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AIP Conference Proceedings 2092, 030024 (2019); https://doi.org/10.1063/1.5096728 2092, 030024
© 2019 Author(s).
Evaluation of encapsulated microparticles ofGarcinia mangostana L. extracts on markerSGOT, SGPT, BUN and creatinine serum ofBALB/c miceCite as: AIP Conference Proceedings 2092, 030024 (2019); https://doi.org/10.1063/1.5096728Published Online: 09 April 2019
Amirah Deandra Diba, Trienty Batari G. Purba, M. Wien Wienarno, Elsa A. Krisanti, Kamarza Mulia, and SitiFarida
Evaluation of Encapsulated Microparticles of Garcinia
mangostana L. Extracts on Marker SGOT, SGPT, BUN and
Creatinine Serum of BALB/c Mice
Amirah Deandra Diba1, Trienty Batari G Purba1, M Wien Wienarno2, Elsa A
Krisanti3, Kamarza Mulia3, and Siti Farida4,a)
1Undergraduate student, Faculty of Medicine, Universitas Indonesia, Jl. Salemba Raya No. 6, Central Jakarta
10430 Indonesia 2National Institute of Health Research and Development, Indonesian Ministry of Health, Jl. Percetakan Negara
No.29, Central Jakarta 10560, Indonesia 3Department of Chemical Engineering, Faculty of Engineering, Universitas Indonesia, Kampus UI Depok, West
Java, 16424 Indonesia 4Department of Medical Pharmacy, Faculty of Medicine, Universitas Indonesia, Jl. Salemba Raya No. 6, Central
Jakarta 10430 Indonesia
f)Corresponding author: siti.farida@ui.ac.id
Abstract. Garcinia mangostana L. (mangosteen) pericarp containing α-mangostin is predicted to have potential as an
anticancer because it can induce apoptosis and has high antioxidant content. Moreover, the preliminary study showed that
this extract has high antioxidant activity and cytotoxicity based on the brine shrimp lethality test. This indicates that
mangosteen has potential as chemotherapeutic agents or additional therapy in cancers such as colorectal cancer, which the
available management is not as effective. To improve its efficacy and bioavailability in the colon as a targeted area, ethyl
acetate fraction of mangosteen extract was then formulated into a controlled release encapsulated microparticles by
chitosan-alginate material. It is intended that the active ingredient to be released after reaching the colon. In this study,
toxicity of mangosteen was investigated because there is still no toxicity data of encapsulated microparticles of the ethyl
acetate fraction of mangosteen extract on function of mice's kidneys (BUN/serum creatinine) and liver (SGOT/SGPT).
Twenty female BALB/c mice were divided into 4 groups. Three groups were given a single dose of 2, 3, and 5 g/kg body
weight (BW) mangosteen via intragastric feeding tube and one control group without mangosteen. The findings show that
marker of BUN, serum creatinine, SGPT, and SGOT are not significantly different from those of the control in the
administration of microparticles ethyl acetate fraction of mangosteen extract at a single dose of 2 and 3 g/kgBW, however,
significant increases of BUN and SGOT levels were found in single dose of 5 g/kgBW. The results indicate that
administration encapsulated of microparticles ethyl acetate fraction of G. mangostana L. extract at single dose of 2 and 3
g/kgBW cause no toxicity on liver and kidneys function, but the single dose of 5 g/kgBW. cause toxicity to the liver and
kidneys. Further studies on histopathology examinations of liver and kidneys are necessary to ensure its safety level.
Keywords: acute toxicity, control released, ethyl acetate fractionation, Garcinia mangostana L., microencapsulation
INTRODUCTION
Colorectal cancer is a cancer that is formed from the tissue of colon which consists of colon and or rectum [1]. In
Indonesia, colorectal cancer is ranked the third cancer with estimated mortality rate 9.5% among other cancer cases
[2-5]. The risk factors of colorectal cancer are genetic, limited activities, obesity, low fiber and high fat diet, low
vitamin D, smoking, alcohol consumption, and drugs [1,6,7]. With the westernization in Indonesia that affect life style
and diet, the incidence of colorectal cancer in Indonesia has increased to 12.8 per 100.000 [6-9]. However, the
3rd Biomedical Engineering’s Recent Progress in Biomaterials, Drugs Development, and Medical DevicesAIP Conf. Proc. 2092, 030024-1–030024-4; https://doi.org/10.1063/1.5096728
Published by AIP Publishing. 978-0-7354-1822-6/$30.00
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treatment of choice, surgery, is usually not affordable and gives many side effects. Other treatment options for
colorectal cancer are chemotherapy and radiotherapy [5]. However, these therapies have some major side effects and
limitations for Indonesia, and the costs are also high.
