Production of medicinal extracts from the leaves of tridax procumbens

Production of medicinal extracts from the leaves of tridax procumbens

1. “PRODUCTION OF MEDICINAL EXTRACTS FROM
THE LEAVES OF TRIDAX PROCUMBENSTRIDAX PROCUMBENS {GADDI
CHAMANTHI}”

2. Introduction
Tridax procumbens commonly known as “Tridax daisy” is a species of flowering plant in
the daisy family. It is best known as a wide spread weed .Tridax procumbens is a
spreading annnual herb found throught India , but unfortunately it is one of the neglected
plant .It is found frequently even in RGUKT ,RKV campus.
The plants with antimicrobial potential has become the need of todays research .Although
several works have been studied for antimicrobial susceptability of tridax procumbens leaf
extracts , we targeted to extract the wide range of phytochemicals with variable solvents in
presence a natural catalyst at optimised operation conditions for comparative study .
Research Objectives:
Objective: Objective: Production of medicinal extracts from hot and cold process for aProduction of medicinal extracts from hot and cold process for a
comparative study.comparative study.
Specific objectives:
➔To harvest the plant leaves and preparation of raw materials.
➔To characterise the selected biomass sample.
➔To conduct the soxhelet extraction process for the medicinal extracts by using
solvents like water,ethanol,ethyl actate,haxane.
➔To conduct maceration(cold) process from mixed and water as solvents.
➔To separate the phytochemicals from solvents by using vacuum oven.
➔To characterise the phytochemical extracted qualitatively.

Materials and Methodology
Materials Required
✔Tridax procumbens plant leaves(that source is harvested from RKV campus)
✔Catalyst(Silica)
Chemical Apparatus

3. ✔ Conical flask
✔ Grinder
✔ Seiving set(for size separation of the particles)
✔ Moisture analyzer(for analyzing of moisture in the sample)
✔ Digital weighing machine
✔ Muffle furnace(for proximate analysis)
✔ Soxhelet Extraction apparatus
✔ Steam distillation apparatus
✔ Vacuum oven
✔ Vacuum pump
✔ Petridishes
✔ Armed funnel
✔ Separate condenser
✔ Magnetic Stirrer
✔ Decicator
✔ Freezer
✔ Beakers
✔ Spatula
✔ Test tubes
✔ Measuring jar
Chemicals
 Distilled water
 Hexane
 Ethanol(99.9%purity)
 Ethyl acetate(98%purity)
Others

4. ✔ Handglouses
✔ Scissors
✔ Filter paper
✔ Tissue paper
✔ Mouth masks
✔ Sample covers

1.PLANT ANATOMY
kingdom : plantae (plants)kingdom : plantae (plants)
family : Aster family family : Aster family
scientific name : Tridax Procumbens scientific name : Tridax Procumbens
common name : Coat Buttons (Gaddi chamanthi) common name : Coat Buttons (Gaddi chamanthi)

Tridax procumbens, commonly known as “coat buttons” or “tridax daisy” is a
species of flowering plant in the daisy family. It is the best known as a widespread
weed and pest plant. Tridax procumbens is a spreading annual herb found
throughout India but unfortunately it is one of the neglected plants.

Physical structure :

Habitat: Tropical and sub tropical regions , perennial herbHabitat: Tropical and sub tropical regions , perennial herb
Stem: Cylindrical, branched, erect Stem: Cylindrical, branched, erect
Leaf: Simple, ovate, serrate margin , reticulate,aromaticLeaf: Simple, ovate, serrate margin , reticulate,aromatic
Inflorescence: head or capitulumInflorescence: head or capitulum
Chemical Constituents:
The leaves of Tridax Procumbens is composed of many active
phytochemicals including the presence of carbohydrates , proteins, tannins, steroids,
alkaloids, luteoline, glucolureolin, glycocide and quercetin. The chemical constituents
were separated and purified by column chromatography with “macroporous
adsorption resin diaion HP-20” and “normal phase silica gel” from these five
chemical compounds were isolated and identified such as 8,3'-dihydroxy-3,7,4'-
Trimethoxy-6-O-B-D-flavone glycocide-6'-acetate , stigmasterol, B-sitosterol, Bis(2-
ethylhexyl)phthalate,apigenin-7-O-B-D-glucocide.

