Biology, Medicine, & Natural Product Chemistry ISSN 2089-6514 (paper)

Volume 12, Number 2, October 2023 | Pages: 601-606 | DOI: 10.14421/biomedich.2023.122.601-606 ISSN 2540-9328 (online)

In Vitro Anti-inflammatory Activity of Bamboo Tali Leaf

(Gigantochloa apus) Ethanol Extract

Nurul Hidayah, Mutista Hafsah*

Chemistry Department, Faculty of Science and Technology, UIN Walisongo,
Jl. Walisongo No.3-5, Tambakaji, Kec. Ngaliyan, Kota Semarang, Jawa Tengah 50185, Tel. (024) 7604554, Indonesia.

Corresponding author*
mutista.hafshah@walisongo.ac.id

Manuscript received: 21 July, 2023. Revision accepted: 10 August, 2023. Published: 07 November, 2023.

Abstract

Inflammation is a physiological process that serves as a defense mechanism for the body against foreign substances, bacteria, or irritants.
Inflammation can be cured with anti-inflammatory drugs. One of the plants that has the potential to be an anti-inflammatory agent is the
bamboo leaf. This research aims to analyze the content of secondary metabolites, determine the inhibition value and IC50 value of the anti-
inflammatory activity of the ethanol extract of bamboo tali leaves. Bamboo tali leaves were macerated using 96% ethanol and subjected to
phytochemical screening. The extract was then tested for anti-inflammatory activity in vitro with the Bovine Serum Albumin (BSA) protein
denaturation inhibition method. Bamboo tali leaf ethanol extract contains flavonoids, alkaloids, saponins, and phenols. The anti-inflammatory
activity of the ethanol extract of bamboo tali leaves with concentrations of 28, 42, 56, 70, and 84 ppm had an inhibition percentage value of
23.14 ± 0.008%; 34.30 0.026%; 54.51 0.060%; 69.07 ± 0.006%; and 87.02 ± 0.021% with an IC50 value of 52.991 ppm. These results
indicate that the ethanol extract of bamboo tali leaves has the potential to be an anti-inflammatory with a strong IC50 value below 100 ppm.

Keywords: Anti-inflammatory; Bamboo tali; Phytochemicals; Protein denaturation; Secondary Metabolites.

INTRODUCTION including alkaloids, saponins, tannins, phenols,

flavonoids, triterpenoids, steroids, and glycosides were
A physiological process called inflammation includes a discovered (Setiawan & Yusransyah, 2018). secondary
living thing's immune system. Inflammation serves as a metabolites with anti-inflammatory effects, including as
physiological defense against certain diseases, such as phenols, saponins, and tannins. These metabolites work
cancer, by shielding the organism from microbial to suppress the production of inflammatory mediators
infection (Bouyahya at al., 2022). There are several signs and increase free radical production (Armadany at al.,
of inflammation in the body, including redness, fever, 2020). Consequently, the bamboo tali's leaves can be
edema, and loss of tissue function (Novika at al., 2021). utilized as an anti-inflammatory medication.
Inflammation can be treated with synthetic medications Through the approach of preventing protein
such ibuprofen, aspirin, diclofenac sodium, and denaturation, anti-inflammatory activity can be tested.
celecoxib. Long-term use of these medications may This is due to the fact that protein denaturation is one of
result in gastrointestinal adverse effects (Izzany at al., the causes of tissue inflammation (Farida, Rahmat, &
2018). Research into anti-inflammatory medications Amanda, 2018). The protein bovine serum albumin
manufactured from natural substances, sometimes known (BSA) was utilized in this study. The way the sample
as herbal medicines, is therefore required. solution interacts with BSA demonstrates the theory
In Indonesia, herbal remedies use is one way to utilize behind protein denaturation inhibition. Free radical
of the country's natural resources. One of the bamboo generation causes body proteins to be more susceptible to
species found in Indonesia, bamboo tali, has been denaturation. The outcome of the inflammatory process
extensively used in the medical field. One component of is the release of inflammatory mediators (Shalihah at al.,
rope bamboo, called bamboo tali leaves, is not 2022).
appropriate for use because it is regarded as waste for the
environment (Fitriani, 2018). Extraction of the bioactive
chemicals present in bamboo leaves is one strategy that MATERIALS AND METHODS
can be used to make use of them (Romansyah at al., Materials and Tools
2019). In bamboo tali (Gigantochloa apus) leaf extract The tools used in this study were volume pipettes, stir
macerated with 70% ethanol, bioactive substances bars, spatulas, thermometers, test tubes, test tube racks,
602 Biology, Medicine, & Natural Product Chemistry 12 (2), 2023: 601-606

