In Vitro Anticancer Potential of Avicennia marina Leave Extract and

Taurin on HeLa Cell Line: An Alternative Approach of Anticancer

Silvia Andriani

1,*

, Endang Linirin Widiastuti

, Evi Kurniawaty

, Suharyani

, Iffa Afiqa Khairani

and Dea Putri Andeska

Faculty of Health, Universitas Muhammadiyah Pringsewu, Lampung, Indonesia

Faculty of Math and Science, Universitas Lampung, Lampung, Indonesia

Faculty of Medicine, Universitas Lampung, Lampung, Indonesia

Faculty of Sciences, Institut Teknologi Sumatera, Lampung, Indonesia

Faculty of Biology, Universitas Gadjah Mada, Yogyakarta, Indonesia

Keywords: Avicennia Marina, Cytotoxic, Antiproliferative, Taurine.

Abstract: The pharmacological activities of Avicennia marina tree leaves, often known as api, have been scientifically

proven to include anti-inflammatory, analgesic, and toxicological effects. The Avicennia marina plant is

thought to have anticancer qualities since it contains high levels of flavonoids, tannins, saponins, and

alkaloids. Taurine's organic acids has antioxidant and anticancer properties, alongside mangrove plants.

Cancer is caused by excessive cell growth, which damages surrounding cells and tissues. Currently, cervical

cancer is the second most common cause of mortality among women globally. The MTT technique (3-(4, 5-

dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide) on HeLa cervical cancer cell cultures showed that

leaf extract and taurine effectively inhibited cancer growth. The findings indicated that the leaf extract of api-

api and taurine had cytotoxic properties, with IC50 values of 206 ppm, 122 ppm, and 603 ppm, respectively.

In contrast, the antiproliferation test demonstrated that the api-api leaf extract and taurine exhibited a longer

period of cell division compared to the control cells, with doubling times of 72 and 19 hours, respectively.

The utilization of api-api leaf extract and taurine exhibited a deleterious impact on HeLa cells as compared to

untreated cells (control cells). Treatment also inhibited cell proliferation, as shown by the longer doubling

time of treated cells compared to control cells.

1 INTRODUCTION

Cancer results from unregulated cell proliferation

in dysfunctional tissues. Cancer ranks as the second

most prevalent cause of mortality, trailing behind

cardiovascular disease. The number of cancer-related

fatalities in 2018 was roughly 9.6 million, according

to the World Health Organization's report in 2020. It

is projected that by 2030, there will be a rise of 11.4

million deaths attributed to cancer cells (Rio, S., Suci,

2017). Indonesia has the second largest number of

cervical cancer cases globally, with a fatality rate of

50% (Kementerian Kesehatan 2020). HeLa cells are

cervical cancer cells that have been infected with the

HPV-18 virus. The current cancer treatment remains

inadequate in achieving a complete cure, primarily

due to the temporary effectiveness of chemical drug-

based treatments. These medications lack selectivity

for target cells, resulting in damage to normal cells in

the body. Exploring diverse natural resources, such as

the mangrove ecosystem, can facilitate the

development of alternative medications (Albinhassan

et al., 2021).

Mangroves are resilient plants that can adjust to

shifting environments with erratic salinity and tidal

patterns. The capacity is attributed to the production

of unique chemicals by api-api leaves (Avicennia

marina) for the purpose of adaptation. The objective

of this study is to expand the availability of

phytopharmaceuticals (medicinal plants) for the

investigation of secondary metabolites from api-api

leaves (Avicennia marina) and taurine as an

anticancer agent, focusing on the components. The

efficacy of this intervention will next be assessed on

HeLa cells, namely those derived from cervical

carcinoma (Rahman, 2021).

Andriani, S., Widiastuti, E. L., Kurniawaty, E., Suharyani, , Khairani, I. A. and Andeska, D. P.

