WOLLO UNIVERSITY

KOMBOLCHA INSTITUTE OF TECHNOLOGY

SCHOOL OF TEXTILE, LEATHER AND FASHION TECHNOLOGY
DEPARTMENT OF TEXTILE ENGINERING

EXTRACTION, APPLICATION, AND CHARACTERIZATION OF

ANTIMICROBIAL FINISHING AGENT FOR COTTON FABRICFROM
SENSEL LEAVES

BY

NAME ID NO
1. Mintesnot kefale ………………………1164/08
2. Moges dessie ………………………….1174/08
3. Muluken gidi ………………………….
4. Khalid sisay

UNDER THE GUIDANCE OF

FENTA MEHABIE
NOV, 2013
WOLLO ETHIOPIA
 CHAPTER ONE
1.1BACKGROUND OF THE STUDY
Textiles have long been recognized as media to support the growth of microorganisms such as
bacteria and fungi. These microorganisms are found almost everywhere in the environment and
can multiply quickly when basic requirements, such as moisture, nutrients and temperature are
met. The growth of microorganisms on textiles inflicts a range of unwanted effects not only on
the textile itself but also on the wearer. These effects include the generation of unpleasant odor,
skin irritation and contamination for cross infections, stains and discoloration in the fabric, a
reduction in fabric mechanical strength and an increased likelihood of contamination. Such bottle
neck problem forced attention towards developing advanced textile based medical products. The
rapid growth in medical and wellness textiles has evolved many opportunities for the application
of innovative functional finishes.

Antimicrobial finishing of textile materials aims to protect the users from the harmful
microorganisms and to ensure a proper functionality of textiles. An ideal antimicrobial treatment
should provide protection against a wide range of bacterial and fungal species. A number of
chemicals such as, benzophenone, organometallics, iodophors (substances that slowly release
iodine, formaldehyde derivatives and amines have been employed to impart antimicrobial
activity to textiles. Majority of these antimicrobial agents utilized in textile industry are of
leaching type and thus their gradual release from textiles into surroundings results in decrease of
their concentration and fail to inhibit the growth of harmful microbes. Also the release of these
agents acts as poison to a wide spectrum of bacteria and fungi. Owing to this, the concentration
of the active substance in the textile decreases and gradually falls under the limit of
effectiveness. To overcome such problem, researchers and the textile industry looks for non-
toxic, non-allergic and eco- friendly natural antimicrobial agents that do not adversely affect the
quality of the textile material as substitute for synthetic toxic chemicals.

The herbal antimicrobial finishes overcome the disadvantages of the chemical finishes because
they are eco-friendly, non-toxic and also non allergic. Even though, the use of antimicrobials
have been known for the decades, it is only the recent couple of years, and several attempts have
been made on finishing textiles with antimicrobial compounds. A number of studies are
mentioned in the literature in which medicinal.

1.2 PROBLEM STATEMENT

Majority of the antimicrobial agents utilized in textile industry are of leaching type and thus their
gradual release from textiles into surroundings results in decrease of their concentration and fail
to inhibit the growth of harmful microbes. Also the release of these agents acts as poison to a
wide spectrum of bacteria and fungi. Owing to this, the concentration of the active substance in
the textile decreases and gradually falls under the limit of effectiveness. In recent years, the
textile industry looks for non-toxic, non-allergic and eco-friendly natural antimicrobial agents
that do not adversely affect the quality of the textile material as substitute for synthetic toxic
chemicals. The herbal antimicrobial finishes overcome the disadvantages of the chemical
finishes. The antimicrobial compounds, which are mostly extracted from plants, include tannins,
flavonoids, Terpenoids, alkaloids, essential oils, phenolic and polyphenols. These components
show, not only antimicrobial, but also antioxidant properties. In addition, some studies have been
reported the antimicrobial activity of Justicia Schimperiana. But, most of the reviewed papers
concentrate on the technical details of applying natural agents, such as Neem extract, chitosan,
tea tree and aloe Vera on textile finish. To overcome consumer health problems faced by the
consumers due to environmental hazards like microbes, it is of great concerns to use of Justicia
Schimperiana for valuable product. In the other side, no study has been undertaken to investigate
the antimicrobial effect of Justicia Schimperianaextract on cotton fabric.

