|Year : 2020 | Volume
| Issue : 3 | Page : 148-156
Isolation of lupeol acetate from fruit peels of Artocarpus camansi
Rosnani Nasution1, Nurul Muhabbah1, Hira Helwati1, Muhammad Bahi1, Marianne Marianne2, Ulil Amna3
1 Department of Chemistry, Faculty of Mathematics and Natural Sciences, Universitas Syiah Kuala, Banda Aceh, Indonesia
2 Department of Pharmacology, Faculty of Pharmacy, Universitas Sumatera Utara, Medan, Indonesia
3 Department of Chemistry, Faculty of Engineering, Universitas Samudra, Langsa, Indonesia
|Date of Submission||16-Jan-2020|
|Date of Decision||12-Mar-2020|
|Date of Acceptance||06-Apr-2020|
|Date of Web Publication||14-Jul-2020|
Prof. Rosnani Nasution
Department of Chemistry, Faculty of Mathematics and Natural Sciences, Universitas Syiah Kuala, Banda Aceh 23111
Source of Support: None, Conflict of Interest: None
The purpose of this research is to find a lupeol acetate from Artocarpus camansi fruit peel. Ethyl acetate extract of A. camansi fruit peel was obtained by maceration process. After the process of fractionation, it results 3 subfractions (A, B, and C). The subfraction B was rechromatographed and yielded B22pure isolate. Based on data from proton nuclear magnetic resonance, Fourier transform–infrared, and mass spectrometry (MS from gas chromatography-MS), the B22isolate was suspected as lupeol acetate compound (in this study, the presence of lupeol acetate in the A. camansi fruit peel has been reported for the first time).
Keywords: Lupeol acetate, Artocarpus camansi, isolation
|How to cite this article:|
Nasution R, Muhabbah N, Helwati H, Bahi M, Marianne M, Amna U. Isolation of lupeol acetate from fruit peels of Artocarpus camansi. J Adv Pharm Technol Res 2020;11:148-56
|How to cite this URL:|
Nasution R, Muhabbah N, Helwati H, Bahi M, Marianne M, Amna U. Isolation of lupeol acetate from fruit peels of Artocarpus camansi. J Adv Pharm Technol Res [serial online] 2020 [cited 2021 May 6];11:148-56. Available from: https://www.japtr.org/text.asp?2020/11/3/148/289705
| Introduction|| |
Artocarpus camansi is known as breadnut (English), castana (Spanish), kamansi, kolo, pakau, ugod (Philippines), kelur, kulor, kulur, curor (Malaya, Java), and others. In Indonesia, the A. camansi plant is often referred as kulu or kluih. The plant of A. camansi is very similar to Artocarpus communis (breadfruit). A marked difference between the plant of A. camansi and the plant of A. communis is found in several parts, such as fruit, where A. camansi fruit has fine spines and seeds, while breadfruit (A. communis) has no seeds and does not have real fine spines on the fruit.
Research on plants of A. camansi is relatively still rarely both in its activity and chemical compounds. However, the research on A. communis, the similar plants to A. Camansi, was relatively completed. Our research reported that A. camansi plant leaf produced β-sitosterol propionate compound, which is lowering blood glucose. Further studies of n-hexane extract of A. camansi bark contained β-amyrin acetate and Cycloeugenol and cycloeucalenol acetate that actively lower blood glucose. The ethyl acetate extract of A. camansi bark contained β-sitosterol which actively lowered blood glucose. Research on dichloromethane extract of A. camansi leaf has also been done, and it contained friedelinol, squalene, β-sitosterol, stigmasterol, and phytol; while, its bark contained polyprenol, cycloartenol, and cycloartenol acetate. Although some of the compounds from A. camansi leaf and bark had been known, we need to study the fruit of A. camansi plant to obtain the chemical compounds that are likely similar to the leaves and barks.
| Subjects and Methods|| |
Plant materials and bioindicators
The sample used in this research is A. camansi fruit peel collected in 2018, from Aceh Besar, Aceh, Indonesia. The plant was identified at Herbarium Medanense, Department of Biology, Universitas Sumatera Utara, Medan.
