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American Journal of  
Life Science and Innovation (AJLSI)

Antioxidant Activities of  Selected Milk Teas and Fruit Teas Using 2,2-Diphenyl-1-
Picrylhydrazyl (DPPH) Assay

Perez, Jeanine Therese V.1, Perez, Lyx Jady Fy N.1, Padroncillo, Femma Joy A.1, Parreño, Kirk Andrew V.1, Policarpio, Marti 
Angela J.1, Porras, Alliah Rose B.1, Salamat, Adriel Amadei S.1, Sararaña, Rachel Joy S.1, Secapuri, Lyle Josh C.1, Tacluyan, Ann 

Marie Heather T.1, Tosoc, Daiel Dwight G.1,Vito, Joel B. Jr.1, Perez, Jose Jr. G.1,
Palmos, R.J.1, Salanio, R.1, Leong-On, Ma. Socorro G.1*

Volume 4 Issue 2, Year 2025
ISSN: 2833-1397 (Online)

DOI: https://doi.org/10.54536/ajlsi.v4i2.5050
https://journals.e-palli.com/home/index.php/ajlsi

Article Information ABSTRACT

Received: June 02, 2025

Accepted: July 08, 2025

Published: October 11, 2025

Milk teas and fruit teas are popular as a refreshment, but their percent antioxidant properties 
need to be determined. In this study, the antioxidant activities of  selected commercialized 
milk teas (Matcha, Okinawa, and Winter Melon) and fruit teas (Green Apple, Kiwi, and 
Lychee) were determined using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay in terms of  
% scavenging activity. In that assay, the plates with milk teas and fruit teas were incubated in 
the dark and covered with aluminum foil for 30 minutes at room temperature before being 
measured for absorbance at 570 nm with a microplate reader. They were kept out of  the light 
until they were evaluated for analysis. Within 30 minutes of  incubation, the discoloration 
from purple to yellow was observed. Three (3) trials with three (3) replicates were carried out 
with negative control (water with sugar and creamer for milk tea and water with sugar for fruit 
tea) and positive control (Vitamin C). Using one-way ANOVA, the Matcha (mean = 62.9, sd 
= 2.6) Okinawa (mean = 46.4, sd = 5.9), and Winter Melon (mean = 61.6, sd = 2.8) milkteas 
have anti-oxidant activities when compared to the negative control (mean = 0.0, sd = 0.0) (p < 
.05) Both Matcha and Winter Melon have the best antioxidant activities, but not comparable 
to the positive control, Vitamin C (mean = 98.7; sd = 0.62) (p < .05). While the Green apple 
fruit tea (mean = 95.5; SD = 2.4) Kiwi fruit tea (mean = 97.3, SD = 1.5), and Lychee fruit 
tea (mean = 94.4; sd = 1.3) (p > .05) have comparable antioxidant activities when compared 
to the negative control (mean sd = 0.0). Kiwi fruit tea has the best antioxidant activity and 
comparable to Vitamin C (mean = 98.7; sd = 0.62). This study scientifically established that 
both Matcha and Winter Melon commercial milk teas have antioxidant activities of  61.6 to 
62.9%. While the commercial   Kiwi, Green apple, and Lychee fruit tea ranged from 94.4 
to 97.3%. These antioxidant activities can be useful in preventing diseases brought by free 
radicals, which can be verified by conducting more studies.

Keywords

2, 2-Diphenyl-1-Picrylhydrazyl 
(DPPH) Assay, Antioxidant 
Activity, Fruit Teas, Milk Teas

1 Medical Laboratory Science, College of  Pharmacy and Medical Technology, University of  San Agustin, Iloilo City, Philippines
* Corresponding author’s e-mail: usa.edu.ph@usa.edu.ph