fig1.1 Tridax Procumbems plantfig1.1 Tridax Procumbems plant
Fig 1.2: stem of tridax Fig 1.2: stem of tridax
procumbensprocumbens

5. Why Tridax Procumbens is preferable ????
Nature has been a source of medicinal agents for thousands
of years and an impressive number of modern drugs have been isolated from natural
resources .Tridax procumbens is well known ayurvedic, ethino botanical and unani
medicinal plant .



Pharmacological studies have shown that Tridax procumbens possess properties like
anti-inflammatory , heptatoprotective, wound heal , antimicrobial , antiseptic ,
hypotensive and bradycardiac effects when compared to other weed plants. These
properties are due to the presence of active phytochemicals in the leaves of tridax
daisy. So this weed plant is much preferable than other weeds.
3.Methods of extractions
1.Supercritical fluid extraction(SFE):
Supercritical fluid extraction is the process of separating one component from
another using supercritical fluids such as water and carbon dioxide as the extracting
solvent.Loss of desired compounds with improper solvent selection, specialized
equipment is required are the main drawbacks of this extraction method.
leaf of tridax daisy- fig 2.2leaf of tridax daisy- fig 2.2
fig 3.1
leaves of tridax daisy- fig 2.1leaves of tridax daisy- fig 2.1

6. 2.Sonication extraction:
Sonication extraction or Ultrasonic extraction involves the use of ultrasound
ranging from 20 kHz to 2000 kHz . The mechanic effect of acoustic cavitation from
the ultrasound increases the surface contact between solvents and samples and
permeability of cell walls.This extraction is efficient tool for large scale commercial
applications but it shows weak effect on the extraction of oil.
3.Microwave assisted extraction(MAE):
MAE utilizes microwave energy to facilitate partition of analytes from the
sample matrix into the solvent. Microwave radiation interacts with dipoles of polar
and polarizable materials (e.g. solvents and sample) causes heating near the surface
of the materials and heat is transferred by conduction. MAE can provide high
returns on capital investment and it is applicable for both industrial and laboraty
scales but it is expensive equipment and difficult to operate.
fig 3.2
fig 3.3

7. 4.Marinated extraction:
Marinated extraction is an ancient extraction method conducted at room
temperature.For this technique the parameters to be considered include the type of
solvent the ratio of dried herbs to solvent, and the extraction time.This method
requires a prolonged extraction time because it doesn't use heat or other forces.The
advantages of this method is that it requires neither special equipment nor a special
location such as a laboratory.The commony cited disadvantage of this method is he
long extraction time.
5.Hydrodistillation extraction:
Hydrodistillation is a variant of steam distillation.For the hydro distillation
extraction,plant material is soaked for sometime in water after which the mixture is
heated and volatile materials are carried away in the steam,condensed and
separated.Hydrodistillation extraction is inexpensive method because of no organic
solvent is required but it is energy consuming and has the risk of thermal
degradation.
6.Steamdistillation extraction:
Steam distilation is a special type of a separation process for temperature
sensitive materials like natural aromatic compounds.Steam distillation extraction can
extract water insoluble compound but has no agitation needed to accelerate the
process.Water is used as a solvent in this extraction method.
fig 3.4

8. 7.Ultra-high pressure extraction:
A novel extraction technique,ultrahigh pressure extraction has been developed
for the extraction of bioactive compounds from plants,in order to shorten the
extraction time,decrease the solvent consumption and enhance the quality of
extracts.Compound impurity issuse may arise in this extraction method.
8.Accelerated solvent extraction(ASE):
Accelerated solvent extraction is a method for extracting various chemicals
from a complex solid matrix.The process uses high temperature and pressure,which
results in the extraction taking less time and requiring less solvent and possibly also
fig 3.6
fig 3.5