In Vitro Anticancer Potential of Avicennia marina Leave Extract and Taurin on HeLa Cell Line: An Alternative Approach of Anticancer.

DOI: 10.5220/0013110000003873

Paper published under CC license (CC BY-NC-ND 4.0)

In Proceedings of the 1st International Conference on Medical Science and Health (ICOMESH 2023), pages 25-31

ISBN: 978-989-758-740-5

2 METHOD

2.1 Extract Preparation

The Api-api leaves were acquired from the Lampung

Mangrove Center (LMC) located in Labuhan

Maringgai, East Lampung. The Api-api leaves were

rinsed with flowing water. Subsequently, the drying

process is carried out by subjecting it to an oven set

at a temperature range of 30-40˚C. Following the

drying process, it is ground into a powder known as

simplicia. Simplicia is macerated in 1:10 methanol

solvent for 24 hours, 100 grams per liter. The

macerate is further strained using filter paper. A

rotary evaporator at 50°C evaporated the extract to a

thick consistency (Nurfitri, W. A., Endang, L. W., &

Endang 2019).

2.2 Hela Cell Cuture

A 10% solution of Fetal Bovine Serum (FBS) was

prepared by adding 5 ml of the solution and 0.5 ml of

Penicillin Streptomycin. This mixture was then

combined with 50 ml of Rosewell Park Memorial

Institute medium (RPMI 1640) according to CCRC

(2009). For cell counting, 10 μl of HeLa cells were

pipetted into a well plate, 10 μl of trypan blue was

added, and the cells were counted on a

hemocytometer. A living cell appears clear, while a

dead cell is red. Hemocytometer calculations are done

in 4 rooms. The subsequent calculations pertain to the

quantity of cells to be cultivated (CCRC, 2009).

Preparation of a stock solution of 10 mg

Avicennia marina extract with 1 ml 1% DMSO for

taurine in 1 ml distilled water. The original solution

was diluted to concentrations of 125 parts per million

(ppm), 100 ppm, 75 ppm, 50 ppm, and 25 ppm

(CCRC, 2009).

2.3 Cytotoxic Test Using the MTT

Method(3-(4,5-dimetiltiazol-2-il)-2,5-

difenil tetrazoliumbromida)

For the cytotoxic test, 100 μl of cells were put to each

well, with each well containing 20,000 cells (CCRC,

2009). After a 24-hour culture period, the cells are

then washed with phosphate buffer saline (PBS).

Extract and taurine were pre-concentrated and

incubated for 24 hours in each well. The solution was

discarded and subsequently washed with a PBS

solution. Combine 10 µl MTT (3-(4,5-

dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium

bromide) with 5 mg/ml phosphate buffer saline.

Subsequently, the sample was placed in a CO

incubator and kept at a temperature of 37°C for a

duration of 2 hours. Living cells will undergo

metabolic processes to convert MTT (3-(4,5-

Dimethylthiazol-2-yl)-2,5-Diphenyl Tetrazolium

bromide) into a purple compound called formazan.

The MTT reaction was halted by adding 100 µl of

100% dimethyl sulfoxide (DMSO) stopper reagent to

each well. The absorbance was measured using an

ELISA reader at a wavelength of 550 nm (CCRC,

2013).

2.4 Antiproliferative Test with the

MTT Method (3-(4,5- dimetiltiazol-

2-il)-2,5-difenil tetrazoliumbromida)

Antiproliferative assay involved 100 µl of HeLa cells

each well, totaling 20,000 cells. The incubation

process lasted for 24 hours at a temperature of 37°C

in a CO

incubator (CCRC, 2009). After 24 hours of

growth in well plates, cells were given 100 µl of

extract and taurine at doses of 125, 100, 75, 50, and

25 ppm. The incubation process was carried out for

24, 48, or 72 hours at a temperature of 37°C in a CO

incubator. Incubation was followed by PBS rinsing of

the wells. 10 µl of MTT solution (with a

concentration of 5 mg/ml in PBS) was applied to the

wells. The wells were then incubated for an additional

2 hours at 37°C in a CO

incubator. Active cells will

transform MTT into a purple formazan compound.