INTRODUCTION
The scientific community is very interested in antioxidants 
and their effects in a variety of  sectors, including food 
engineering, medicine, and pharmacy (Munteanu & 
Apetrei, 2021). According to Xiao et al. (2020), reactive 
oxygen species (ROS) have the potential to harm 
biological macromolecules, resulting in oxidative stress-
related diseases, lipid, protein, and DNA damage, as 
well as cell aging. An imbalance between the production 
and neutralization of  oxidants leads to oxidative stress. 
Oxidative stress causes a variety of  illnesses and disorders, 
including cancer, chronic renal disease, neurological 
diseases, cardiovascular diseases (CVDs), and chronic 
obstructive lung disease (Sharma et al., 2022). Oxidative 
stress could be avoided using antioxidants.
The antioxidant properties of  several types of  tea 
have been recognized. As a beverage, milk teas grew 
in popularity (Ong et al., 2021). By experimenting with 
different creamers, flavors, and sweeteners, it opened the 
door for tea lovers to enter the industry (Bastasa et al., 
2022). On the other hand, fruit teas gained popularity as 
cool alternatives. A range of  fruits, herbal infusions, and 
occasionally even boba pearls for texture are frequently 

included. These beverages transcended tradition and 
became a phenomenon in society and culture. Millennials 
and Generation Z Filipinos become much more interested 
in milk tea (National Nutrition Council, 2020).
Matcha tea, one of  the milk teas, was examined for 
its nutritional makeup and demonstrated antioxidant 
qualities (Koláčková et al., 2019). High concentrations 
of  catechins, including EGCG, which have anti-
inflammatory and antioxidant properties, were found in 
matcha. There are numerous possible health advantages 
of  matcha green tea. Compared to other green tea kinds, 
it had higher quantities of  phenolic acids, quercetin, 
rutin, theanine, and chlorophyll (Kochman et al., 2020). 
Additionally, general studies on matcha tea indicated 
that it had positive effects on anti-tumorigenesis, cardio-
metabolic health, and cognitive function (Sokary et al., 
2023). 
On the other hand, the primary constituents of  Okinawa 
milk tea are often tapioca pearls, black tea, milk, and 
kokuto, a unique Okinawan brown sugar (Saiful, 2022). 
In the Zaiyar et al. study. Using the DPPH assay, black tea 
showed high antioxidant contents in 2020. Using DPPH, 
the ABTS test (2,2′-azino-bis 3-ethylbenzothiazoline-6-



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sulfonic), and FRAP (ferric reducing antioxidant power), 
Stobiecka et al. (2022) found that cow milk, the most widely 
used type of  milk worldwide, had the lowest antioxidant 
potential when compared to other animal milk. In the 
Kinjo et al. (2019) study, kokuto, a non-centrifuged cane 
sugar, showed antioxidant and anti-stress properties. 
Tapioca pearls are made from cassava (Manihot 
esculenta). Zekarias et al. (2019) stated that it was thought 
to be a treatment for a number of  ailments, such as 
diabetes, celiac disease, bone and neurological health, 
cardiovascular disease, allergens and prostate issues, GIT 
issues, and blood pressure issues. 
Winter melon (Benincasa hispida), was evaluated as a 
nutrient-dense vegetable in Alsaadi and Abass’s (2020) 
study because it was a good source of  natural sugars, 
amino acids, organic acids, minerals, and vitamins. 
According to the pharmacological investigations, the 
plant had a wide range of  pharmacological properties, 
such as analgesic, antioxidant, anti-inflammatory, and 
central nervous system effects. antibacterial, antidiabetic, 
nephroprotective, diuretic, and hypolipidemic properties. 
Triterpenes, phenolics, sterols, glycosides, and other 
functionally significant bioactives and medicines are 
abundant in it. Using water-immersing restraint stress 
(WRS) and Indometacin [INDO]-induced gastric 
mucosal damage in rats, as well as DPPH to investigate 
the antioxidation effect and free radical scavenging of  
watermelon seed methanol extract, winter melon seed 
extract demonstrated an improved correlation between 
dependent DPPH concentration and pyloric association 
action (Purohit et al., 2019).
Fruit tea is another type of  tea offered in milk tea 
establishments. Dried fruit, flowers, or leaves are combined 
to make fruit teas. Antioxidant qualities are also present 
in the fruits used to make these teas. Green apples (Malus 
domestica) have been shown in laboratory experiments to 
have a potent antioxidant impact that lowers cholesterol, 
lipid oxidation, and the growth of  cancer cells. Quercetin, 
catechin, fluorine, and chlorogenic acid are among the 
many phytochemicals found in apples that have strong 
antioxidant properties (Aksoy & Ötleş, 2022). 
Conversely, kiwi (Actinidia deliciosa) fruits are incredibly 
rich in vitamin C and also contain a variety of  other 
nutrients, such as nutritionally significant amounts of  
dietary fiber, potassium, vitamin E, and folate, as well 
as several bioactive substances, such as a variety of  
antioxidants, phytonutrients, and enzymes, that contribute 
to their metabolic and functional advantages. A rising 
body of  evidence from human intervention research was 
drawing special attention to the role that kiwi fruit plays 
in digestive health (Richardson et al., 2018). 
Additionally, phytochemical components with anticancer, 
antibacterial, antioxidative, and antiglycating effects have 
been discovered in lychee (Litchi chinensis Sonn.). In 
addition to improving insulin resistance, lychee seed, 
lychee seed extracts, and related compounds have been 
shown to have promising antihyperglycemic properties, 
including lipid regulation, neuroprotection, anti-