Thus, the aim of this present study will be to identify the phytochemical constituent and
investigate the antimicrobial activity of aqueous extract of Justicia Schimperiana on cotton
fabric finish.
 1.3 OBJECTIVE
1.3.1 General objective
 Extraction, application, and characterization of antimicrobial finishing agent for
cotton fabric from sensel leaves

1.3.2 Specific objective

The specific objective of this research will be
 To determine the yield of aqueous extract of sensel
 To identify the bioactive metabolites present in the leaf of sensel
 To treat cotton fabric by the extracted components
 To see the effect of concentration on selected test microorganism (Staphylococcus aureus and
Escherichia coli) finished cotton fabric with extract of sensel
 To characterize the treated cotton fabric (tensile strength, wash durability etc.).

1.4. SCOPE OF THE STUDY

This research will be focused on extraction of antibacterial agents from the sensel leaf, treat
cotton fabric by the extracted active components and the antibacterial activity of aqueous extract
of sensel leaf on cotton fabric.
 CHAPTER TWO
LITERATURE REVIEW
Functional fabrics have become an important issue in the textile industry, especially the
antibacterial ones. Generally, fabrics provide a good contact area and can absorb moisture, both
required for microbial growth. This growth can lead to malodours, dermal infections, allergic
responses and fabric deterioration (Christie Jennifer 2016) Thus, the incorporation of
antibacterial agents on textile products, able to overcome these problems, is of atmost
importance.

Plants and plant product are traditionally used for healing of wounds, burn injuries, anti-fungal,
anti-viral, anti-bacterial and anti-microbial activity against skin infections. Plant extract for
antimicrobial finishing in textiles because of the excellent antimicrobial and ecofriendly
properties exhibited by them. There are several methods of extraction used for plant extracts. The
sole purpose of basic extraction procedures is to obtain the therapeutically desirable portion and
eliminate the inert material by treatment with a selective solvent known as the Menstruum

Various approaches have been used for antimicrobial functionalization of textile materials
depending on the particular active agent and fibre type. In general, two different antimicrobial
finishing methods can be distinguished. Antimicrobial agents can be either applied as an after-
treatment process or incorporated into the polymer solution prior to extrusion or into the spinning
bath. The effectiveness of anti-bacterial activity of treated fabric was evaluated qualitatively and
quantitatively.

2.1 BACTERIA
Bacteria are unicellular organisms which can further be subdivided as either gram positive (e.g.
Staphylococcus aureus) or gram negative (e.g. E. coli), or spore forming and non-spore forming
types. Bacteria grow very rapidly under warmth and moisture (Sood 2014). They have a short
generation time, lasting from minutes to hours, being able to rapidly respond to changes in their
environment (Morais, Guedes, and Lopes 2016). The bacteria population, will double every 20 to
30 min under ideal conditions (36–40 °C or 77–98 °F, pH 5–9). At this rate, one single bacteria
cell can increase to 1048 576 cells in just 7 hours (Boryo 2013). Estimations have shown that the
production of antimicrobial textiles was in the magnitude of 30,000 tones in Western Europe and
100,000 tones worldwide in 2000 (Zanoaga 2014). Furthermore, it was estimated that the
production rate will increase, making it one of the fastest growing sectors of the textile market.

2.2 SOURCE OF BACTERIA

The main source of bacteria is in the air we breathe, in the soil, in our skin and bodies &
everywhere. The human skin is usually crowded with innumerable microbes. In favorable
conditions certain bacteria can grow from a single germ to million in a very short period of time.
They can double every 20 to 30 minutes in a warm and moist micro climate that has plenty of
food for them, e.g. perspiration and other body secretion, skin particles, fats and leftovers from
air we breathe, in our skin and bodies worn out threads (Pannu, 2013). The common promoters
are;

 Moisture  Food particles

 Receptive surface  Textile finishes
 Perspiration

2.2.1 Effects of bacteria on textile material and wearer

Textiles provide an excellent medium for the adherence, transfer and propagation of infection-
causing bacteria. The detrimental effects of microbes can be seen on the wearer and on the fabric
itself. Due to the microbial attack, the results are loss of the performance properties of textile
substrate, odour formation, contamination risk, strength reduction, quality loss.