The process was halted by introducing 100 µl of

100% DMSO into each well. The measurement of

absorbance for each well was conducted using an

ELISA reader at a specific wavelength of 550 nm.

Next, statistical analysis was used to compare viable

cell counts during different incubation times (CCRC,

2013).

2.5 Data Analysis

Analysing cytotoxic test data on HeLa cells involves

calculating the proportion of viable cells. The

percentage is transformed into a probit number in

order to obtain the IC50 value. Antiproliferative test

data analysis was used to estimate the doubling time

of extract and taurine at varied doses and incubation

times. The estimates were derived using linear

regression, which involved correlating the incubation

time with the logarithm of the number of viable cells.

A statistical analysis was performed using the One-

way ANOVA test with a 95% confidence level to

assess the influence of concentration on the average

number of live cells. If there are substantial disparities

between treatments, the analysis will proceed with the

Least Significant Difference (LSD) test.

ICOMESH 2023 - INTERNATIONAL CONFERENCE ON MEDICAL SCIENCE AND HEALTH

3 RESULT AND DISCUSSION

3.1 Cytotoxic Test of Avicennia Maria

Extract

Cytotoxic studies using Avicennia marina leaf extract

and taurine against HeLa cervical cancer cells yielded

a graph connecting extract concentration to cell

viability. Figure 1 and Figure 2 display these graphs.

Figure 1: Comparison of extract concentration with

percentage of Hela cell viability

As shown in Figure 1, Avicennia marina leaf extract

affects cell viability compared to the control group.

The leaf extract, when present at a concentration of

125 ppm, exhibited the lowest viability percentage of

62.4%. This value was significantly lower compared

to the other concentrations and the control group of

cells. Administration of Avicennia marina extract at a

dosage of 125 ppm led to a greater degree of cell

inhibition than the inhibitory effect of the control drug

at a concentration of 5 ppm.

Taurine has distinct properties compared to

Avicennia marina leaf extract. Evidence indicates that

taurine elicits varying reactions based on the level of

cell viability. Taurine had the lowest percentage of

viability (74.92%) and cell inhibition (24.07%) at a

dose of 100 ppm. However, the inhibition at 100 ppm

was still below the control drug's 5 and 10 ppm effects.

When compared to the drug control at a dose of 1 ppm,

the inhibition value was still greater.

All treatments have shown considerable cytotoxic

action, which reduces test cell viability relative to the

control group. Figures 3–5 show how this cytotoxic

action alters cell shape and structure. Under normal

circumstances, HeLa cells often have a polygonal

morphology and closely interact with their

extracellular matrix. However, in the event of a

disruption or the initiation of apoptosis, the cellular

morphology and structure will undergo a

transformation. Indications of these alterations

comprise a reduction in cell dimensions and cell

contraction. Hutomo et al. (2016) have provided

additional clarification for this occurrence.

Figure 2. Relationship between taurine concentration and %

cell viability

Figure 3 (A) illustrates a significantly increased

density of viable HeLa cells in comparison to

untreated control cells. Living cells have a high

density due to cells that develop without impediments

and meet nutritional needs. The cells have a flat

epithelial morphology, with a spherical and compact

nucleus placed centrally. They possess a basal lamina

that serves to bind them to the substrate, and it is

structurally intricate. Deceased HeLa cells exhibit an

uneven shape and lack luminescence (Nurani 2011).

In the pharmacological control group, cell density

was observed to be low due to apoptosis, which

resulted in cell death.

(A)

(B)

Note: Apoptosis

Figure 3: HeLa cell morphology (A) cell control and (B)

drug control with Doxorubicin.