Mass spectra were characterized using a Shimadzu gas chromatography–mass spectrometry (GC-MS) QP2010 Ultra. The 1D nuclear magnetic resonance (1 H-NMR) spectrum was measured in a CDCl3 solvent with 400 MHz JEOL spectrophotometer. The infrared (IR) spectra were recorded on PerkinElmer Fourier transform-IR spectrophotometer, using a KBr disc in the range 4000–400 cm−1. Column chromatography was performed on silica gel G60 (70–230 mesh Merck). Thin-layer chromatography (TLC) analysis was carried out using precoated silica gel G60-F254 on aluminum foil (Merck).
The method used for testing the phytochemicals can be found in phytochemical methods, a guide to modern techniques of plant analysis. Testing of triterpenoids is with the Liebermann–Burchard reagent (anhydrous acetic acid and concentrated sulfuric acid), phenolic testing with ferric chloride reagent.
Extraction of Artocarpus camansi fruit peels
As much as 30 g of ethyl acetate extract was fractionated using gravity column chromatography. The eluent system used was n-hexane:ethyl acetate with a ratio of 8:2; 7:3; and 5:5, obtained as many as 95 fractions. All fractions are monitored by TLC. The fractions that have the same mode pattern are combined to obtain three subfractions (A, B, and C). The subfractions of the extract of ethyl acetate of the A. camans i fruit peels are shown in [Table 1].
|Table 1: Subfraction of the extract of ethyl acetate of the Artocarpus camansi fruit peels|
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Based on the results of the grouping in [Table 1], further separation is focused on subfraction B because it is cleaner, then sub-fraction B as much as 2.18 g is separated by the column Chromatography with n-hexane eluent: ethyl acetate (8: 2) and produced 23 fractions which can be seen in [Figure 1].
|Figure 1: The chromatogram of fractions from subfraction B with eluent of n-hexane:ethyl acetate (8:2)|
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From chromatogram in [Figure 1], fractions 12 and 13 (subfraction B2) which are relatively pure are combined and obtained as much as 0.8 g, the B2 subfraction was rechromatographed again with eluent of n-hexane:ethyl acetate (7:3), and 23 fractions were obtained; the chromatogram of B2 subfraction is shown in [Figure 2].
|Figure 2: The chromatogram of fractions from subfraction B2 with eluent of n-hexane:ethyl acetate (7:3)|
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From chromatogram in [Figure 2], fractions 10–21 (subfraction B22) which are relatively pure are combined and tested for their purity with three different eluents, n-hexane:ethyl acetate (a) 8:2 (b) 7:3 (c) 6:4. The yield of TLC chromatogram under ultraviolet light shows one stain pattern that indicated as pure compound. The chromatogram of subfraction B22 is shown in [Figure 3].
|Figure 3: The chromatogram of isolates of B22with eluent of n-hexane:ethyl acetate (a) 6:4, (b) 7:3, and (c) 8:2|
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The B22 isolated in [Figure 3] shows a pattern of one stain. To confirm the structure, the isolate was then analyzed using1 H-NMR and IR.
| Results and Discussions|| |
Phytochemical test results
Phytochemical tests of A. camansi fruit peel showed secondary metabolites: triterpenoids, which were present in fresh samples, ethyl acetate extracts, subfractions A, B, C, and isolates of B22. The isolate of B22 showed a negative result to the ferric chloride test, and a positive result was observed to the Liebermann–Burchard test [Table 2], thus confirming that the compound is a steroid/triterpenoid type.
Characterization of the ethyl acetate extract of Artocarpus camansi fruit peels using gas chromatography–mass spectrometry
Ethyl acetate extract of A. camansi fruit peel was characterized using GC-MS. The results of GC are shown in [Figure 4], while the results of characterization with MS, and after being analyzed based on Library: NIST14.lib data on MS, the compounds are shown in [Table 3].
|Figure 4: Chromatogram of ethyl acetate extract of A. camansi fruit peel by gas chromatography|
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|Table 3: The compounds contained in the ethyl acetate extract of Artocarpus camansi fruit peel (characterization by gas chromatography-mass spectrometry)|
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Based on the chromatogram in [Figure 4] and characterization with MS, it was found that the ethyl acetate extract of A. camansi fruit peel contained 25 compounds. These compounds are shown in [Table 3].