9. giving better analyte recovery,than traditional methods that use less extreme
conditions.The elevated temperature is employed to increase extraction efficiency of
the analyte of interest and the elevated procedure is used to keep the solvent in a
liquid state as the temperature is increased above its boiling point.But this extraction
process is only suitable for the extraction of high temperature stable compound.
9.Hot water extraction:
Hot water extraction is a method of carpet cleaning.It involves a
combination of hotwater and cleaning agents being injected into the fibres of a
carpet at high pressure and all lifted soil being removed by a powerful vaccum.It is
inexpensive method because no organic solvent is required but it is not suitable for
heat sensitive compounds.
fig 3.8
fig 3.7

10. 10.Soxhlet extraction:
A soxhlet extraction is a form of continuous solid-liquid extraction where a
desired compound is extracted from solid material (containing unwanted products )
using a solvent.Soxhlet extraction is used when the desired compound has a limited
solubility in a solvent, and the impurity is insoluble in that solvent.It allows for
unmonitered and unmanaged operation while efficiently recycling a small amount of
solvent to dissolve a larger amount of material.Normally a solid material containing
some of the desired compound is placed inside a timble made from thick filter
paper,which is loaded into the main chamber of the soxhlet extractor.The soxhlet
extractor is placed on to a flask containing the extraction solvent.The soxhlet is then
equipped with a condenser.


Advantages
➢Soxhelet extraction is very simple and inexpensive method.
➢Temperature in the extraction system can be maintained.
➢It is able to extract a difficult to separate solute from insoluble impurities that
otherwise might be inseparable.
➢It has a wide variety of solvents from which to choose allowing optimization
of extraction for a particular solid solute.
➢It is mechanically gentle on the samples bu still efficient in separation.
➢It is often used as a benchmark when deveoping new extraction methods
Soxhelet Extraction set up- fig 3.9Soxhelet Extraction set up- fig 3.9
Feed in the soxhelet extractor- fig 3.10 Feed in the soxhelet extractor- fig 3.10

11. 11.Maceration:
Maceration is an old technic used for medicinal preparation. It is consider as
a widely and low-cost way to get phytochemical’s from plant material. The
maceration is a method is a solid–liquid extraction where the bioactive compound
(solute) inside the plant material is extracted by
soaking the plant material in a s pecific solvent for a period of time. The efficiency
of maceration process is determined by two main factors, solubility and effective
diffusion.
4.FEED SELECTION:



The fine powder of dry leaves of Tridax procumbens is selected as feed for this extraction
process, because Tridax plant has high medicinal content when compared to other medicinal plants
like spinach. Some of the medicinal plants grow at specific conditions and they are not available
at all seasons , but Tridax daisy is available in all the seasons and in all parts of India.It is
unbelievable that a weed plant which commonly grows at roadside can produce useful
phytochemicals, which are useful in pharmaceutical industries and fertiliser industries.We can
prove that a waste plant can act an intermediate substance for pharmaceuticals by selecting “Tridax
Procumbens “ as feed.
5.FEED PREPARATION:
Picking and Drying:
The fresh leaves of Tridax procumbens were collected from the local fields of RGUKT ,RK
Valley campus.The materials were washed with distilled water and air dried at an average
tridax daisy
fig 4.1

12. temperature of 33
0
C

in the sunlight for 10 hours and 30 minutes.The dried leaves were grounded
to coarse powder.
Catalyst preparation:
Silica was selected as catalyst because this extraction is a Catalytic Soxhelet Extraction. 3062.8
grams of dry silica had taken. Contaminants and dust particles were removed by cleaning with
water.The resultant cleaned silica was dried for 4 hours under sunlight.