In Vitro Anticancer Potential of Avicennia marina Leave Extract and Taurin on HeLa Cell Line: An Alternative Approach of Anticancer

(A)

(B)

Note: Apoptosis

Figure 4: HeLa Cell Morphology in Api-api Leaf Extract

Treatmen (A) 25 ppm; (B) 125 ppm.

Figure 4 indicates that 25 ppm and 125 ppm

extracts significantly change cell density. The cell

density was greater at a concentration of 25 ppm

compared to 125 ppm. The cell density is relatively

low when the api-api leaf extract is present at a

concentration of 125 ppm. Certain cells undergo

apoptosis, resulting in abnormal cellular morphology.

The application of 125 ppm api-api leaf extract was

deemed efficacious in suppressing the percentage of

cell viability in comparison to other concentrations.

These findings demonstrate the toxicity of api-api leaf

extract towards HeLa carcinoma cells. At 25 ppm,

cell density was lower than control cells, but at 125

ppm, cell density was rarely visible and color and

shape changed, indicating Hela was undergoing

apoptosis. At a dosage of 125 parts per million (ppm),

it exhibits the highest level of effectiveness in

suppressing cell development and is highly toxic to

HeLa cells.

(A)

(B)

Note: Apoptosis

Figure 5: HeLa Cell Morphology in Taurin (A) 25 ppm dan

(B) 100 ppm.

Cell density exhibited a significant decrease when

treated with a concentration of 100 ppm, in contrast

to a concentration of 25 ppm. However, viable HeLa

cells were still detectable at this concentration.

Among all the concentrations tested, taurine at 100

ppm had the best response in terms of inhibiting the

development of HeLa cells. After calculating cell

viability with the extract and taurine, the IC50 value

is calculated using the reference (CCRC, 2013).

Table 2: Test Compounds' IC50 Cytotoxic Activity against

HeLa Cervical Cancer Cells

Test

compound

Concen-

tration

(ppm)

Cell

viability

(%)

(ppm)

25 94,48 206

50 89,52

.marina 75 80,55

100 73,76

125 61,44

25 106,4

603

50 114,5

Taurin 75 91,4

100 75,9

125 109,9

1 113,38

12,35

Doxorubicin 5 73,71

10 27,46

IC50 is the concentration at which a drug inhibits

test cell growth by 50%. A lower IC50 value of the test

material indicates a higher level of toxicity and a better

potential for use as a medication. The American

National Cancer Institute (NCI) defines the cytotoxic

activity criterion for crude extracts as having an IC50

value of less than 30 µg/ml, as stated by (de Oliveira

et al. 2016).

Table 2 demonstrates that api-api leaf extract,

taurine, and doxorubicin, at different doses, result in a

ICOMESH 2023 - INTERNATIONAL CONFERENCE ON MEDICAL SCIENCE AND HEALTH

reduction in cell viability. This suggests that the test

chemical exhibits cytotoxic action against HeLa cells.

The regression calculations indicate that the IC50

value for api-api seed extract is 206 ppm, taurine is

603 ppm, and doxorubicin is 12.35 ppm. The IC50

value of the api-api leaf extract plus taurine is

significantly higher when compared to doxorubicin.

Table 3 indicates the doubling time values varied

across different treatment concentrations of api-api

leaf extract and taurine. Cell proliferation is measured

by linear regression equation slope in the doubling

time test. The control cell yielded a slope value of

0.0042. This value functions as a point of reference for

clusters of cells undergoing therapy. Treatment slope

lower than control cell slope increases doubling time.

However, the doubling time is reduced when the slope

is higher than that of the control cells (Meiyanto et al.,

2008). According to the information provided in Table

3, the slope values of all treatment cells are lower than

the slope values of the control cell. According to

Haryoto et al. (2013), the research shows that treated

HeLa cells have a longer period of time between each

cell division compared to untreated HeLa cells.