The area and peak height in GC can be used for quantitative analysis so that the levels of each compound can be determined. Based on [Table 3], there are several compounds that have a wide area and a high peak, which makes it possible to be isolated because these compounds are obtained at high levels, such as zonarone (57.85%), and 9,19-Cyclolanost-24-en-3-ol (12.71%).
The composition of compounds in A. camansi fruit peel extract contains, among others, straight and cyclic chains. Cyclic compounds are generally as terpenoid, monoterpenoid (trans-geraniol), and triterpenoid (methyl commate C; 9,19-Cyclolanost-24-en-3-ol; 9,19-Cyclolanost-23-ene-3,25-diol, 3-acetate), and steroids (Spiro androst-5-ene-17,1'-cyclobutane-2'-one, 3-hydroxy; DELTA.5-Ergostenol (δ.5-ergostenol); trans-stigmasta-5,22-dien-3 β.-ol). For compounds with high similarities with compounds in library MS, it is very close to the actual compound.
Characterization the isolate B22(lupeol acetate)
The B22 isolates were characterized using1 H-NMR, and the results of characterization are shown in [Figure 5].
Based on the1 H-NMR spectrum, the signal showed 8 methyls at dH 2.04 (3H, s, H- 2∋), 1.74 (3H, s, H-30), 0.97 (3H, s, H-25), 0.89 (3H, s, H-28), 0.88 (3H, s, H-23), 0.85 (3H, s, H-24), 0.84 (3H, s, H-26), dan 0.83 ppm (3H, s, H-27). Doublet at 4.85 and 4.94 ppm shows C-29 atomic shifts (2H, dd, H-29a and H-29b) and methyl singlets at 1.74 for C-30, indicating that the B22 isolate is triterpenoid type. The multiplet at δH 4.31 ppm is a typical signal for proton C-3 α-orientation and dH 2.04 ppm, for C 2', indicating that the B22 isolate is a triterpenoid derived from lupeol ester.
The1 H-NMR spectrum for aliphatic protons (CH3, CH2, and CH) in triterpenoid compounds is usually seen in the chemical shift (δH) 2 ppm. Aliphatic protons are cyclic protons from the basic triterpenoid framework that are not well separated.
Based on the above reason, the isolate B22 as ester lupeol compared with lupeol acetate compound. A comparison of the chemical shift of B22 isolate compound with lupeol acetate compound is shown in [Table 4].
|Table 4: Comparison of δ proton nuclear magnetic resonance of the isolated B22 with lupeol acetate of standard|
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[Table 4] shows that there is a proton that absorbs around δH1.5 ppm like protons at H-1, H-2, H-5, H-6, and H-7 atoms. The proton at H-3 absorbs more below the field at δ = 4.13 ppm as it is influenced by the O atom having a large electronegativity. The lupeol acetate compound has a double bond on H-29, thus causing absorbing absorption at δ = 4.31 ppm and δ = 4.85 ppm. This is because the proton is attached to a carbon sp2 (C = C).
Based on the above data, the B22 isolate suspected to be lupeol acetate compounds. This is confirmed by the presence of the MS spectrum which shows a similarity in the fragmentation pattern of B22 isolates with lupeol acetate compounds. The spectrum of the mass spectrometry of the isolate B22 is shown in [Figure 6].
The mass spectrometry of the B22 spectrum shows the peaks of m/z: 453, 408, and 393 and peaks at 218, 203, and 189, as well as the peak base at m/e 43 (100%), showing fragmentation patterns similar to compounds of lupeol acetate.
The presence of acetate groups in the lupeol compound was amplified by the presence of carbonyl uptake (C = O) at the wave number at 1716 cm−1 and the absorption of CO at wave number of 1248 cm-1 at IR [Figure 7].
The ester of the compound was studied has a band at 1640 cm−1 indicated C = C vibrations, (C29), and C-O band found in the fingerprint area 1110–1300 cm−1 so that the B22 isolates were predicted as lupeol acetate [Figure 8].