Knowing the limitations of these treatments, alternative treatment that costs less and has effective effects for
colorectal cancer is needed. One of the potential substances is from a fruit plant, Garcinia mangostana L. Many studies
found that the extract of G. mangostana L. is beneficial as an anticancer agent for cancers, including colorectal cancer
[10-15]. As colorectal cancer is located in the gastrointestinal tract, the medicine should be encapsulated with polymer
substances so the drugs can be slowly released in the colon. Chitosan-alginate encapsulation microparticles are proven
to be the basic formulation for controlled drug release [11].
In this research, the extract of G. mangostana L. which is encapsulated by chitosan-alginate microparticles was
given to 4 groups of BALB/c mice to find the oral acute toxicity of the drug with different doses on marker function
of mice’s kidneys using BUN (Blood Urea Nitrogen) and serum creatinine and liver using SGOT (serum glutamic
oxaloacetic transaminase) and SGPT (serum glutamic pyruvate transaminase). The objectives are to evaluate the effect
of this drug to SGOT, SGPT, BUN, and creatinine serum levels so it can be used as a consideration for a safe anti-
colon cancer agent. Elevation of marker levels of function kidneys and liver indicate toxicity and reduction of their
function.
MATERIALS AND METHODS
Plant material and extract preparation
The extract of chitosan-alginate microparticles encapsulated mangosteen was obtained as a result of a research that
was done by Faculty of Engineering, Universitas Indonesia. The pericarp of mangosteen was obtained in 2014 from
Solo and was labeled as Garcinia mangostana Linn. by Herbarium Bogoriense, Research Center for Biotechnology-
Indonesian Institute of Sciences (LIPI). Standard α-mangostin (98%) was from China and the chitosan (medical grade;
deacetilation degree of 93.6%; viscosity of 23.3 cp) was obtained from Biotech Surindo, Indonesia. Other materials
such as calcium chloride and sodium alginate were obtained from Merck. While Sodium tripolyphosphate (food grade)
was obtained from Brataco Chemical, Indonesia. The extract that was already within ethyl acetate fraction was then
suspended in Department of Medical Pharmacy, Faculty of Medicine Universitas Indonesia.
The dry pericarp of mangosteen was extracted using a modified procedure stated by Jung, et al [12]. Subsequently,
the extract was macerated using ethanol 96% for seven days applying mangosteen powder to ethanol ratio of 1:3 (m/v)
and periodic stirring was done to the mixture. The macerated mixture was then filtrated and evaporated under a reduced
pressure to be crude ethanolic extract. Afterwards, the mixture was fractionated using a mixture of ethyl acetate and
water with 1:1 volume ratio.The evaporation of this ethyl acetate fraction under reduced pressure generated mangostin
powder/paste [11]. The mangostin powder was then emulsified using gum arabic to form a more soluble drug to be
administrated.
Experimental Animals
Female BALB/c mice were obtained from Central of Health Research Development, Ministry of Health of the
Republic of Indonesia and acclimatized to the laboratory conditions. Twenty 6-8 weeks, nulliparous and not pregnant
mice, 25-30 g of body weight with 20% maximum of weight distribution were prepared. All mice were housed 5 per
cage in a room maintained at 25°C, 50 ± 10% relative humidity and a 12-hour-light/12-hour-dark cycle throughout
the experiment. All mice had free access to water and food before the beginning the experiment. All of the procedures
in this study have been approved by the Health Research Ethic Committee Faculty of Medicine, Universitas Indonesia
(Approval no. 0430/UN2F1/ETIK/2018).