inflammatory, anti-neurotoxic, and renoprotective effects 
(Zhang et al., 2021). To completely grasp the potential of  
these many teas and tastes, more research is needed, as the 
total health impact depends on individual circumstances 
and consumption habits.
The antioxidant qualities of  several plant extracts have 
been the subject of  numerous investigations. Nevertheless, 
no research had been done on the antioxidant content of  
commercial milk teas or fruit teas, which were becoming 
more and more popular. Thus, this investigation of  the 
antioxidants found in particular fruit and milk teas has 
been carried out.
This study aimed to determine the antioxidant activities 
of  the selected milk teas (matcha, okinawa, and winter 
melon) and fruit teas (green apple, kiwi, and lychee)  using 
a 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay in terms 
of  % scavenging activity. The study also determined 
the best milk tea and fruit tea that exhibited the highest 
antioxidant, and they were compared to the positive 
control used (Ascorbic Acid).
This study established the antioxidant activities of  one 
commercial brand of  milk teas (matcha, okinawa, and 
winter melon) and fruit teas (green apple, kiwi, and 
lychee). This may benefit drinkers of  these milk teas and 
fruit tea 

MATERIALS AND METHODS
Only a post-test experimental design was used in this 
study. Using the DPPH assay, various milk and fruit teas 
were screened for antioxidants. The Ethics Committee 
of  a private university in Iloilo City reviewed this study, 
which was carried out there. This research was verified 
by a licensed chemist. In order to ensure dependability, 
this study used three replicates with three trials. Every 
instrument had the proper calibration.

Acquisition of  Milk Tea and Fruit Tea
The selected milk and fruit teas were acquired from a 
local seller in Iloilo City

Dpph Assay
Three distinct flavored milk teas, three distinct flavored 
fruit teas, as well as positive and negative controls, were 
evaluated for antioxidant activity in this study. 180 μL of  
DPPH solution and 20 μL of  the commercially available 
fruit teas (green apple, kiwi, and lychee) and milk teas 
(matcha, okinawa, and winter melon) were added to each 
well of  a 96-well plate. The stock solution was created 
by dissolving 0.00591 g of  DPPH (0.3 mM) in 50 mL of  
methanol. Another stock solution was made with 50 mL 
of  methanol and 0.5 g of  ascorbic acid (10 mg/mL). 
In the wells of  the 96-well plate, 180 μL of  DPPH 
solution was mixed with 20 μL of  distilled water with 
creamer and sugar (a negative control for milk teas). To 
prevent false positive results in the antioxidant activity of  
the samples, a non-dairy creamer was employed in the 
production of  negative control for milk teas from the 
local vendor. Additionally, 180 μL of  DPPH solution and 



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Table 1: Antioxidants of  selected milk teas and fruit teas using a 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay through
% scavenging activity
Milk Teas and Fruit Teas % Scavenging Activity