From the results of GC-MS, the abundant compound in ethyl acetate extract was 9,19-Cyclolanost-24-en-3-ol (area: 12.71%; retention time: 57,748 min; and similarity: 84%), and this compound is a triterpenoid. Besides the presence of zonarone (retention time of 63,371 min and area of 57.85%; similarity of 67%), its structure as the initial skeletal precursor of lupan (the lupeol acetate has lupan framework). Of the two compounds present in GC-MS, this has a relatively large similarity, as a framework supporting the skeleton of lupan so that the compound B22 is lupeol acetate which is an abundant compound in the ethyl acetate extract of A. camansi fruit peel.
A literature shows that secondary metabolites isolated from plants A. camansi are groups of terpenoids or steroid, which have one-way pathways, So it is very relevant to find terpenoid compounds in the peel of A. camansi fruit because, in other parts of this plant, steroids have been found. Both of these secondary metabolites have a one-way biosynthetic pathway. The structure of the lupeol acetate is included in the skeleton lupan-type triterpene, and the presence of lupeol is reported for the first time in the A. camansi peels.
Based on the literature, it is known that lupeol acetate has many benefits, including antinociceptive and anti-inflammatory.,, Lupeol acetate also has an antimicrobial, anti-inflammatory, antimalarial, and antituberculosis activity.,, Lupeol compounds can reduce the activity of α-amylase and can inhibit Tyrosine phosphatase 1B. In addition, lupeol also showed moderate inhibitory activity against glutathione S-transferase and acetylcholinesterase. Umbelliferone and lupeol studies (100 and 200 mg/kg BW) of banana flowers decreased fasting hyperglycemia activity in diabetic rats given for 4 weeks.
| Conclusions|| |
Based on the phytochemical test, secondary metabolite in ethyl acetate extract of A. camansi fruit peel is triterpenoid. Characterization by GC-MS and ethyl acetate extract of A. camansi fruit contained 25 chemical compounds with the main compounds of zonaron (retention time: 63,371 min; area of 57.85%; and similarity: 67%) and 9,19-Cyclolanost-24-en-3-ol (area: 12.71%; retention time: 57,748 min, 84%: similarity). The results of B subfraction separation by column chromatography obtained isolate B22, based on spectra analysis of1 H-NMR, IR, and MS, the isolate B22 was suspected as a lupeol acetate compound. In this study, the presence of lupeol acetate (B22) has been reported for the first time.
The author would like to thank Universitas Syiah Kuala, Ministry of Research, Technology and Higher Education in accordance with the Letter of Agreement for Research of Professor Candidates.
Financial support and sponsorship
This stud was financially supported by the Ministry of Research, Technology and Higher Education.
Conflicts of interest
There are no conflicts of interest.
| References|| |
Ragone D. Artocarpus camansi
(Breadnut) ver 2.1. In: Elevitch CR, ed. Species Profiles for Pacific Island Agroforestry. Permanent Agriculture Resources(PAR). Holualoa, Hawaii. 2006.p. 1-11.
Nasution R, Barus T, Nasution P, Saidi N. Isolation and structure elucidation of steroid from leaves of Artocarpus camansi
(Kulu) as antidiabetic. Int J Pharm Tech Res 2014;6:1279-85.
Nasution R, Marianne, Nur H. ß-Amyrin acetate of ethyl acetate extract of the bark plant Artocarpus camansi
and its antidiabetic activity. Der Pharma Chemica 2015;7:71-8.
Nasution R, Marianne, Bahi M, Saidi Junita N. β-Sitosterol from Bark of Artocarpus camansi
and its Antidiabetic Activity. Proceedings of the 5th
Annual International Conference, Syiah Kuala University (AIC Unsyiah), In Conjunction with the 8th
International Conference of Chemical Engineering on Science and Applications (ChESA). Banda Aceh. Indonesia; 9-11 September, 2015.
Tsai PW, Cruz DC, Shen CC, Chiou CT, Raga YC. Chemical constituents of Artocarpus camansi
. Pharm J 2013;5:80-2.