Erlenmeyer (IWAKI), porcelain cups, volumetric flasks This procedure is continued until the combination turns a
(100 mL, 100 mL, 25 mL, and 10 mL) (IWAKI), vials, light shade of clear, provided that it is filtered and the
cuvettes, dark glass bottles, pH meter, analytical balance solvent is changed once every 24 hours in order to
(AND), blender (COSMOS), Centrifuge (PLC SERIES), acquire the ethanol extract and determine the yield level.
1 set of distillation apparatus, one set of vacuum rotary 𝑬𝒙𝒕𝒓𝒂𝒙𝒕 𝒘𝒆𝒊𝒈𝒉𝒕
evaporator, and UV-Vis spectrophotometry instrument %𝑌𝑖𝑒𝑙𝑑 = 100% (Eq. 2)
𝑺𝒊𝒎𝒑𝒍𝒊𝒄𝒊𝒂 𝒘𝒆𝒊𝒈𝒉𝒕
(ORION AQUAMATE 8000).
The juvenile leaves of Gigantochloa apus, a species
of rope bamboo, were the materials employed in this Phytochemical Screening Test
investigation. In this study, additives with proanalytical Flavonoid (Romansyah at al., 2019)
quality were employed. 96% technical ethanol, distilled Two milliliters of the sample in total, heated for 5
water, Mg, HCl, Wagner reagent, FeCl3, bovine serum minutes. The material is heated before being combined
albumin (BSA), tris buffer saline (TBS), NaCl (Merck), with five drops of concentrated HCl and 0.1 g of Mg
and CH3COOH are the other ingredients. metal. Positive reaction is indicated by a yellowish-
orange to reddish solution.
Methods
Making powder from bamboo tali leaf (Wahyuni at al., Alkaloid (Wahid & Safwan, 2020)
Wagner's reagent was added in three separate drops to
2020)
Bamboo leaves are removed, cleaned under running the 2 mL extract sample. If a brown hue shows up, the
sample is positive for alkaloids.
water, and allowed to air dry for about five days. Using a
blender, the little bits of dried leaves were mixed into a Saponin (Wahid & Safwan, 2020)
powder. A test tube containing 3 mL of the extract sample was
then filled with 10 mL of hot water, cooled, and
vigorously shaken for 10 seconds. One drop of pure HCl
was then added after that. The positive sample includes
saponins if the foam that forms does not vanish.

Tannin
The extract sample, a total of 3 mL, was heated for 5
minutes before being combined with 5 drops of 1%
FeCl3. According to Romansyah et al. (2019), the
development of dark blue or blackish green is a sign that
tannins are present.

Phenol (Putri, 2018)