Mean SD
Matcha with tea 62.9c 2.6
Okinawa with tea 46.4b 5.9
Winter melon with tea 61.6c 2.8
Negative Control for Milk Teas (Distilled water with creamer and sugar) 0.0a 0.0
Green apple with tea 95.5d 2.4
Kiwi fruit with tea 97.3d,e 1.5
Lychee with tea 94.4c 1.3
Pure Tea 91.5b 0.5
Negative control for Fruit Teas (water with sugar) 0.0a 0.0
Positive Control (Ascorbic acid) 98.7e 0.62

Note. Significant if  p-value <0.05

20 μL of  distilled water with sugar (the negative control 
for fruit teas) were poured into the 96-well plate’s wells. 
The positive controls were 180 μL of  DPPH solution and 
20 μL of  ascorbic acid (10 mg/mL) that were dispersed in 
the 96-well plate’s wells. 
A microplate reader was used to measure the absorbance 
at 570 nm after the plate had been incubated for 30 
minutes at room temperature in the dark while covered 
with aluminum foil (Suarez et al., 2021). The materials are 
kept out of  the light until they are assessed for analysis 
since DPPH is photosensitive (Lim et al., 2023). The 
purple-to-yellow staining was noticed within 30 minutes 
of  incubation. Three replicates and three trials were used 
to conduct the test. The following formula was used 
to calculate the chosen milk teas’ and fruit teas’ DPPH 
radical scavenging activity.
The plate was incubated in the dark, covered with 
aluminum foil, for 30 minutes at room temperature 
before being measured for absorbance at 570 nm 
with a microplate reader (Suarez et al., 2021). DPPH is 
photosensitive; the materials are kept out of  light until 
they are evaluated for analysis (Lim et al., 2023). Within 30 
minutes of  incubation, the discoloration of  the purple-
to-yellow color was observed. The test was carried out 
in three replicates with three trials. The DPPH radical 
scavenging activity of  the selected milk teas and fruit teas 
was determined using the formula:
% DPPH Radical Scavenging Activity = {(absorbance of  
negative control - absorbance of  sample)/ 
(absorbance of  negative control)}X100

Waste Disposal
Segregation and proper garbage disposal procedures 
were adhered to. For local rubbish collection, general 
waste was disposed away in the appropriate bins. 
Chemical wastes were divided into different containers 
and categorized based on their content. Ascorbic acid 
and DPPH were categorized as organic halogenated trash 
and were electronically and physically recorded with the 
appropriate waste codes before being submitted to the 
Pollution Control Officer.

Statistical Analysis
In this study, the mean and standard deviation were 
computed, and comparisons were performed using One-
Way Analysis of  Variance (ANOVA) followed by Tukey’s 
Honest Significant (p ≤ 0.05). 

RESULTS AND DISCUSSION
In contrast to the negative control (mean sd = 0.0), Table 1 
demonstrates that the milk teas made from matcha (mean 
= 62.9, sd = 2.6), okinawa (mean = 46.4, sd = 5.9), and 
winter melon (mean = 61.6, sd = 2.8) exhibit antioxidant 
properties. = 0. Winter melon and matcha both exhibit 
the highest levels of  antioxidant activity, however they are 
not on par with ascorbic acid, the positive control. 
In contrast to the negative control (mean sd = 0.0), the 
antioxidant activities of  the green apple fruit tea (mean = 
95.5; SD = 2.4), kiwi fruit tea (mean = 97.3, SD = 1.5), and 
lychee fruit tea (mean = 94.4; sd = 1.3) are comparable. 
The greatest antioxidant activity, on par with vitamin C, is 
seen in kiwi fruit tea (mean = 98.7; sd = 0.62). 