Harborne JB. Phytochemical Methods: A Guide to Modern Techniques of Plant Analysis. 3rd
ed. London: Chapman and Hall; 1998. p. 302.
Sparkman OD, Penton Z, Fulton G. Gas chromatography and mass spectrometry: A practical guide, Elsevier 2011.
Muktar B, Bello IA, Sallau MS. Isolation, characterization and antimicrobial study of lupeol acetate from the root bark of Fig-Mulberry Sycamore (Ficus sycomorus LINN). J Appl Sci Environ Manage 2018;22:1129-33.
Hingkua SS, Julaeha E, Kurnia D. Terpenoid compound from the stem of Mangrove plant Avicennia marina against human pathogenic Bacteria staphylococcus aureus and Pseudomonas aeruginosa. Proceedings of MatricesFor IITTEP – ICoMaNSEd 2015 ISBN: 978-602-74204-0-3.
Nejma BA, Besbes M, Guerineau V, Touboul D, Jannet HB, M'hamed AH. Isolation and structure elucidation of acetylcholinesterase lipophilic lupeol derivatives inhibitors from the latex of the Tunisian Periploca laevigata
. Arabian J Chem 2017;10:S2767-72.
Fessenden RJ, Fessenden JS, Organic chemistry, 2nd
ed, Williard Grant Press, Boston, massachusets 1982.
Jamal AK, Yaacob WA, Din LB. A chemical study on Phyllanthus reticulatus
. J Phys Sci 2008:19:45-50.
Perfeito JP, Santos ML, Lopez KS, Paula JE, Silveira D. Characterization and biological properties of Pouteria torta
extracts: A preliminary study. Rev Bras Farmacogn 2005;15:183-86.
Chen YF, Ching C, Wu TS, Wu CR, Hsieh WT, Tsai HY. Balanophora spicata and lupeol acetate possess antinociceptive and anti-inflammatory activities in vivo
and in vitro
. Hindawi Publishing CorporationEvidence-Based Complementary and Alternative MedicineVolume 2012, Article ID 371273, 10 pages. doi:10.1155/2012/371273.
Patocka J. Biollogically active pentacyclic triterpenes and their current medicine signification. J Appl Biomed 2003;1:7-12.
Tahany MA, Hegazy AK, Sayed AM, Kabiel HF, El-Alfy T, El-Komy SM. Study on combined antimicrobial activity of some biologically active constituents from wild Moringa peregrina
Forssk. J Yeast Fungal Res 2010;1:015-24.
Subhadhirasakul S, Takayama H, Kitajima FM, Aimi NF. Triterpenoids from Thai medicinal plant, Willughbeia firma.
Nat Med 2000;54:155-57.
Prachayasittikul S, Saraban P, Cherdtrakulkiat R, Ruchirawat S, Prachayasittikul V. New bioactive triterpenoids and antimalarial activity of Diospyros rubra
Lec. EXCLI J 2010;9:1-10.
Lucetti DL, Lucetti EC, Bandeira MA, Veras HN, Silva AH, Leal LK, et al
. Anti-inflammatory effects and possible mechanism of action of lupeol acetate isolated from Himatanthus drasticus
(Mart.) Plumel. J Inflamm (Lond) 2010;7:60.
Ali H, Houghton PJ, Soumyanath A. Alpha-amylase inhibitory activity of some Malaysian plants used to treat diabetes; with particular reference to Phyllanthus amarus
. J Ethnopharmacol 2006;107:449-55.
Na M, Kim BY, Osada H, Ahn JS. Inhibition of protein tyrosine phosphatase 1B by lupeol and lupenone isolated from Sorbus commixta
. J Enzyme Inhib Med Chem 2009;24:1056-9.
Ramu R, Shirahatti PS, Swamy N, Zameer F, Lakkappa Dhananjaya B, Prasad NM. Assessment of in vivo
antidiabetic properties of umbelliferone and lupeol constituents of banana (Musa
sp. var. Nanjangud Rasa Bale) flower in hyperglycaemic rodent model. PLoS One 2016;11:e0151135.
[Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6], [Figure 7], [Figure 8]
[Table 1], [Table 2], [Table 3], [Table 4]