It is becoming increasingly important to maintain hygiene through the effective use of textile
materials. With a view to improve the performance properties of the textile substrate and protect
the wearer, the antimicrobial finish is applied to textile materials (Sood 2014). Bacteria causes,
some fibre damage, unpleasant odours, and a slick, slimy feel. In extreme cases, microorganisms
can cause serious problems, include fabric rotting, staining, unpleasant odors and health concerns
ranging from simple discomfort to physical irritation, allergic sensitization, toxic responses,
infection and disease.
Many of the characteristic malodors associated with the human body are due to the presence of
large populations of bacteria. Most importantly, biodeterioration of textile materials is noticed as
staining, discoloration, and pigmentation, as well as loss of tensile strength, elasticity, and
integrity of the products (Sun 2016).

2.2.2 Antibacterial agent

Antibacterial agents are substances which are intended to kill or inhibit bacteria. Substances
which shall prevent harmful effects of bacteria on human beings, animals or material are called
biocides (Pannu 2013). An ideal antimicrobial finish must control, destroy or suppress the
growth of microorganisms. Only this can prevent their negative effects, such as odour, staining
and deterioration. A common hazard in hospitals and other health care centres is bacterial
contamination of textile fabric surfaces. Fabric contamination can lead to infection and cross-
infections. So, it is very important to reduce the migration of pathogenic bacteria between people
and clothing, by using antimicrobial finished textiles (Zanoaga 2014).

2.2.3 Types of antibacterial finishing agent

According to the origin, there are two types of antimicrobial finishing agents, these are natural
and synthesis. According to the mod of action also leaching and none leaching and based on the
antimicrobial power they are divided into biostatic and biocides.

I. Synthetic antibacterial agents

synthetic agents are triclosan, polybiguanides, N-halamine and halogenated phenols (Naveed
2018). Quaternary ammonium compounds, silver, polyhexamethylene biguanides (PHMB)
(Coman 2015). Use of these antimicrobial compounds may result in a shift in the skin microflora
composition and lead to outgrowth of pathogen.

The synthetic antimicrobial agent shows good antibacterial power and durability, but toxic nature
of some artificial agents and their non-degradability in the environment has shifted the
inclination of their utilization towards natural products as viable alternatives.
 II. Natural antibacterial agents
Plants possess a wide variety of secondary metabolites which have been found in vitro to have
antimicrobial properties. Extracts from different parts of diverse species of plants like roots,
leaves, flowers, and seeds exhibit antimicrobial properties. With a rising interest in personal
health and hygiene, the market for functional textiles is steadily increasing.

Leaves of eucalyptus (Eucalyptus globulus), Tinospora (Tinospora cordifolia) and tridax (Tridax
procumbens (Byadgi 2018). Curcuma aromatica (Kasthuri manjal), Camellia sinensis (Green tea)
and Azadirachta indica (Neem) herbal powders extracted withdistilled water (Sumathi 2017).
These all showed varying degrees of antimicrobial activity against Gram positive and Gram-
negative micro-organisms.Bidenspilosa (whole plant), Bischofiajavanica (leaves),
Elmerilliapapuana (root) and SigesbekiaOrientalis (whole plant) with petrol, dichloromethane,
ethyl acetate and tested these extracts for antibacterial efficacy by the disc diffusion method. The
crude ethanolic extracts and all the obtained fractions showed a broad spectrum of antibacterial
activity whereas the ethyl acetate fractions and the petrol fraction of E. papuana were the most
effective (Sood 2014).

2.3 EXTRACTION AND APPLICATION TECHNIQUES FOR PLANT EXTRACTS

2.3.1 Extraction techniques for plant extracts
There are several methods of extraction used for plant extracts. The main purpose of basic
extraction procedures is to obtain the therapeutically desirable portion and eliminate the inert
material by treatment with a selective solvent known as the Menstruum. The knowledge of the
pharmacognosy of the drug is essential for selection of the extraction process that will give the
best results. The factors which affect the choice of extraction process include character of the
drug, therapeutic value of the drug and stability of drug. The techniques used for extraction of
plant extracts are as follows:

Maceration: This is the most common and simple method of extraction. It involves occasional
shaking of pulverised material overnight with a solvent followed by filtration and evaporation of
the solvent. Repeated maceration may be more efficient than a single maceration, since an
appreciable amount of active principle may be left behind in the soaked plant material after
pressing for the first-time during maceration process. The repeated maceration is more efficient
in cases where active constituents are more valuable (Seidel 2012).