The doubling time of api-api leaf extract

increases with extract concentration. At 25 ppm, api-

api leaf extract doubles in time. Above 25 ppm,

doxorubicin did not produce the same doubling time

values as the control agent. This occurrence can be

ascribed to the negative coefficient in the linear

regression equation. A negative slope value indicates

the absence of growth as a result of cellular mortality

(Nurani 2011). The cells that were exposed to taurine

had diverse doubling time values, which were

significantly greater than those of the control cells.

This fact implies that api-api leaf extract and taurine

have the potential to function as anti-proliferative

agents in HeLa cervical cancer cells.

Table 3. Doubling Time Value in Antiproliferation Test

Test compound

Concentration

(ppm)

Equation of incubation time

line and log of cell number

Slope value

Doubling Time

value (hours)

A.Marina 25 0,0019x + 4,194

0,0019 170

50 0,0013x +4,232 0,0013 253

75 0,0012x + 4,168

0,0012 325

100 0,0007x + 4,145 0,0007 482

125 0,0002x + 4,136

0,0002 1884

25 0,0014x+4,2937

0,0014 218

50 0,001x + 4,3142

0,001 285

Taurin 75 0,0013x+4,2752

0,0013 249

100 0,0004x + 4,259 0,0004 852

125 0,0016x+4,2878

0,0016 195

Cell control 0 0,0021x + 4,306 0,0042 73,17

1 -0,0037x+4,3688

-0,0037 Tidak ada

Doxorubicin 5 -0,0187x + 4,462

-0,0187 Tidak ada

10 -0,0208x+4,2854

-0,0208 Tidak ada

Table 4: Average Number of Cells in Test Compound Treatment

Test Compound

Concentration

(ppm)

Number of livin

cells ( x 1000 Sel)

24 hours 48 hours 72 hours

Extract Avicennia

marina

25 18,9±1,48a 21,0±1,30a 26,3±0,52 a

50 18,9±0,79a 19,2 ±0,72a 23,8±0,94 a

75 16,5±0,60ab 15,6±0,92b 21,4±0,83 ab

100 15,4±0,30b 14,7±1,21b 24,1±2,88 a

125 15,0±1,34b 13,3±0,30b 18,9±0,69b

Taurin

25 21,3±0,30ab 21,8±1,42 25,7±0,60ab

50 22,9±1,48a 22,9±1,60 24,2±0,35b

75 18,3±0,66bc 22,1±0,99 23,6±1,80b

100 15,2±1,99c 21,4±3,08 19,4±1,12c

125 22,0±1,43ab 19,5±1,28 27,8±0,21a

In Vitro Anticancer Potential of Avicennia marina Leave Extract and Taurin on HeLa Cell Line: An Alternative Approach of Anticancer

The mean count of viable cells in all api-api leaf

extract groups varied significantly, according to One-

way ANOVA. Subsequent analyses utilizing the

Least Significant Difference (LSD) revealed that the

greatest quantity of viable cells was detected at a

concentration of 125 ppm following a 72-hour

incubation period. Conversely, the minimum value

was documented at a concentration of 100 ppm for a

duration of 24 hours. Different taurine dosages caused

changes in 24-hour and 72-hour viable cell numbers.

In addition, the taurine concentration reached its peak

at 125 ppm after 72 hours, whereas the lowest level

was measured at 100 ppm after 24 hours.

The anticancer action of the active chemicals in

the methanol extract of api-api seeds (Avicennia

marina) may be attributed to various probable

pathways. A. marina, a natural source abundant in

medicinal properties, has been recognized for its

potential to function as an anti-cancer agent. Api-api

seeds consist of a diverse range of chemical

constituents, including cyclic triterpenoids,

flavonoids, iridoids, naphthaquinones, polyphenols,

polysaccharides, and steroids. The majority of these

substances have demonstrated substantial anticancer

efficacy, so validating the promise of A. marina as a

natural agent in cancer treatment. The reference is

from Tian et al. (2020). Studies conducted on various

solid tumor models, both in vivo and in vitro, have

demonstrated that the activity of the treatment is

dependent on the dosage. Furthermore, the treatment

exhibits selectivity towards cancer cells, hence

minimizing the occurrence of adverse effects caused

by non-specific distribution.

