Research Article

Acute toxicity of ethanolic extract of Turkcap ( arboreus Cav.) leaves Nyi Mekar Saptarini*, Resmi Mustarichie

ABSTRACT

Background: Turkcap (Malvaviscus arboreus Cav., ) leaves, empirically, are used as a hair growth stimulant in the Salawu area, Tasikmalaya district, Indonesia. Ethanolic extract of Turkcap leaves contains polyphenols, essential oils, terpenoids, saponins, and sapogenin. Aim: The purpose of this study was to determine the acute toxicity of Turkcap leaves ethanolic extracts on Swiss Webster mice. Materials and Methods: Mice were given Turkcap leaves ethanolic extract at a dose of 500, 2500, 5000, 10000, and 15000 mg/kg body weight mice. Results: The safety level of Turkcap leaves ethanolic extract was 6th category, i.e., relatively harmless, based on the classification of the OECD Guidelines for the Testing of Chemicals. Conclusion: Turkcap leaves were safe. KEY WORDS: Acute toxicity, Ethanolic extract, Non-toxic, Safe, Turkcap leaves

INTRODUCTION MATERIALS AND METHODS Turkcap (Malvaviscus arboreus Cav.) is a perennial Experimental Animals herb or shrub belongs to Malvaceae, which introduced Swiss Webster mice, 2–3 months old and weigh to several tropical and subtropical areas in Australia, 20–30 g, were obtained from the School of Pharmacy, [1] Asia, and Africa. Conventionally, this is Institute of Technology Bandung, Indonesia. Mice known to cure bronchitis, cystitis, diarrhea, dysentery, were acclimated for 1 week and their body weight fever, gastritis, hypertension, kidney diseases, liver (BW) was measured every day. Mice were said to be and gallbladder problems, stomachache, sore throat, healthy if the weight increased or decreased by no tonsillitis, and wounds.[2,3] more than 10% of the previous weight with normal Turkcap leaves are used by the people of Kampung Naga, activity during the acclimation period. This study was Salawu area, Tasikmalaya, West Java, to stimulate the approved by the Health Research Ethics Committee, hair growth empirically. Its contain asparagine, Faculty of Medicine, Universitas Padjadjaran. anthocyanin, resin, and tannin,[4] while its leaves contain Extraction saponins and sapogenin.[5] Ethanolic extract of Turkcap leaves has antifungal[6] and antibacterial activity to Turkcap leaves were collected from Manoko, Lembang Staphylococcus epidermidis and Staphylococcus district, West Java. A total of 1 kg of Turkcap leaves aureus.[7] Its ethanolic extract has anti-alopecia activity in was extracted by the maceration method through 3 × rabbits as experimental animals.[8] There is no scientific 24 h with 96% as the solvent. Every day, the solvent evidence regarding the acute toxicity of Turkcap leaves was replaced by the fresh one. All macerate was ethanolic extract. In this study, the acute toxicity assay collected and concentrated using a rotary evaporator, of Turkcap leaves ethanolic extract in mice was carried then calculated the yield.[9] out to obtain an overview of its safety for further development as a standardized herbal medicine. Acute Toxicity Assay Swiss Webster mice were divided into six groups (n Access this article online = 5), i.e., control (2% Arabic gum [PGA] suspension) and five treated groups (Turkcap leaves ethanolic extract Website: jprsolutions.info ISSN: 0975-7619 500, 2500, 5000, 10000, and 15000 mg/kg BW). Mice

Department of Pharmaceutical Analysis and Medicinal Chemistry, Faculty of Pharmacy, Universitas Padjadjaran, Jl Raya Bandung Sumedang Km 21, West Java, Indonesia

*Corresponding author: Nyi Mekar Saptarini, Department of Pharmaceutical Analysis and Medicinal Chemistry, Faculty of Pharmacy, Universitas Padjadjaran, Jl Raya Bandung Sumedang Km 21, West Java 45363, Indonesia. E-mail: [email protected]