Hot continuous extraction Soxhletion: This method is commonly used to prepare the crude
extracts using solvents. In this method, the material to be extracted is placed in a thimble made
of cellulose or cloth in a central compartment with a siphoning device and side arm connected to
a lower compartment. The solvent is placed in a lower compartment and a reflux condenser is
attached above the central compartment. The warm solvent percolates through the powder and
the extract gradually collects in the central compartment. Once the extract reaches the top of the
siphon, the entire liquid circulates through the central container and back into the lower
compartment. The process is repeated till the powder decolorizes (Handa 2008).

Ultrasonication: It involves suspending the pulverized material in the solvent and subjecting it
to cavitations under the influence of ultrasonic vibrations. After the filtration and evaporation of
the extractor solvent, the dry extract can be obtained (Sood 2014).

Microwave assisted extraction: Microwaves are made up of two oscillating perpendicular

field’s i.e. electric field and magnetic field and the former is responsible for heating. The
moisture when heated up inside the plant cell due to microwave effect, evaporates and generates
tremendous pressure on the cell wall due to swelling of the plant cell that bursts and the
extractable leach out in the solvent (Doughari 2012).

2.3.2 Application techniques for plant extracts

Various approaches have been used for antimicrobial functionalization of textile materials
depending on the particular active agent and fibre type. In general, two different antimicrobial
finishing methods can be distinguished. Antimicrobial agents can be either applied as an after-
treatment process or incorporated into the polymer solution prior to extrusion or into the spinning
bath (Sood 2014). For antimicrobial finishing of natural as well as synthetic fibers, conventional
exhaust and pad-dry-cure methods have been used as after-treatment processes (Doughari 2012).
2.4 GENERAL TEST METHODS FOR ANTIMICROBIAL ACTIVITY
2.4.1. Agar diffusion test
Agar diffusion test is a preliminary test to detect the diffusive antimicrobial finish. It is not
suitable for non-diffusive finishes and textile materials other than fabrics. The agar diffusion
tests include AATCC 147-2004 (American Association of Textile Chemists and Colorists), JIS
1902-2002 (Japanese Industrial Standards) and SN 195920- 1992 (Swiss Norm). They are only
qualitative, but are simple to perform and are most suitable when a large number of samples are
to be screened for the presence of antimicrobial activity (Moustafa 2013).

2.4.2. Suspension test (bacterial reduction methods)

Suspension test method isillustrated by AATCC 100-2004, JIS L 1902-2002 and SN 195924-
1992.These methods provide quantitative values on the antimicrobial finishing, but are more
time-consuming than agar diffusion tests. Typically, a small volume (e.g. 1 ml) of bacterial
inoculums in a growth media is fully absorbed into fabric samples of appropriate size without
leaving any free liquid. This ensures intimate contact between the fabric and the bacteria. After
incubating the inoculated fabrics in sealed jars at 37°C or 27°C for up to 24 h, the bacteria in the
fabric are eluted and the total number is determined by serial dilution and plating on nutrient agar
plates.

Antimicrobial activity, expressed as percentage of reduction, is calculated by comparing the size

of the initial population with that following the incubation. Appropriate controls, e.g. samples
that have gone through the same processing except the antimicrobial finishing, should be
included in each experiment to ascertain that the observed decrease in bacterial number is truly
due to the antimicrobial finishing (Moustafa 2013).

The dynamic shake flask test method

The dynamic Shake Flask Test Method is designed to evaluate the resistance of non-leaching
antimicrobial treated specimens to the growth of microbes under dynamic contact conditions.
This dynamic shake flask test was developed for routine quality control and screening tests in
order to overcome difficulties in using classical static antimicrobial test methods to evaluate
substrate-bound antimicrobials. These difficulties include ensuring contact of inoculum to treated
surface (as in AATCC 100), flexibility of retrieval at different contact times, use of
inappropriately applied static conditions (as in AATCC 147), sensitivity and reproducibility (Sun
2016).

Mechanism of antibacterial agent

Table2.1 Mechanism of antibacterial agent
Phytochemical Active components Mechanism
s
Quercetin & chrysin Inactivate enzyme, Bind to adhesin,
Flavonoids complex with cell wall
Quinones Hypercin Inactivate enzyme, Bind to adhesin
Enzyme inhibition, complex with cell
Tannins Ellagitannin wall, metal ion complexion, membrane
disruption
Polyphenols Catechol & catechin Inactivate enzyme, substrate deprivation
Terpenoids Capsaicin Membrane disruption, antiviral activity
Alkaloids Berberine Intercalate into cell walls

The finished antibacterial compounds in the fabric samples disruptthe cell membrane of the
bacteria through the physical and ionic phenomenon. The finishing agent inhibited the growth of
test organisms by using an electrochemical mode of action to penetrate and disrupt their cell
walls. When the cell walls are penetrated, leakage of metabolites occurs and other cell functions
are disabled, thereby preventing the organism from duplication (Sumathi 2017).