Antioxidant activity depends on extract phenolic

and flavonoid concentration. Higher phenolic content

increases antioxidant activity (Gaffar et al. 2022). The

active chemical component acts as an inhibitor of

signal transduction. Growth factor-induced signal

transduction begins with external stimulation and is

subsequently detected by receptors. Signal

transduction cascades can be hindered by a variety of

test compounds, including phosphatase inhibitors and

kinase inhibitors. Flavonoids, like ATP, can interfere

with the phosphorylation process, leading to its

inhibition (Meiyanto et al., 2008). Saponin

compounds possess the capability to inhibit the

synthesis of Bcl-2. Bcl-2 is a protein with anti-

apoptotic properties, which means it prevents cell

death and promotes cell proliferation (Nitami, 2019).

Research has shown that the use of Avicennia marina

extract can trigger apoptosis in cancer cells and

enhance the expression of p53 in these cells

(Momtazi-Borojeni, 2013).

4 CONCLUSION

1. Methanol extract from api-api leaves and taurine

have a cytotoxic effect on HeLa cervical cancer

cells.

2. Methanol extract of api-api leaves and taurine

inhibit HeLa cervical cancer cell growth, as

evidenced by slower cell doubling rates compared

to controls.

3. Test compounds showed variations in cytotoxic

and antiproliferative activity. Api-api leaf extract

stands out with higher activity against HeLa

cervical cancer cells than taurine.

REFERENCES

Albinhassan, TahaniH et al. 2021. Anticancer, Anti-

Proliferative Activity of Avicennia Marina Plant

Extracts.” Journal of Cancer Research and

Therapeutics 17(4): 879.

https://journals.lww.com/10.4103/jcrt.JCRT_659_19.

CCRC (Cancer Chemoprevention Research Center). 2009.

Prosedur Kultur Sel. Fakultas Farmasi Universitas

Gadjah Mada. Yogyakarta

CCRC (Cancer Chemoprevention Research Center). 2009.

Prosedur Perhitungan Sel. Fakultas Farmasi

Universitas Gadjah Mada. Yogyakarta.

CCRC (Cancer Chemoprevention Research Center). 2009.

Prosedur Preparasi Sampel. Fakultas Farmasi

Universitas Gadjah Mada. Yogyakarta.

CCRC (Cancer Chemoprevention Research Center). 2009.

Prosedur Uji Proliferasi Sel (Doubling Time). Fakultas

Farmasi Universitas Gadjah Mada. Yogyakarta.

CCRC (Cancer Chemoprevention Research Center). 2013.

Prosedur Uji Sitotoksik. Fakultas Farmasi Universitas

Gadjah Mada. Yogyakarta.

Gaffar, Shabarni et al. 2022. “Aktivitas Antioksidan Dan

Sitotoksik Terhadap Sel Kanker HeLa Dari Ekstrak

Daun Vernonia Amygdalina (Asteraceae).” Chimica et

Natura Acta 10(1): 6–14.

https://jurnal.unpad.ac.id/jcena/article/view/36779.

Haryoto et al. 2013. “Aktivitas Sitotoksik Ekstrak Etanol

Tumbuhan Sala (Cynometra Ramiflora Linn) Terhadap

Sel HeLa, T47D Dan WiDR.” Jurnal Penelitian Saintek

18(AKTIVITAS SITOTOKSIK EKSTRAK ETANOL

TUMBUHAN SALA AKTIVITAS SITOTOKSIK

EKSTRAK ETANOL TUMBUHAN SALA

(Cynometra ramiflora Linn) TERHADAP SEL HeLa,

T47D dan WiDR): 21–28.

