown promising activity in many human clinical trials (Abdelrahim et al., 2002, Adeyemi

9
et al. 2009). Flavonoids extracted from guava leaves including quercetin and were reported to
have strong antibacterial action (Rattanachaikunsopon and Phumkhachorn, 2007; Arima and
Danno, 2002). Kutsay contains an Organophosphate derivative, Diallyl sulfide (Mnayer,
Petitcolas&Hamieh, et. al. (2014). Sampa-sampalukan contains alkaloids, lignans, phyllanthin,
hypophylanthin, flavonoids, triterpenoids, glycosides, and tannins (Valle et. al. 2015). Ginger
contains a monocyclic sequiterpene hydrocarbons like -sesquiphellandrene, caryophyllene and
zingiberene in ginger rhizome oil (El-Baroty, Abd El-Baky,Farag& Saleh, 2010). Lantana
contains lantadenes, phenolics, Glycosides like anthocyanidins, proanthocyanidins (Agrawal,
Varma & Goyal, 2012). Lemon grass contains terpenoids like citral (geranial), citral
(neral) (Shah, Shri &Panchal,et. al. 2011). Mainly the essential oil of oregano contains it
antibacterial property (Sienkiewicz , Wasiela&Gowacka, 2012) For Tsaang gubat studies
suggest that triterpene mixture, -amyrin, -amyrin and baurenol (Chandrappa,
Govindappa&Anil Kumar2012).

Table 2a.Test of significance of the zone of inhibition against various test microorganisms
using ANOVA

Test Organisms Test Statistics F-value p-value

Staph aureus 6.567 0.00*
Bacillus cereus 20.220 0.00*
E. coli 218.35 0.00*
P. aeruginosa 44.296 0.00*
*significant @ p-value < 0.05

Table 2a shows the significance of the zone of inhibition against test microorganisms. For
Staph. aureus, Bacillus cereus, E. coli and P. aeruginosa have f-values of 6.567, 20.220, 218.35
and 44.296 respectively. All the test microorganisms gave a p-value of 0.00 indicating a
significant difference on the zone of inhibition. This means that the combination of povidone-
iodine and the plants are effective against the four microorganisms.

10
 Table 2b.Post hoc analysis on zone of inhibition using Tukey Procedure

On Staphylococcus aureus

Subset for alpha = 0.05

Extract N 1 2 3
lantana 3 6.00
lemon grass 3 6.00
oregano 3 7.67 7.67
ginger 3 8.00 8.00
dama de noche 3 8.67 8.67
Povidone Iodine 3 8.67 8.67
tsaang gubat 3 9.33 9.33 9.33
kutsay 3 10.00 10.00 10.00
garlic 3 12.00 12.00
guava 3 13.00 13.00
sampa-sampalukan 3 14.67

The table illustrates the post hoc analysis on the zone of inhibition of Staph. aureus.
Sampa-sampalukan is significantly different with povidone-iodine, dama de noche, ginger,
oregano, lemon grass and lantana. Garlic and guava is also significantly different with lantana
and lemon grass. For the zone of inhibition on Staph. aureus, sampa-sampalukan gave the best
result and is the most significantly different with most of the plants.

11
 Table 2c.Post hoc analysis on zone of inhibition using Tukey Procedure

On Bacillus cereus

Subset for alpha = 0.05

Extract N 1 2
oregano 3 0.00
lantana 3 8.00
sampa-sampalukan 3 8.00
tsaang gubat 3 8.00
Povidone Iodine 3 8.33
garlic 3 9.33
lemon grass 3 9.33
dama de noche 3 9.33
ginger 3 10.00
kutsay 3 10.00
guava 3 10.67

The table displays the post hoc analysis on zone of inhibition on Bacillus cereus.
Oregano showed no antibacterial activity towards Bacillus cereus and is significantly different
with the rest of the plants. The rest of the plants showed the same effectivity with Bacillus
aureus but is not significantly different with each other. However, statistically, guava yielded the
highest results of antibacterial activity with Bacillus cereus.