Research gaps identified

In present project Justicia schimperiana extracts will be apply to textile as preliminary
experiment. Justicia schimperiana not only used as an antimicrobial agent but also heals different
skin disease. Antimicrobial finishing agents used in textiles are mostly created in laboratory
(synthetic) no matter how or where the finish originates but eco-friendly nature for wearer and
for environment were not fully studied. Natural products particularly derived from plants; on the
account of their plenty availability, green approach, low toxicity, biocompatibility and eco-
friendly nature are accepting popularity all the world-wide for their utilization in textiles (Jaswal
2017).
 CHAPTER THREE
MATERIAL AND METHODS
3.1. CHEMICALS AND MATERIALS
Mainly the following chemicals and materials we be used to accomplish this research.
Table 3.1 Chemicals and materials
Materials Function
Justicia schimperiana leaf Plants for extract
Distilled water Extractive solvents
(100%) Pretreated cotton woven fabric Test specimen(fabrics for treatment)
Citric acid Cross linking agent
Agar powder (to prepare agar media (solidifying agent)
Staphylococcus aureus (S.aureus) Gram positive bacteria
Escherichia coli (E.coli) Gram negative bacteria

3.2 SAMPLE SELECTION

For this study herbal plant parts which is Justicia schimperiana was selected for obtaining the
antimicrobial extract. The herb and the vegetable part were selected because they exhibited anti-
microbial activity properties, locally available and low cost.

3.3 SAMPLE COLLECTION AND PRETREATMENT

The plant material which is Justicia schimperiana was collected from its natural habitat in
November 2013 around kemissie town 20km away from kombolcha, Amhara regional state. The
collected sample from the available area was washed in tap water and rinsed in distilled water to
remove dust and other impurities. Then the rinsed samples were dried in shade for 14 days and
the dried samples were subjected for size reduction to powder (finely) by high speed
multifunctional grinder.
 Figure3.1 jstica schimpriana
3.4 EXTRACT PREPARATION
Extraction will be done using a water bath method. The dried powder, 20 gram of sample will
soaked in 400 ml (MLR 1:20) of distilled water for 6 hour at 80 0c temperature. After extraction,
the perspective sample will filtered through Whatman filter paper (No.1). After filtration, the
filtrates will be placed by plastic bottle and stored in a refrigerator at 4℃ waiting for treat cotton
fabric.The yield of the extraction will be calculate by

w 1−w 2
Yield ( % )= 100
w2

3.5 ANTIMICROBIAL FINISH APPLICATION ON COTTON FABRIC

The fabrics will be treated in solution prepared by different concentration of extracted of Justicia
schimperiana for 30 minutes and padded on padding mangle individually in the presence of citric
acid to maintain 5.5pH to get a wet pick up of 80% on weight of the fabric. The fabric will be
then dried at 80 ℃ for 3 minute and cured at 130 ℃ for 2 minute on theoven drier.
3.6 ANTIMICROBIAL ACTIVITY ASSESSMENT
AATTC Test Method 147-2004 Parallel Streak Method (Pinho et al. 2011)Specimens of the test
material including corresponding untreated controls of the same material are placed in intimate
contact with the agar surface which has been previously streaked with an inoculums of a test
bacterium. After incubation a clear area of interrupted growth underneath and along the sides of
test material indicates antibacterial activity of the specimen. A standard strain of bacteria is used
which is specific to the requirements of the material under test. The average width of a zone of
inhibition along a streak on either side of the test specimen was calculated using the following
equation:
T −D
w=
2
Where: W is width of clear zone of inhibition in mm T is total diameter test specimen and clear
zone in mm D is diameter of the test specimen in mm.

3.7 WASH DURABILITY TEST

In this study, the wash durability antimicrobial activity of the treated cotton fabric will be
evaluated at different wash cycles. Each sample will be washed with 5% neutral soap solution for
20 minutes. Each of the washed samples will be tested for the retention of antimicrobial activity
after 1, 5 and 10 launderings by AATCC-100 test method.
 CHAPTER FOUR
RESULT AND DISCUSSION
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