Hastuti, Endah Dwi, Fina Irodatul Afiyah, and Munifatul

Izzati. 2023. “Potensi Mangrove Avicennia Marina

(Forsk.) Sebagai Agen Fitoremidiasi Kadmium (Cd) Di

Tambak Dan Laut Mangunharjo, Kecamatan Tugu,

Kota Semarang.” Buletin Anatomi dan Fisiologi 8(1):

71–78.

https://ejournal2.undip.ac.id/index.php/baf/article/view

/13743.

ICOMESH 2023 - INTERNATIONAL CONFERENCE ON MEDICAL SCIENCE AND HEALTH

Kementerian Kesehatan. 2020. “Panduan Penatalaksanaan

Kanker Serviks.” Kementerian Kesehatan Republik

Indonesia. Kanker.kemkes.go.id.

Meiyanto, E., R. A. Susidarti, S. Handayani, and F Rahmi.

2008. “Ekstrak Etanolik Biji Buah Pinang (Areca

Cathecu L.) Mampu Menghambat Proliferasi Dan

Memacu Apoptosis Sel MCF-7.” Majalah Farmasi

Indonesia 19(1): 12–19.

Momtazi-Borojeni, Amir Abbas, Mandana Behbahani, and

Hojjat Sadeghi-Aliabadi. 2013. “Antiproliferative

Activity and Apoptosis Induction of Crude Extract and

Fractions of Avicennia Marina.” Iranian Journal of

Basic Medical Sciences 16(11): 1203–8.

Nurani, Laela Hayu. 2011. “Cytotoxicity, Antiproliferatif

Assays, and Expresion of P53 and BCl2 of Ethanolic

Fraction from Tea (Camellia Sinensis (L.) O.K.) Leaves

Infuse to HeLa Cells.” Majalah Obat Tradisional 16(1):

2011.

Nurfitri, W. A., Endang, L. W., dan Endang, N. C. 2019.

“Efek Ekstrak Metanol Daun (Acanthus Ilicifolius L.)

Serta Buah Jeruju Dan Taurin Dalam Menurunkan

Kadar Glukos Darah Dan Kolesterol Serta Fertilitas

Mencit Jantan (Mus Musculus L.) Yang Diinduksi

Aloksan.” In Prosiding Seminar Nasional Tumbuhan

Obat Indonesia Ke-55. Magelang,.

de Oliveira, Pollyanna Francielli et al. 2016. “Study of the

Cytotoxic Activity of Styrax Camporum Extract and Its

Chemical Markers, Egonol and Homoegonol.”

Cytotechnology 68(4): 1597–1602.

Rahman, Mahbubur. 2021. “The Effect Of Dosage Of

Mangrove Leaf Extract Avicennia Marina On The

Viability Of Hela Cells.” Journal of Stem Cell Research

and Tissue Engineering 5(1): 41.

Rio, S., Suci, E. S.T. 2017. “Persepsi Tentang Kanker

Serviks Dan Upaya Prevensinya Pada Perempuan Yang

Memiliki Keluarga Dengan Riwayat Kanker.” Jurnal

Kesehatan Reproduksi 4(3): 159–69.

Siska Febdian Nitami, Rifki Febriansah. 2019.

“Penambatan Molekular Senyawa Tangeretin Dan

Kampferol Pada Protein Antiapoptosis Bcl-XL: Studi In

Silico.” Acta Pharmaciae Indonesia 7(2): 42–50.

Tian, Shan et al. 2020. “Anti-Cancer Activity of

Biosynthesized Silver Nanoparticles Using Avicennia

Marina against A549 Lung Cancer Cells through

ROS/Mitochondrial Damages.” Saudi Journal of

Biological Sciences 27(11): 3018–24.

https://doi.org/10.1016/j.sjbs.2020.08.029.

WHO. 2020. “Cervical Cancer.” World Health

Organization. www.who.int.

In Vitro Anticancer Potential of Avicennia marina Leave Extract and Taurin on HeLa Cell Line: An Alternative Approach of Anticancer