Table 2d.Post hoc analysis on zone of inhibition using Tukey Procedure on Escherichia coli

Extract N Subset for alpha = 0.05

1 2 3
ginger 3 0.00
lantana 3 0.00
garlic 3 0.00
kutsay 3 0.00
oregano 3 0.00
tsaang gubat 3 0.00
dama de noche 3 0.00
Povidone Iodine 3 0.00
lemon grass 3 8.00
sampa-sampalukan 3 14.00
guava 3 14.67

12
 Table 2d shows the post hoc analysis on zone of inhibition on Escherichia coli. Ginger,
lantana, garlic, kutsay, oregano, tsaang-gubat, dama de noche and povidone-iodine showed no
antibacterial activity on E. coli. Guava and sampa-sampalukan have the same effectivity and are
significantly different with lemon grass and the rest of the plants. However, guava showed the
largest result of zone of inhibition towards E. coli.

Table 2e.Post hoc analysis on zone of inhibition using Tukey Procedure on Pseudomonas
aeruginosa

Extract N Subset for alpha = 0.05

1 2 3 4
ginger 3 0.00
garlic 3 0.00
lemon grass 3 0.00
Povidone Iodine 3 0.00
tsaang gubat 3 2.67
kutsay 3 8.00
dama de noche 3 8.00
oregano 3 8.67
lantana 3 12.00 12.00
sampa-sampalukan 3 14.00 14.00
guava 3 18.00

The table illustrates the post hoc analysis on zone of inhibition on Pseudomonas
aeruginosa. Guava is significantly different with all of the plants except sampa-sampalukan.
Sampa-sampalukan is significantly different with oregano, dama de noche, kutsay, tsaang gubat,
lemon grass, garlic and ginger. Ginger, garlic, lemon grass, povidone-iodine and tsaang gubat are
not significantly different with each other however, the statistical results for tsaang gubat showed
a minimal zone of inhibition compared to the others.

To summarize, guava and sampa-sampalukan gave a consistent results on its antibacterial

activity with the four test microorganisms. Considering the zone of inhibition, Sampa-
sampalukan gave the best antibacterial activity with Staph. aureus while guava was the most
effective against Bacillus cereus, E. coli and P. aeruginosa.

13
 Table 3. Day of Scab Formation

Extract DAY OF SCAB FORMATION

1.Guava DAY 3
2.Ginger DAY 3
3.Lantana DAY 9
4.Garlic DAY 3
5.Lemon Grass DAY 9
6.Sampa-sampalukan DAY 3
7.Kutsay DAY 3
8.Oregano DAY 6
9.Tsaang-gubat DAY 3
10.Dama de Noche DAY 3
Positive control: Povidone-iodine (Betadine) DAY 3
Negative control DAY 6

The table shows the day of scab formation of the different plants. The results show that
the day of scab formation of guava, ginger, garlic, sampa-sampalukan, kutsay, tsaang-gubat and
dama de noche was at day 3. This shows that the scab formation usually starts at the earlier phase
of wound healing where the wound is still fresh.

Conclusion

This study has shown that all of the plant samples exhibited antibacterial properties but
when mixed with PVP-I, the wound healing activity varies. Out of the 10 Philippine plant
samples, only guava, lantana and ginger showed significant wound healing activity. Guava
exhibited the most effect of all of the plant samples and this shows that guava, when mixed with
PVP-I, results in an increase in efficacy in wound healing process and have synergistic effect
with PVP-I. Thus, from the results obtained, guava extracts promote wound healing and can be
more effective when mixed with PVP-I. In conclusion, PVP-Is antiseptic effects could be
further improved when mixed with plant extracts especially guava. With this in mind, the use of
the naturally available plant extracts can be utilized further as adjuncts of PVP-I which can
promote better wound healing properties.

14
Recommendation

There are certain recommendations that are associated with this research. These
recommendations are listed below:

1. Assessment of wound contraction must be monitored by metric measurement and be

measured as a percentage decrease of original wound size.
2. Histopathological study must be performed wherein the granulation tissue obtained after
day 21 from the test and the control group animal to be compared.
3. Determination of the minimum inhibitory concentration must be done to assess more the
antibacterial activity of the plant extracts.
4. Varying concentration of the plant extracts with 10% PVP-I must be performed to
determine if there is a correlation between the dose and rate of wound healing process.
5. Consideration of other possible test parameters regarding the assessment of wound
healing activity like getting the area of the wound instead of just the length could also be
added to support the results of the observations.
6. Further studying of the combination of PVP-I and guava extracts using other test methods
can also be done to determine its full potential towards wound healing.