Discussions
There was antioxidant activity in every flavor of  milk tea, 
matcha, okinawa, winter melon, and fruit tea, as well as in 
green apple, kiwi, and lychee. This is because flavonoids, 
a subgroup of  polyphenols, are present. A wide range 
of  fruits and plant-based flavorings include them. Much 
research has focused on their ability to scavenge free 

radicals and reduce oxidative stress. Flavonoids have the 
ability to neutralize free radicals such as DPPH radicals 
and give hydrogen atoms or electrons (Kumar & Pandey, 
2013). 
Because fruits, even when powdered, are rich in 
flavonoids, fruit teas have higher antioxidant activity 
than milk teas. Our study’s results demonstrate this, since 



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while both displayed the highest antioxidant activity 
within their respective groups, kiwi fruit with tea had 
a higher antioxidant activity than matcha with tea. The 
concentration of  antioxidant chemicals during drying 
procedures may be the reason for the release of  flavonoids 
from the cell’s fruits (Nunes et al., 2016). Catechins are 
abundant in tea, which was used as one of  the flavorings.
Tea can also help reduce inflammation and cancer (Musial 
et al., 2020).The findings indicated that matcha tea had the 
highest percentage of  scavenging activity among the milk 
teas. This is consistent with a comparable study that was 
previously carried out in which matcha tea’s nutritional 
composition was examined and antioxidant qualities 
were found (Koláčková et al., 2019). High concentrations 
of  catechins, including EGCG, are found in matcha 
(Kochman et al., 2020).
Out of  all the fruit teas examined, kiwi fruit had the highest 
antioxidant activity. According to a related study about the 
fruit’s potential medical benefits, the kiwi fruit itself  has 
significant levels of  phenolic acid, with protocatechuic 
acid having the highest concentration (Zhang et al., 2021). 
One kind of  phytochemical that is frequently referred to 
as a secondary metabolite of  plants is phenolic acid, a non-
nutritive substance that may improve human cell function 
(Yoo et al., 2018). Numerous bioactivities, including anti-
tumor, anti-inflammatory, antioxidant, hypoglycemic, and 
hypolipidemic effects, are also attributed to them. The 
total antioxidant qualities of  the fruit tea are enhanced 
by the inclusion of  tea, which contains phytochemical 
qualities of  its own (Bag et al., 2022).
Ascorbic acid, which is the positive control of  this 
study, appears in the body in the form of  ascorbate, 
which is recognized for its powerful antioxidant and 
anti-inflammatory characteristics. It is clear that the 
radical anion monodehydroascorbate largely reacts with 
radicals, while ascorbic acid functions mostly as a donor 
of  single hydrogen atoms (Njus et al., 2020). All of  these 
characteristics work together to provide ascorbic acid 
with exceptional antioxidant benefits that are higher 
compared to milk teas and fruit teas, which makes it ideal 
to use as a positive control.
So far, this study is the first attempt to determine the 
antioxidant of  the commercial of  selected milk tea and 
fruit teas.
Thus, future researchers could try a different approach 
to address the limitations of  our study, which includes 
selected samples of  milk teas (matcha, okinawa, and 
winter melon) and fruit teas (green apple, kiwi, and 
lychee) along with pure tea. In addition to our analysis 
of  the data, this study was limited to using two negative 
controls specific to milk teas and fruit teas. Distilled water 
with creamer and sugar was utilized as a negative control 
for milk teas. Distilled water and sugar were utilized as 
negative controls for fruit teas.
This adds to our growing research body, further enriching 
our understanding of  antioxidant activities of  commercial 
milk teas (matcha, okinawa, and winter melon) and fruit 
teas (green apple, kiwi, and lychee).

CONCLUSION
The study demonstrated the antioxidant activity of  the 
selected milk teas (matcha, okinawa, and winter melon) 
and fruit teas (green apple, kiwi, and lychee) in terms of  
% scavenging activity. The best milk tea and fruit tea that 
exhibited antioxidant activity were matcha and winter 
melon and kiwi, for fruit tea, respectively. They can be 
useful in preventing diseases like cancer, inflammatory 
and cardiovascular disorders, and other conditions 
brought on by free radicals, subject to further studies. It 
is recommended to explore other health benefits of  milk 
teas and fruit teas aside from their antioxidant properties, 
and may also opt to use vitamin E as a positive control.

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10(15), 5195. https://doi.org/10.3390/app10155195