Two milliliters of the extract sample were mixed with
Figure 1. Bamboo tali leaf.
five drops of 1% FeCl3. A positive sample that contains
phenol is indicated if a solid black or bluish green color
Calculation of Bamboo Tali Leaf Powder's Moisture is produced.
Content (Fitri & Anita, 2014)
Two grams of powdered rope bamboo leaf were baked An anti-inflammatory effect Test
for 30 minutes at 105 degrees. It was then placed in a TBS (Tris Buffer Saline) production (Barr at al., 2006)
desiccator to chill for 15 minutes. After cooling, the 900 mL of distilled water was added after 8.7 g of NaCl
resulting weight is weighed to ascertain the water and 1.21 g of tris base were weighed in total. After then,
content. the pH was maintained at 6.2–6.5 by adding glacial
CH3COOH. A 1000 mL volumetric flask is filled with a
𝑏−(𝑐−𝑎) stable NaCl and tris base mixture, and the pH is then
% 𝑀𝑜𝑖𝑠𝑡𝑢𝑟𝑒 𝑐𝑜𝑛𝑡𝑒𝑛𝑡 = 𝑏
(Eq. 1)
raised to the desired level using distilled water.
Information: BSA (bovine serum albumin) production 0.2%
a : weight of the cup (g) (Williams at al., 2008)
b : sample weight (g) 0.2 g of BSA was added to a 100 mL volumetric flask,
c : cup weight + sample (g) and the volume was subsequently raised to 100 mL with
TBS solution.
Producing Bamboo tali Leaf Extract from Tali leaf
(Wahyuni at al., 2020) Positive Controls Creation (Rusli & Setiawan, 2020)
Bamboo tali leaf powder was macerated with 96% Two milliliters of 0.2% BSA solution were combined
ethanol that has undergone a 1:5 (w/v) distillation ratio. with 1.5 mL of TBS and 1.5 mL of distilled water to
 Hidayah & Hafsah – In Vitro Anti-inflammatory Activity of Bamboo Tali Leaf … 603

create a 5 mL volumetric flask. Without any additional Developing the primary and test solutions (Muliati,
processing, the mixture's absorbance value was instantly 2014)
determined using a UV-Vis spectrophotometer at the In order to create mother liquor, a 10 mL volumetric
optimized wavelength of 289 nm. Duplicate runs of the flask containing up to 70 mg of an ethanol extract of
experiment were conducted. bamboo tali leaves was combined with 10 mL of ethanol.
This resulted in a concentration of 7,000 ppm. Following
Negative Control Creation (Rusli & Setiawan, 2020) that, the mother liquor concentration was adjusted to 840,
Two milliliters of 0.2% BSA solution, 1.5 mL TBS, and 700, 560, 420, and 280 ppm.
enough distilled water to reach the boundary mark should
be added to the 5 mL volumetric flask. The mixture was Testing for Anti-Inflammatory (Novika at al., 2021)
cooked for 30 minutes at 37°C and then for another 10 To a mixture of 2 mL of 0.2% BSA solution, 1.5 mL of
minutes at 72°C. Following the completion of the heating TBS, and distilled water, a total of 0.5 mL of the test
phase, the mixture was centrifuged at 6,000 rpm for 10 solution at various concentrations was added.
minutes while maintaining a temperature of 32 °C. At a Concentration changes from the solution were 84, 70, 56,
wavelength of 289 nm, the absorbance value was 42, and 28 ppm. The temperature of each concentrated
measured twice (duplo). A 5 mL volumetric flask was solution was raised for 10 minutes at 72°C after 30
filled to the boundary mark with distilled water, 1.5 mL minutes at 37°C. Following the completion of the heating
TBS, and 2 mL of 0.2% BSA solution. The mixture was operation, the mixture's temperature was maintained at
cooked for 30 minutes at 37°C and then for another 10 32°C for 30 minutes before being centrifuged at 6,000
minutes at 72°C. Following the completion of the heating rpm for 10 minutes. A UV-Vis spectrophotometer was
phase, the mixture was centrifuged at 6,000 rpm for 10 used to measure the absorbance, which was repeated
minutes while maintaining a temperature of 32 °C. At a twice (duplo). (Shalihah at al., 2022). Once the
wavelength of 289 nm, the absorbance value is measured absorbance value has been read, each solution's percent
twice. inhibition value has to be calculated. If a natural
product's % inhibition value is greater than 20%, it is
considered to have anti-inflammatory activity.

