Determination of the antioxidant

property from the leaves extract

of Decaspermum fruticosum
(Daniri)
Baluyut, Karla
Baluyut, Patricia
Basilio, Marnie
Dagal, Reygie
Esguerra, Katherine
Yun, Elma
 Extraction
Schematic Diagram Sequential Extraction
Collection of Daniri leaves Weigh 200 g powder extract (sequence for 8 hrs.
⬇ with phenol using soxhlet apparatus)
Wash with water ⬇
⬇ Filter
Wt. 50 g (air dried) pulv. ⬇
⬇ Filtrate (concentrated to obtained phenol extract)
Macerate with 250 mL phenolic solution (24 hrs) ⬇
⬇ Extract conc. With rotary evap and dried oven
Filter @35 – 40 °C
⬇
50 mL filtrate (evaporate to dryness) 105°C and
weigh)
⬇
Calculate % Phenolic extract
 Extraction
PREPARATION OF PHENOLIC
COLLECTING/DRYING METHOD COMPOUND ASSAY
Collection of Daniri leaves 1mL extract + 2mL D. Water + 0.5 Folin Ciocalteu
⬇️ rgnt (3mins)
10g of leaves wash w/ 100mL ultra pure water ⬇️
⬇️ + 0.5mL 10% NaCO3 soln (mix and stand for 1hr
Water bath ➡ boiled @ 250 mL erlenmayer flask @ room temp and dark room)
(20mins) ⬇️
⬇️ Measure __nm w/ UV Visible spectrophotometer
Filter and stored @ -15C (1wk) ⬇️
⬇️ Caffeic â soln; construction of calibration curve
Filter w/ Whatman no.1 filter paper
 Phytochemical Screening
• Test for alkaloids
Solvent free extracts, 50 mg was stirred with few ml of dilute hydrochloric acid and
filtered separately. The filtrate was tested carefully with various alkaloid reagents as
follows:
• Mayer’s test
To 1 ml filtrate of the extract, 0.5 ml of Mayer’s reagent (potassium mercuric iodide)
was added by the side of the test tube. A white or creamy or yellow colour precipitate
was formed and that was indicated as the presence of alkaloids.
• Wagner’s test
To 1 ml filtrate of the extract, 0.5 ml of Wagner’s reagent was added by the
side of the test tube. Reddish brown precipitate was formed and that was indicated as
the presence of alkaloids.
 Phytochemical Screening
• Hager’s test
To 1 ml filtrate of the extract, 0.5 ml of saturated picric acid solution (Hager’s
reagent) was added by the side of the test tube. Yellowish precipitate was
formed and that was indicated as the presence of alkaloids.
• Dragendroff’s test
To 1 ml filtrate of the extract, 0.5 ml of Dragendroff’s reagent (bismuth nitrate)
was added by the side of the test tube. A prominent yellow or orange brown
precipitate was formed and that was indicated as the presence of alkaloids.
• Test for flavonoids
The extracts were treated with few drops of 10% lead acetate solution. The
formation of yellow precipitate confirmed the presence of flavonoids.
 Phytochemical Screening
• Test for reducing sugars
• Extracts were dissolved individually in 5 ml of distilled water and filtered. The
filtrates were used to test the presence of carbohydrates.
• Benedict’s test
• Filtrate was treated with Benedict’s reagent and heated on water bath.
Formation of an orange red precipitate indicated the presence of reducing
sugars.
• Fehling’s test
• Filtrate was hydrolyzed with dilute hydrochloric acid, neutralized with alkali
and heated with Fehling’s A and B solutions. A red precipitate was formed
which indicated the presence of carbohydrates.
 Daniri Study
• Decaspermum fruticosum or
commonly called Daniri in tagalong • Flowers: white or pink
and Dugayon is found, often common, • Fruit : Rounded, flattened 5-8mm
throughout the Philippines in all or thick, and terminated by calyx
most islands and provinces. It is also crown which turns red when ripe.
occurs in visayas in India and China
• Properties:
through Malaya to tropical Australia
and Polynesia. >Astringent
• Decaspermum fruiticosum is an erect >Antydysenteric
shrub or small tree. the leaves are • Studies:
opposite, ovate or lanceolate -ovate,