15
Acknowledgement

It is our pleasure to express our gratitude to all those who blessed us with their presence
in the form of support, encouragements and guidance.

We are indebted to Dr. Evelyn Oda, the Dean of School of Natural Sciences, for the
support not only to us but to all students currently taking up Research subject and to the School
of Natural Sciences.

Our deepest thanks to Mrs. Arlyn Rebogio, the undergraduate program research
coordinator for always guiding and reminding us in the completion and submission of our
outputs.

We will always be grateful to Mrs. Ana Fe B. Revilla, our faculty research promoter, for
constantly reminding and guiding us all throughout the process, from writing up until the
experimentation despite of all the delays and hindrances. We owe you this completion maam.

This thesis would not have been possible without the help of Dr. Ana Mendoza, Dean of
the Veterinary Medicine in Benguet State University for the processing of our Animal Permit.
Deepest thanks to Dr. Eric R. Evangelista, for the help in the incision of the rats also to Mrs.
Rhoda S. Lumang-ay, Mr. Rommel B. Consolacion for assistance and guidance during the
experimentation at Natural Sciences Research Unit (NSRU), to several people who help us
whenever we are confused on what to do, we would like to offer our sincere thanks to all of
them.

Our sincerest gratitude to our parents for their love and encouragement, for being our
constant support system and for providing our needs. We owe you this success.

And most of all to God Almighty, for providing us this opportunity and granting us the
capability to proceed successfully. All the glory is Yours.

16
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20
 APPENDICES

Raw data on Length of Wounds (mm), Animal Permit & Certificate of Authentication of
Plants

Day Day Day Day Day Day Day Day

SAMPLE 1 3 6 9 12 15 18 21
Guava T1 22 15 0 0 0
T2 26 17 4 4 0
T3 17 13 0 0 0
Ginger T1 20 20 18 11 0 0
T2 20 15 13 10 3 0
T3 20 15 9 8 5 0
Lantana T1 25.4 20.3 12.7 10.2 10.2 10.2 10.2 10.2
T2 12.7 10.2 7.6 5.08 0 0 0 0
T3 12.7 7.6 0 0 0 0 0 0
Garlic T1 20 13 12 10 7 4 3 0
T2 17 13 9 5 3 0 0 0
T3 16 13 11 6 4 0 0 0
Lemon T1 20 16 13 10 3 0 0 0
grass T2 14 12 11 10 5 3 0 0
T3 15 13 12 11 8 7 7 6
Sampa- T1 20 18 13 6 3 1 0 0
Sampalukan T2 20 19 13 9 5 5 3 0
T3 25 25 18 15 10 10 8 4
Kutsay T1 17 13 10 8 6 5 3 0
T2 16 14 11 7 5 4 2 0
T3 18 16 13 10 9 7 0 0
Oregano T1 20 17 13 10.5 6.5 3 1.5 0
T2 20 19 16 14.5 11.5 6 3 2
T3 20 18.5 14 12 7 4 2.5 0
Tsaang T1 23 21 20 17 15 died died died
gubat T2 15 14 13 12 10 10 died died
T3 21 19 18 7 6 5 3 0
Dama de T1 25 20 20 16 13 10 6 5
noche T2 20 17 17 16 15 15 12 10
T3 10 7 7 7 16 5 4 3
Povidone T1 15 15 12 6 5 1 0
iodine T2 25 25 15 10 5 1 0
(+) control T3 12 10 10 7 5 0 0
Negative (-) T1 10.2 10.2 7.6 5.1 2.5 1.7 0 0
control T2 15.2 15.2 10.2 10.2 5.1 2.5 1.7 0
T3 17.8 17.8 12.7 10.2 7.6 5.1 2.5 0

21