𝑡𝑒𝑠𝑡 𝑎𝑏𝑠𝑜𝑟𝑏𝑎𝑛𝑐𝑒−𝑛𝑒𝑔𝑎𝑡𝑖𝑣𝑒 𝑐𝑜𝑛𝑡𝑟𝑜𝑙 𝑎𝑏𝑠𝑜𝑟𝑏𝑎𝑛𝑐𝑒

%𝐼𝑛ℎ𝑖𝑏𝑖𝑡𝑖𝑜𝑛 = × 100% (Eq. 3)
𝑝𝑜𝑠𝑖𝑡𝑖𝑣𝑒 𝑐𝑜𝑛𝑡𝑟𝑜𝑙 𝑎𝑏𝑠𝑜𝑟𝑏𝑎𝑛𝑐𝑒−𝑛𝑒𝑔𝑎𝑡𝑖𝑣𝑒 𝑐𝑜𝑛𝑡𝑟𝑜𝑙 𝑎𝑏𝑠𝑜𝑟𝑏𝑎𝑛𝑐𝑒

RESULTS AND DISCUSSION ethanol has a high boiling point (79.37°C), using a
temperature of 60°C during the evaporation process aims
Sample preparation to prevent harming the bioactive components. A small
Using a blender, the dried bamboo leaves were chopped amount of oil and a dark, viscous extract are produced by
and ground into a fine, light-weight powder that was the evaporation process. 12.34 g of the extracted viscous
light green in color. Bamboo tali leaf powder has a material had a yield value of 12.34%. To estimate the
moisture level of 3.42%. 10% of the sample must be value of bioactive components that can be extracted
water-free to maintain sample quality (Wijaya & using solvents, the percent yield is calculated (Dewatisari
Noviana, 2022). These facts demonstrate that the sample at al., 2018).
of rope bamboo leaves satisfies the water content
requirements. The sample's water content has a Phytochemical ScreeningTest
significant impact on its quality and shelf life, making it As can be seen in table 1, the phytochemical screening of
more susceptible to damage and microbial contamination the ethanol extract of bamboo leaves revealed the
(Hutauruk at al., 2014). presence of flavonoids, alkaloids, saponins, and phenols
(Table. 1).
Obtainable Yield
At a temperature of 60 °C and a rotational speed of 70
rpm, the macerated filtrate was concentrated. Because

Table 1. Results of Phytochemical Testing.

Bioactive Substances Description of Phytochemical Test Results Information

Flavonoid orange-yellow +
Alkaloid Brown with developed black clumps +
Saponin Formed Stable Foam +
Tannin Brown -
Phenol Solid Black +
Information: (+) : Has bioactive ingredients, (-) : Does not include any bioactive ingredients
604 Biology, Medicine, & Natural Product Chemistry 12 (2), 2023: 601-606

Inflammatory-Reduction Capacity denaturation, which will allow the phospholipase enzyme