Acute Oral Toxicity Study and Dose Administration
The mice were randomly divided into 4 groups (n=5) i.e. three experimental groups and one control group, with
each group contains five mice [16]. The extract in doses of 2, 3 and 5 g/kgBW. Mice are fasted for 3-4 hours (except
water) before treatment. The administration of mangosteen extract dissolved in a solvent (distilled water and gummi
arabicum), and it was administered through intragastric in a single oral dose by gavages using a feeding needle with
maximum volume 1 ml/100 g mice. The control group received an equal volume of solvent. The animals were then
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weighed and given a single dose of mangosteen extract based on their body weight. In the circumstance where it is
not possible to administer a dose with a single feeding, the extract of mangosteen can be administered several times
within 24 hours. The feeds may be given 1-2 hours after treatment. The mice were maintained within 14 days
observations. After administration, the mice were observed during the first day and each of the following 14 days to
look for any signs and symptoms of toxicity and death. After it was done, all of the remaining mice were sacrificed to
see the clinical biochemistry in their blood specimen.
Biochemical Analysis
After the 14 days observations, the remining mice were anesthetized for blood collection. Blood samples were
centrifuged and the supernatant were stored. Examination of clinical biochemistry including SGOT and SGPT levels
are used to determine the liver function, while BUN and serum creatinine concentration levels are used to determine
the kidney function. The blood samples was examined using Spectrophotometry.
Statistical analysis
All data was processed using Stat view 5.0 software. Statistical significance of the difference of clinical
biochemistry in the four groups was tested using one-way analysis of variance (ANOVA). Significance of difference
was considered at p <0.05. The difference among groups was analyzed using Fisher’s Paired Least Significant
Difference (PLSD) post hoc. If the P value is less than 0.05, the result will be considered significant.
RESULTS AND DISCUSSION
Table 1 shows that all mice groups given single dose of mangosteen extract had higher BUN levels than the control
group, but a significant increase occurred at the administration of 5 g/kgBW. Moreover, a significant increase of
SGOT levels was found in the 5 g/kgBW group. On the other hand, serum cretainine and SGPT did not show any
significant dose-related effects. This finding indicates that the administration of single dose encapsulated
microparticles of the ethyl acetate fraction of mangosteen extract at 2 and 3 g/kgBW did not affect the marker of
kidneys and liver function, but a single dose of 5 g/kgBW caused toxicity and reduction of the function of the liver
and kidneys of mice. These findings were different from study by Kosem who conducted repeated administration of
crude methanolic extract during a 14-day study showed renal and hepatic cell injury, resulting a significant increase
of BUN in ≥500 mg/kg BW dose, SGPT in ≥250 mg/kg BW dose, and SGOT in ≥200 mg/kg BW dose.
CONCLUSION
From this research, it can be concluded that G. mangostana L. extract with ethyl acetate fraction encapsulated by
chitosan-alginate microparticles is safe seen from kidneys and liver function at a dose until 3 g/kg BW. Therefore, this
extract is potential to be a safe anti-colon cancer drug. However, further research on histopathological examination
needs to be done to ensure the safety of this drug.
Dose administered
(g/kgBW)
Marker
BUN
Serum
creatinine SGPT SGOT
Control 13.03 ± 1.88 22.36 ± 0.58 4.60 ± 2.74 7.22 ± 1.14
2 15.84 ± 2.17 21.88 ± 0.21 5.29 ± 0.24 8.73 ± 1.78
3 15.44 ± 3.79 21.80 ± 0.20 5.24 ± 0.71 8.73 ± 1.81
5 19.45 ± 5.79* 21.92 ± 0.71 4.42 ± 1.66 12.22 ± 1.43*
TABLE 1. The marker level of function of kidneys and liver after administration of the G. mangostana L. extract
*p < 0.05 vs. control group (0 g/kg BW)
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ACKNOWLEDGMENTS
The research was funded by Hibah Publikasi Terindeks International untuk Tugas Akhir Mahasiswa UI (PITTA)
2018. This study, which is an experimental study using mice, has been approved by the ethical commissary of Faculty
of Medicine of University of Indonesia with serial number of 0430/UN2.F1/ETIK/2018. The authors would like to
appreciate also dr. Siti Farida, M.Kes for the support, assistance, and advices during this research. The funding of this
research was supported by the PITTA Grand from Universitas Indonesia with number of contract
2130/UN2.R3.1/HKP.05.00/2018.
