Profil Metabolit Fraksi Teraktif Daun Ketapang (Terminalia catappa L.) dalam Menghambat Enzim α-Glukosidase

Abstract

Ketapang (Terminalia catappa L.) merupakan salah satu tumbuhan obat yang telah banyak digunakan dalam pengobatan tradisional karena memiliki beberapa aktivitas biologis, salah satunya sebagai antidiabetes. Tumbuhan ini sudah banyak diteliti aktivitas biologisnya akan tetapi, belum ada penelitian yang mengevaluasi pengaruh perbedaan tiap fraksi berdasarkan perbedaan kepolaran yang memiliki aktivitas inhibisi α-glukosidase. Penelitian ini bertujuan untuk mengidentifikasi profil metabolit dari fraksi teraktif pelarut yang memiliki aktivitas inhibisi α-glukosidase. Daun ketapang diekstraksi menggunakan metode maserasi dengan pelarut etanol 96% dilanjutkan dengan fraksionasi cair-cair menggunakan pelarut n-heksana dan etil asetat. Aktivitas penghambatan enzim α-glukosidase menggunakan pengujian aktivitas inhibisi enzim α-glukosidase dilakukan menggunakan microplate reader dengan substrat, yaitu p-nitrofenil-α-D-glukopiranosida (p-NPG) dan α-glukosidase yang berasal dari bakteri Bacillus stearothermophillus. Profil metabolit dari fraksi teraktif diidentifikasi menggunakan kromatografi cair spektrometri massa/spektrometri massa (LC-MS/MS). Simplisia daun ketapang dalam penelitian ini memiliki kadar air sebanyak 7,5±0,3%. Ekstrak daun ketapang memiliki rendemen tertinggi pada fraksi etanol sebanyak 10,78±3,39%, dilanjut fraksi etil asetat dan n-heksana. Hasil analisis aktivitas inhibisi α-glukosidase tertinggi ditunjukkan dari fraksi etanol dengan nilai persen inhibisi 20,17% pada uji konsentrasi ekstrak 100 μg/mL dibandingkan fraksi n-heksana dan etil asetat. Konsentrasi uji ditingkatkan hingga 5000 μg/mL dan diperoleh nilai inhibisi 67,24% dan nilai IC50 yang diperoleh pada penelitian ini 945,52 µg/mL. Meskipun nilai IC50 belum efektif, pada penelitian ini telah terkonfirmasi dugaan profil metabolit fraksi etanol menggunakan UPLC-Q-ToF-MS sebanyak 17 senyawa, yang terdiri dari golongan furan, tanin, flavonoid, benzena, terpenoid, alkaloid, lignan dan streroid. Metabolit sekunder dalam fraksi etanol ketapang yang menghambat enzim α-glukosidase, yaitu diellagilactone, ellagic acid, vitexin, 2’’-O-galloylvitexin dan astragalin 7-rhamnoside. Hasil penelitian ini menunjukkan bahwa daun ketapang kurang efektif sebagai antidiabetes melalui mekanisme penghambatan enzim. Sehingga dapat menjadi acuan untuk pencarian agen antidiabetes dari ketapang dengan menggunakan mekanisme lainnya untuk menghasilkan nilai IC50 lebih efektif.

Ketapang (Terminalia catappa L.) is a medicinal plant that has been widely used in traditional medicine because it has several biological activities, one of which is antidiabetic. This plant has been widely studied for its biological activity, however, there has been no study evaluating the effect of differences in each fraction based on differences in polarity which have α-glucosidase inhibitory activity. This study aims to identify the metabolite profile of the most active solvent fraction which has α-glucosidase inhibitory activity. Ketapang leaves were extracted using the maceration method with 96% ethanol solvent followed by liquid-liquid fractionation using n-hexane and ethyl acetate solvents. α-Glucosidase enzyme inhibition activity using α-glucosidase enzyme inhibition activity testing was carried out using a microplate reader with substrates, namely p-nitrophenyl-α-D-glucopyranosida (p-NPG) and α-glucosidase derived from the bacterium Bacillus stearothermophillus. The metabolite profile of the most active fraction was identified using liquid chromatography-mass spectrometry (LC-MS/MS). Ketapang leaf simplicia in this study had a water content of 7.5±0.3%. Ketapang leaf extract had the highest yield in the ethanol fraction of 10.78±3.39%, followed by the ethyl acetate and n-hexane fractions. The results of the analysis of the highest α-glucosidase inhibitory activity were shown from the ethanol fraction with a percent inhibition value of 20.17% in the 100 μg/mL extract concentration test compared to the n-hexane and ethyl acetate fractions. The test concentration was increased to 5000 µg/mL and an inhibition value of 67.24% was obtained and the IC50 value obtained in this study was 945.52 µg/mL. Although the IC50 value was not effective, in this study the alleged metabolite profile of the ethanol fraction was confirmed using UPLC-Q-ToF-MS of 17 compounds, consisting of furan, tannin, flavonoid, benzene, terpenoid, alkaloid, lignan and steroid groups. Secondary metabolites in the ethanol fraction of ketapang inhibit the α-glucosidase enzyme, namely diellagilactone, ellagic acid, vitexin, 2''-O-galloylvitexin, and astragalin 7-rhamnoside. The results of this study indicate that ketapang leaves are less effective as an antidiabetic through the mechanism of enzyme inhibition. So that it can be a reference for the search for antidiabetic agents from ketapang by using other mechanisms to produce more effective IC50 values.