The BSA protein denaturation inhibition method was to convert phospholipids into arachidonic acid.
used to assess the ethanol extract of bamboo tali leaves Cyclooxygenase (COOX) enzymes will then break down
for anti-inflammatory effects. BSA is a protein derived arachidonic acid to create prostaglandins, which can
from cow albumin. BSA works in enzymatic processes to activate pain receptors and increase capillary
stabilize proteins. BSA has a pH between 6.0 and 8.5 permeability (Fitriyani & Fatahillah, 2022). Secondary
(Borzova at al., 2016). Because BSA is simple to metabolites have the ability to limit the synthesis of
produce, easily soluble, affordable, and minimizes the prostaglandins. Bamboo tali leaf extract has anti-
use of live specimens in the drug development process, it inflammatory properties due to the presence of secondary
is used in this study (Novika at al., 2021). One of the metabolites including flavonoids, alkaloids, saponins,
reasons why the cell tissues of the body get inflamed is and phenols.
protein denaturation. Protein denaturation is the process Through the action of the phospholipase enzyme,
by which pressure or chemicals from the outside result in flavonoids prevent the synthesis of prostaglandins
an alteration of the tertiary and secondary structure of (Bustanul & Sanusi, 2018). Because of their capacity to
proteins or nucleic acids. The approach of protein suppress the creation of cytokines that promote
denaturation inhibition was chosen because it is non- inflammation, alkaloids have the potential to be anti-
acidic, inexpensive, and simple to use (Aditya at al., inflammatory medications. Saponins' anti-inflammatory
2015). mechanism, which involves preventing the breakdown of
glucocorticoids and the release of inflammatory
Table 2. results of Bamboo tali leaf extract's % BSA inhibition. mediators (Mohammed at al., 2014). Phenol is a
No Concentration (ppm) %Inhibition secondary metabolite with OH groups, which can protect
1. 28 23,14±0,008
membranes, prevent the production of inflammatory
2. 42 34,30±0,026 mediators, and deactivate free radicals (Novika at al.,
3. 56 54,51±0,060 2021).
4 70 69,07±0,006 The anti-inflammatory efficacy of the dicoflenac
5. 84 87,02±0,021 sodium medication is shown in Table 3. According to the
data in table 3, all of the variants employed satisfied the
requirements for % protein denaturation inhibition.
Table 3. Results of % inhibition of BSA by diclofenac sodium. Diclofenac sodium was chosen since it is one of the most
effective anti-inflammatory medications (Novika at al.,
No Concentration (ppm) %Inhibition 2021). Diclofenac sodium is a heteroarylacetate salt of
1. 10 26,38%±0,006 sodium (2,6-dichlorophene) (Fitri Yani & Reynaldi,
2. 20 35,77%±0,001
2021). Figure 2 depicts the structure of diclofenac
3. 30 44,54%±0,000
4 40 56,31%±0,001 sodium. Diclofenac sodium can inhibit the
5. 50 66,73%±0,001 cyclooxygenase COX-1 and COX-2 enzymes, lowering
prostaglandin, prostacyclin, and thromboxane synthesis
(Altman at al., 2015).
When BSA protein is heated, protein denaturation
Cl
takes place (Fitriyani & Fatahillah, 2022). The protein's
molecules travel quicker and sustain damage as a result
of the rising temperature's increased kinetic energy NH
(Shalihah at al., 2022). As a buffer solution, BSA was
Cl O
dissolved in Tris Buffer Saline (TBS). BSA's ideal pH is
7, so TBS solution was selected because it has a pH of
6.25 to 6.5 (Abidin at al., 2020). Percent inhibition is
ONa
used to express the protein denaturation activity. When Figure 2. Diclofenac Sodium Structure.
the proportion of inhibition is greater than 20%, anti-
inflammatory action is present (Farida, Rahmat, & Widia
Amanda, 2018). Tables 2 and 3 show the findings of After acquiring data on the percentage inhibition
research on the anti-inflammatory effects of diclofenac value, the IC50 value is calculated. The anti-inflammatory
sodium and an ethanol extract of bamboo tali leaves. test results are interpreted using the IC50 value. The IC50
The percentage of inhibition for each extract value denotes the concentration of the test substance that
concentration is shown in Table 2. The % inhibition can reduce inflammation by 50%. As illustrated in
values for concentrations of 28, 42, 56, 70, and 84 are Figures 3 and 4, the obtained % inhibition value data is
known to already meet the criterion for % inhibition of entered into the linear regression equation with the
protein denaturation, namely > 20%, based on Table 2. extract concentration (ppm) as the x-axis and the %
The cell membrane will be harmed by protein inhibition value as the y-axis.
 Hidayah & Hafsah – In Vitro Anti-inflammatory Activity of Bamboo Tali Leaf … 605

inflammatory properties. Furthermore, dicloofenac

100,00% sodium has a faster absorption and pain alleviation than
y = 0,0116x - 0,1147
other nonsteroidal anti-inflammatory medicines like
80,00% ibuprofen or naproxen (Altman at al., 2015).
R² = 0,9939
Inhibition (%)

60,00%

40,00% CONCLUSIONS
20,00% Bamboo tali leaves (Gigantochloa apus) ethanol extract
0,00% contains secondary metabolites including flavonoids,
0 20 40 60 80 100 alkaloids, saponins, and phenols. The anti-inflammatory
Concentration (ppm) activity of the ethanol extract of bamboo tali leaves
(Gigantochloa apus) at concentrations of 28, 42, 56, 70,
Figure 3. Linear Regression Curve of Anti-inflammatory Activity of and 84 ppm was 23.14%, 34.48%, 55.12%, 69.30%, and
Bamboo Tali Leaf Ethanol Extract. 87.41%, respectively, with an IC50 value of 52.991 ppm.

Competing Interests: The authors declare that there are

80,00%
y = 0,0101x + 0,1549 no competing interests.
60,00% R² = 0,9974
Inhibition (%)

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