Received on: 10-07-2019; Revised on: 14-08-2019; Accepted on: 16-09-2019

2966 Drug Invention Today | Vol 11 • Issue 11 • 2019 Nyi Mekar Saptarini and Resmi Mustarichie were given a single-dose preparation orally and observed ethanolic extract. Daily BW was observed for 14 days the toxic symptoms for 14 days; then, the mice were to determine the effect of Turkcap leaves ethanolic sacrificed. The toxic symptoms include effects on the extract on BW [Figure 1]. The mouse’s BW at the central nervous system, i.e., motoric effects, hanging, beginning of the study was in the range of 19–26 g and retention, catalepsy, sedatives, tremors, convulsions, decreased to 16–23 g after 14 days. The decreased BW straub, flexion, Hafner, pineal, and breathing; and in the control (2.4 g) was smaller than in the treated effects on the autonomic nervous system, i.e., the effects group (2.8–3.8 g), but there was not proportional to of piloerection, salivation, lacrimation, defecation, and the increased in extract dose. There was no statistically urination. The observation was carried out as before significant difference (P = 0.0033), so it was concluded treatment, then at 0.5, 1, 2, 4, and 24 h after treatment.[10] that a single dose of 500–15000 mg/kg did not affect Acute toxicity was evaluated based on the criteria of the the mouse’s BW for 14 days of observation. globally harmonized classification system for chemical substances and mixtures listed in the 13th addendum to All mice do not experience pain or death during the the OECD Guidelines for the Testing of Chemicals.[11] 14 days of observation in all treated groups [Table 1]. The administration of a single dose of Turkcap leaves Statistical Analysis ethanolic extract at the largest dose (15000 mg/kg

Data were presented as the mean ± standard deviation. BW) did not cause death until 14 days. The LD50 value One-way analysis of variance was conducted to was more than 15 g/kg BW, so the Turkcap leaves statistical analysis. Values were considered statistically ethanolic extract was categorized as having a toxicity significant at P < 0.05. level of six with a relatively harmless classification based on the classification of the OECD Guidelines RESULTS AND DISCUSSION for the Testing of Chemicals.[10,11] The yield of concentrated extract of Turkcap leaves The observation on the appearance kidney, liver, by the maceration method was 15.28%. The extract and stomach of the treated mice did not show any was dark green in color with distinctive odor and bitter abnormal changes in texture, shape, size, or color in taste. The water content of the ethanolic extract was comparison to that of the control [Table 2]. These 2.76%. This value indicated that the ethanolic extract organs were observed, due to their role in removing was good quality, due to meet the requirement, i.e., no of xenobiotics and waste products from the metabolic more than 4.1%.[12] process, so they are vulnerable to the side effects of xenobiotics. Nephrotoxicity on renal function can be The acute toxicity was evaluated through mice caused by the poisonous effects of some substances.[13] mortality and BW after 14 days, and behavior for The liver is susceptible to injury from xenobiotics, due the first 24 h after treatment with Turkcap leaves to its unique metabolism and close relationship with

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15 weight (g

body control 10 extract 500 mg/kg extract 2500 mg/kg extract 5000 mg/kg extract 10000 mg/kg 5 extract 15000 mg/kg

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Figure 1: The mouse’s body weight during observation (n=5)

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Table 1: Mice mortality during the observation Mice (n=5) Cumulative mortality (%) 2 h 4 h 24 h 48 h 72 h 7 d 14 d Control 0 0 0 0 0 0 0 Dose 500 mg/kg BW 0 0 0 0 0 0 0 Dose 2500 mg/kg BW 0 0 0 0 0 0 0 Dose 5000 mg/kg BW 0 0 0 0 0 0 0 Dose 10000 mg/kg BW 0 0 0 0 0 0 0 Dose 15000 mg/kg BW 0 0 0 0 0 0 0 BW: Body weight