>Inhibitory activity on ligand
up to 8cm or more in length, 1 to 3 cm
in width, pointed to blunt at the base,
Binding on 5HT1a receptor
pointed at the tip, smooth when old, >Anti-Babesial activity
and often silky beneath when young.
 Family with antioxidant property
• Family • Myrtaceae
• Makopa
• Syzygium malaccense
• Botany:
Makopa is a tree reaching a height of 10 meters. Leaves are pinkish when
young. Older leaves are large, drooping, elliptic-oblong to broadly oblong-
lanceolate, 15 to 30 centimeters long, 7 to 15 centimeters wide, narrowed and
pointed at both ends. Flowers are large, showy, crimson, 5 to 6 centimeters in
diameter, borne on the branches below the leaves, clustered on short, few-
flowered racemes, 6 centimeters long or less. Fruit is shiny, oblong or pear-
shaped, 5 to 7.5 centimeters long, either white splashed, striped with pink, or
wholly crimson to purplish, and slightly shiny, seedless or one-seeded. Flesh is
white, pithy, juicy. Although rather tasteless, some varieties have a pleasant
flavor.
• Study:
Antioxidant: Study of 58 underutilized Malaysian fruits of 32 different species,
showed fruits from some genera, including Syzygium, had higher antioxidant
capacity compared to other genera.
Antioxidant / Leaves: A methanolic extract of fresh leaves exhibited high
antioxidant activity with DPPH and hydroxyl radical scavenging assays. A strong
correlation was noted with phenolic and flavonoid contents.
Antioxidant / Pulp in Ripe and Unripe State: Study of both extracts of ripe and
unripe rose apple fruits showed high antioxidant activity and free radical
scavenging; however, the unripe rose apple showed greater activity
 Family with antioxidant property
• Family • Myrtaceae
• Bayabas
• Psidium guajava Linn.
• Botany:
Bayabas is a somewhat hairy plant reaching a height of 8 meters. Young
branches are 4-angled. Leaves are opposite, oblong to elliptic, and 5 to 1
centimeters long, the apex being pointed, and the base usually rounded.
Peduncles are 1- to 3-flowered. Flowers are white, 3 to 3.5 centimeters across,
with in-curved petals, coming out solitary or two to three in the leaf axils.
Numerous stamens form the attractive part of the flower. Inferior ovaries
develop into round or obovoid green fruits 4 to 9 centimeters long, turning
yellow on ripening and have edible, aromatic, seedy pulp.
• Properties
Antidiarrheal, antiseptic, antispasmodic, antioxidant hepatoprotective, anti-
allergy, antimicrobial, antigenotoxic, antiplasmodial, cardioactive, anticough,
antidiabetic, antiinflammatory, antinociceptive.
Bark and leaves are astringent and vulnerary.
• Study:
Antioxidant / Leaves: Study investigated the antioxidant activity of Psidium
guajava leaf extract for antioxidant activity by DPPH free radical scavenging
method using ascorbic acid as standard. The leaf extract showed strong
antioxidant activity. IC50 of the P. guajava extract was 45.5 ± 0.044 µg/mL
compared to ascorbic acid standard of 25.8 ± 0.204 µg/mL. (55)
 Terminologies

1. Folin & Ciocalteu′s phenol reagent - is most commonly used in the Lowry method
for determining protein concentration.It has also been used for the quantification of
total phenolics. In this method, protein is pretreated with copper(II) in a modified
biuret reagent (alkaline copper solution stabilized with sodium potassium tartrate).

http://www.sigmaaldrich.com/catalog/product/sial/f9252?lang=en&region=PH
 Terminologies
• 2. Antinociceptive - reducing sensitivity to painful stimuli.
• http://medical-dictionary.thefreedictionary.com/antinociceptive
 Terminologies
3. Anti-babesial activity - countering or preventing babesia
infection.
http://www.wordsense.eu/antibabesial/
Picture of the plant