Seiving:
172.4 grams of the coarse powder of Tridax leaves had taken for seiving by using seiving machine
grinding the grinding the
leaves of Tridax- leaves of Tridax-
fig 5.2fig 5.2
picking the leaves of Tridax Daisy- fig picking the leaves of Tridax Daisy- fig
5.15.1
cleaning the silica-fig 5.3cleaning the silica-fig 5.3
drying the silica- fig drying the silica- fig
5.45.4

13. . 103 grams of the seived powder having 150 micrometer [mesh number =100] of particle size is
considered for the soxhelet extraction. And 3063.6 grams of dry silica was seived to get 876.6
grams of seived silica having 150 micrometer of particle size is also considered for this extraction.
6.Feed Characterization:
The sieved powder of Tridax Daisy leaves and silica was tested for physical properties like
moisture analysis , Bulk Density , Specific gravity etc.,
Moisture analysis:

Bulk Density calculation:
For Tridax powder:
Density = sample weight of tridax sample weight of tridax/ / sample volume of tridaxsample volume of tridax = 10 g/ 27 ml = 0.370 g/ml
For water:
Density = sample wt of water / sample volume of water sample wt of water / sample volume of water = 10 g/11ml = 0.909 g/ml
Specific Gravity calculation:
moisture analyser- fig moisture analyser- fig
6.16.1
moisture analysis of Tridax moisture analysis of Tridax
powder -fig 6.2powder -fig 6.2
moisture analysis of sandmoisture analysis of sand
-fig 6.3-fig 6.3
seiving machine- fig 5.5seiving machine- fig 5.5

14. Density of tridax / Density of waterDensity of tridax / Density of water = 0.370 / 0.909 = 0.40

7.Solvent Selection:
Many literatures and research institutions used many solvents like ethanol,ethyl
acetate,hexane,water,chloroform,methanol etc., in the soxhelet extraction.The efficiency of
extraction depends on the type of solvent used.
We used monosolvents like ethanol,ethyl acetate,hexane etc. and mixture of solvents and
with water in this extraction process.
8.Extraction Method Selection :
“Hot continuous catalytic solvent extraction “ is the best extraction method among all other
methods. Because it is inexpensive and simple method .We can use various solvents in this
extraction.Temperature in the extraction system can be maintained.It acts as a
benchmark to develop other extraction methods.
Procedure for Hot process i.e.Soxhelet Extraction:
➢Soxhelet extraction is a continuous process of extraction with a hot organic
solvents like ethanol, ethyl acetate, hexane and water etc,.
➢20 g of the powdered tridax plant material is taken along with the 40 g of
natural catalyst i.e.silica (we can extract without using catalyst) in a thimble
which is placed in the soxhlet extractor.
➢ The extractor,which has a siphoning system,is fitted on the top of a round
bottom flask.A condenser is fitted at the top of the extractor.
➢500 ml of the extracting solvent is poured into the flask placed on a heating
mantle.
➢On heating, the solvent evaporates,raises to the condenser,where it condenses
and drains back to the extractor holding the thimble with the feed.
➢When the extractor becomes full with the hot solvent.The solvent siphons
down to the flask along with the extracted constituents.
➢The recycling of the evaporated solvent is allowed to continue until the
extraction is complete.

15. Note: Same procedure was followed for all solventsNote: Same procedure was followed for all solvents
Vaccum Woven:
The extracted solution in distillation flask was taken into small petridishes and kept in vacuum
woven to vapourise the solvents and to remain extract .We can save some amount of solvent from
vapourisation by attaching a condenser to the vaccum woven.

The resulted extracts were stored in refrigerator.
Test For Phytochemicals:
The selected plant extracts were subjected to quantitative chemical investigation
to test for the presence of various phytochemicals in extracts. The resulted were noted in
the following table:
Procedure for analysis of Extract extracted by using the solvents :
s.no Experiment Procedure Inference(pres
ent/ absent)
extraction process- fig 8.1extraction process- fig 8.1
vaccum woven- fig 8.2