REFERENCES
1. Scholefield JH, Eng C. Colorectal cancer: Diagnosis and clinical management (Wiley Blackwell, Hoboken,
2014).
2. Ferlay J, Soerjomataram I, Ervik M, Dikshit R, Eser S, Mathers C, Rebelo M, Parkin DM, Forman D, Bray F.
GLOBOCAN 2012: estimated cancer incidence, mortality and prevalence worldwide in 2012. Geneva World
Health Organization, International Agency for Research on Cancer no 11. (2012).
3. Haggar FA, Boushey RP. Colorectal cancer epidemiology: incidence, mortality, survival, and risk factors. Clin
Colon Rectal Surg. 22 (4), 191 (2009).
4. Centers for Disease Control and Prevention. Leading cancer cases and deaths, male and female. CDC; 2018 p. 1
(2014).
5. Komite Penanggulangan Kanker Nasional, Pedoman nasional pelayanan kedokteran kanker kolorektal
(Kementrian Kesehatan Republik Indonesia, Jakarta, 2017).
6. Tamakoshi K, Tokudome S, Kuriki K, Takekuma K, Toyoshima H. Gan To Kagaku Ryoho. Cancer &
chemotherapy. 28 (2), 146-150 (2001).
7. Schmoll HJ, Van Cutsem E, Stein A, Valentini V, Glimelius B, Haustermans K, Nordlinger B, Van de Velde CJ,
Balmana J, Regula J, Nagtegaal ID. ESMO Consensus Guidelines for management of patients with colon and
rectal cancer. A personalized approach to clinical decision making. Annals of oncology. 23 (10), 2479-2516
(2012).
8. Van Cutsem E, Nordlinger B, Cervantes A, ESMO Guidelines Working Group. Advanced colorectal cancer:
ESMO clinical practice guidelines for treatment. Annals of oncology. 21 (suppl 5), 93-97 (2012).
9. Kosem N, Ichikawa K, Utsumi H, Moongkarndi P. In vivo toxicity and antitumor activity of mangosteen extract.
J Nat Med. 67 (2), 255-263 (2013).
10. Watanopokasin R, Jarinthanan F, Jerusalmi A, Suksamrarn S, Nakamura Y, Sukseree S, Uthaisang-
Tanethpongtamb W, Ratananukul P, Sano T. Potential of xanthones from tropical fruit mangosteen as anti-cancer
agents: caspase-dependent apoptosis induction in vitro and mice. Appl Biochem Biotechnol. 162 (4), 1080-1094
(2010).
11. Mulia K, Halimah N, Krisanti E. “Effect of alginate composition on profile release and characteristics of chitosan-
alginate microparticles loaded with mangosteen extract” in AIP Conference Proceedings 2017 Mar 17. 1823 (1),
p. 020010. (2017)
12. Jung HA, Su BN, Keller WJ, Mehta RG, Kinghorn AD. Antioxidant xanthones from the pericarp of Garcinia
mangostana. J Agric Food Chem. 54 (6), 2077-2082 (2006).
13. Palakawong C, Sophonodora P, Pisuchpen S, Phongpaichit S. Antioxidant and antimicrobial activities of crude
extracts from mangosteen (Garcinia mangostana L.) parts and some essential oils. IFRJ 17 (3), 583-589 (2010).
14. Pedraza-Chaverri J, Cárdenas-Rodriguez N, Orozco-Ibarra M, Pérez-Rojas JM. Medicinal properties of
mangosteen (Garcinia mangostana). Food and chemical toxicology . Food Chem Toxicol. 46 (10), 3227-3239
(2008).
15. Putri IP. Effectivity of xanthone of mangosteen (Garcinia mangostana L.) rind as anticancer. J Majority. 4 (1),
33-38 (2015).
16. Badan Pengawas Obat dan Makanan Republik Indonesia. Peraturan kepala badan pengawas obat dan makanan
republik indonesia nomor 7 tahun 2014 tentang pedoman uji toksisitas nonklinik secara in vivo (Badan Pengawas
Obat dan Makanan Republik Indonesia, Jakarta, 2014).
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