Table 2: Observation on kidney, liver, and stomach Group (n=5) Kidney (g) Liver (g) Peptic ulcer Control 0.2384±0.0448 0.9156±0.2484 None Dose 500 mg/kg BW 0.2468±0.0496 0.9803±0.2047 None Dose 2500 mg/kg BW 0.2579±0.0349 0.9068±0.1097 None Dose 5000 mg/kg BW 0.2302±0.0204 0.8560±0.0346 None Dose 10000 mg/kg BW 0.2112±0.0089 0.8148±0.0736 None Dose 15000 mg/kg BW 0.2525±0.0407 0.8816±0.1276 None BW: Body weight the gastrointestinal tract. Hepatocyte and bile duct 1985;13:139-56. cell injuries lead to accumulation of bile acid in the 3. Zamora-Martínez MC, de Pascual Pola CN. Medicinal [14] used in some rural populations of oaxaca, puebla and veracruz, liver, which promote liver damage. Peptic ulcers can Mexico. J Ethnopharmacol 1992;35:229-57. caused by xenobiotics that can damage the inner lining 4. Chooi OH. Ornamental Plants: Medicinal Properties and other of stomach, the first part of the small intestine, or the Uses. Kuala Lumpur: Utusan Publication and Distributors Snd lower esophagus.[15] No lesion was noted in these organs Bhd; 2006. 5. Rakhmani SIW, Wina E, Pasaribu T.Preliminary Study on in all groups. There was no statistically significant Several Indonesian Plants as Feed Additive and Their Effect on difference in weight of the kidney (P = 0.38) and the Eimeria tenella Oocytes. Vol. 2 Proceedings of the 16th AAAP liver (P = 0.42) compared to the control. Animal Science Congress; 2014. p. 656-9. 6. Boughalleb N, Jannet HB, Debbabi N, Gannoun S. Antifungal The pharmacological screening was observed before activity of volatile components extracted from leaves, stems, and flowers of four plants growing in Tunisia. Phytopathol and at 0.5, 1, 2, 4, and 24 h after treatment. The results Mediterr 2005;44:307-12. showed that all mice still had motoric, hanging, 7. Widayanti MA. Antibacterial Activity Test of Ethanol reestablishment, flexion, Hafner, and pineal activity. Extract of (Hibiscus rosasinensis L.) and Bud All mice did not experience catalepsy, sedation, tremor, (Malvaviscus arboreus Cav.) Against Staphylococcus epidermidis and Staphylococcus aureus. Thesis. Surakarta: convulsions, straub, ptosis, writhing, piloerection, Universitas Sebelas Maret; 2016. breathing, salivation, lacrimation, defecation, and 8. Mustarichie R, Wicaksono IA, Hayati C. Anti-alopecia urination. This result showed that there is no difference characteristics of ethanol extract, n-hexane, ethyl acetate and in activity before and after treatment with the Turkcap water fractions of Malvaviscus arboreus Cav. Res J Pharm Tech 2018;11:5066-72. leaves extract compared to control. It concluded that 9. The Indonesian Ministry of Health. Indonesian Herbal Turkcap leaves ethanolic extract was safe. Pharmacopoeia. 1st ed. Jakarta: The Indonesian Ministry of Health; 2015. 10. Available from: http://www.jdih.pom.go.id/Produk/PERATUR CONCLUSION AN%20KEPALA%20BPOM/PerKBPOM%20_Nomor%20 7%20Tahun%202014%20tentang%20in%20vivo.pdf. [Last The Turkcap leaves ethanolic extract was safe in the accessed on 2019 Sep 17]. dose range of 500–15000 mg/kg BW mice. 11. Organization for Economic Cooperation and Development. OECD Guidelines for Testing of Chemicals; 2001. Available from: http://www.oecd.org/chemicalsafety/testing/ ACKNOWLEDGMENT oecdguidelinesforthetestingofchemicals.htm. [Last accessed on 2019 Jul 15]. The authors would like to thank the Academic 12. Food and Drug Supervisory Agency of Republic of Indonesia. Leadership Grant of Universitas Padjadjaran in 2019 Monograph of Indonesian Medicinal Plant Extracts. Vol. 1. for the financial support. The authors also thank Dina Jakarta: Food and Drug Supervisory Agency; 2004. Sembiring for technical assistance. 13. Galley HF. Can acute renal failure be prevented? J R Coll Surg Edinb 2000;45:44-50. 14. Jaeschke H, Gores GJ, Cederbaum AI, Hinson JA, Pessayre D, REFERENCES Lemasters JJ, et al. Mechanisms of hepatotoxicity. Toxicol Sci 2002;65:166-76. 1. Lim TK. Edible Medicinal and Non-medicinal Plants. Vol. 8. 15. Lanas A, Chan FK. Peptic ulcer disease. Lancet 2017;390:613-24. Netherlands: Springer; 2014. 2. Domínguez XA, Alcorn JB. Screening of medicinal plants used Source of support: Nil; Conflict of interest: None Declared by huastec mayans of northeastern Mexico. J Ethnopharmacol

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