16. 1Test for CarbohydratesMolisch’s Test:
To 2ml of extract 2-3 drops of alpha
naphthalene solution in
alcohol was added, shaken for 2 min and
1 ml of concentrated sulphuric acid was
added slowly from the sides of the test
tube. A deep violet colour at the junction
of two layers indicates the presence of
carbohydrates.
2Test for Proteins Xanthoprotein Test:
To 3 ml of extract 1ml of concentrated
sulphuric acid was
added. The appearance of white
precipitate which turn to yellow on
boiling and orange on addition of
ammonium hydroxide (1ml) indicates
the presence of proteins containing
tyrosine tryptophan.
3Test for Tannins and
Phenols
Lead acetate Test:
To 3ml of extract, 3 ml of lead acetate
solution was added.
The occurrence of white precipitates
indicates the presence of tannins and
phenols
4Test for saponins Foam Test:
The extract (2g) was shaken vigorously
with 20 ml of water and
observed for persistent foam, which
indicates the presence of saponins.
5Test for TriterpenoidsSalkowaski Test : To 2 ml of extract 5
drops of concentrated sulphuric acid was
added, shaken and allowed to stand.
Appearance of greenish blue colour
indicates the presence of triterpenoids.
Analysis for Extracts extracted by using the solvents
S.NoName of the ExtractExperiment Inference(present/absent)
1 Ethanol Test for carbohydrates
Test for Proteins
Test for Tannis and
Phenols
Test for saponins
Present
Absent
Absent
Present

17. Test for TriterpenoidsAbsent
2 Ethyl Acetate Test for carbohydrates
Test for Proteins
Test for Tannis and
Phenols
Test for saponins
Test for Triterpenoids
Present
Absent
Present
Present
Present
3 Hexane Test for carbohydrates
Test for Proteins
Test for Tannis and
Phenols
Test for saponins
Test for Triterpenoids
Present
Absent
Present
Present
Absent
Conclusion:
Although several works have been studied for antimicrobial susceptability of tridax
procumbens leaf extracts, we targeted to extract the wide range of phytochemicals with
variable solvents in presence a natural catalyst at optimised operation conditions for
comparative study .
The leaves of this plant have been claimed to be useful in the treatment of
inflammatory conditions and have tendency to heal wound , antidiabetic activity
,preventing hairloss, diarrhoea and serve as insect repellent . The future scope of this work
may include antibacterial studies for applied industries, drug formulation and process
simulation etc.,
It possess antiseptic insecticidal parasiticidal and hepate protective properties and has
marked depressent action on respiration. This extract can widely used in cancer treatment .
It can also useful to prevent insects which affect agricultural fields.
We have proved that a weed plant which actually used to make ayurvedic medicines can
also useful in making Allopathic medicines.
Prepared By ,
B.Ravi Kumar,
K.Pushpa Jyothi,
P.Nandini,
D.Harshitha
Under the guidance of Prof.A.Ramesh Babu, Dept. Of Chemical Engineering ,RGUKT,
RK Valley.

18. GRATITUDE:
We are very thankful to our Ramesh babu sir,who paved the path for our project.We are
sincerely thankful to the HOD of chemical engineering and HOD of chemistry,as they
permitted us to use the available lab equipments.A gratitude towards our honourable
Director and the Administrative Officer as they permitted us to exhibit our project.We
have completed our successfully because of the help of all the Lab Assistants.
Reference:
1] plant anatomy ;
https://plants.usda.gov/core/profile?symbol=trpr5
physical structure;
https://www.cabi.org/isc/datasheet/55072
chemical constitutions:
https://www.researchgate.net/publication/42973055_Chemical_Profile_of_Tridax_procumbens_Li
nn
2] why tridax is selected???
https://www.researchgate.net/publication/271111486_TRIDAX_PROCUMBENS_MEDICINE_F
OR_LIFE_AN_OVERVIEW
3] methods of extraction:
Appel, B.R. et al. (1980). Evaluation of techniques for sulfuric acid and particulate
strong acidity measurements in ambient air. Atmos. Environ. 14, 559-563.
Bristol, D.N., Crist, M.L., Lewis, R.G., MacLeod, K.E. and Sovocool, G.W.
(1982). Chemical analysis of human blood for assessment of environmental
exposure to semivolatile organochlorine chemical contaminants. J. Ana!. Toxico!.
6(6), 269-275.
8] procedure for soxhelet extraction:
https://www.slideshare.net/AbarnaAbi1/soxhlet-apparatus
9] list of phytochemicals:
http://www.phytochemicals.info/plants-list.php