Skripsi

Pengaruh Toksisitas in vitro β-Asaron dan Ekstrak Rimpang Jeringau (Acorus calamus L.) terhadap Larva Aedes aegypti: Mortalitas, Enzim Detoksifikasi, dan Histopatologi Midgut = Toxicity Effect of β-Asarone and Sweet Flag (Acorus calamus L.) Rhizome Extract against Aedes aegypti Larvae in vitro: Mortality, Detoxification Enzymes Activity, and Midgut Histopathological Changes.

Latar belakang: Penyakit demam berdarah dengue (DBD) masih menjadi masalah kesehatan masyarakat di Indonesia. Masalah ini diperparah dengan timbulnya resistensi vektor DBD (Aedes aegypti) terhadap insektisida sintetis secara global sehingga insektisida alternatif dari tanaman sangat dibutuhkan. Studi farmakologi menunjukkan bahwa rimpang jeringau / dringo (Acorus calamus L.) dengan kandungan senyawa fitokimia aktif β-asaron memiliki aktivitas neuroproteksi dan antioksidan sehingga banyak digunakan sebagai obat tradisional. Namun, aktivitas larvisidal dan mekanisme toksisitasnya masih belum banyak diteliti. Penelitian ini bertujuan untuk mengevaluasi aktivitas larvisidal dari β-asaron dan ekstrak rimpang jeringau terhadap larva Ae. aegypti dengan mekanisme perubahan aktivitas enzim detoksifikasi dan histopatologi midgut. Metode: Larva Ae. aegypti instar III-IV dipajankan dengan dua perlakuan, yaitu β-asaron dan ekstrak rimpang jeringau, masing-masing dalam 4 konsentrasi berbeda (0,25, 1,25, 6,25, dan 24,25 ppm) selama 24, 48, dan 72 jam. Pengamatan mortalitas dilakukan sesuai panduan WHO. Aktivitas asetilkolinesterase (AChE), glutation-S-transferase (GST), dan oksidase dianalisis dengan metode biokimia sesuai protokol CDC. Histopatologi midgut dievaluasi dengan pemeriksaan rutin histopatologi menggunakan pewarnaan H&E. Hasil: β-asaron dan ekstrak rimpang jeringau masing-masing berhasil menyebabkan mortalitas larva sebesar 66,4% dan 52% pada konsentrasi 24.25 ppm dalam waktu 72 jam. β-asaron memperlihatkan toksisitas yang lebih tinggi (LC50 = 6,768 ppm, LC90 = 14,408 ppm) dibandingkan dengan ekstrak rimpang jeringau (LC50 = 54,566 ppm, LC90 = 130,884 ppm). β-asaron dan ekstrak rimpang jeringau menghambat aktivitas AChE dan oksidase serta meningkatkan GST. Kedua perlakuan menyebabkan kerusakan masif pada seluruh bagian midgut larva Ae. aegypti: lisis food bolus (FB), kerusakan integritas membrane peritropik (PM), pecahnya lapisan epitelium (EL) pada beberapa bagian midgut, metaplasia sel lapisan epitel (EC), dan atrofi mikrovili (MV). Kesimpulan: Penelitian ini membuktikan β-asaron dan ekstrak rimpang jeringau bersifat toksik dan mampu membunuh > 50% larva Ae. aegypti pada konsentrasi rendah sekalipun (24,25 ppm). β-asaron memperlihatkan aktivitas larvisida yang lebih tinggi dibanding ekstrak rimpang jeringau dengan mekanisme menghambat enzim AChE dan oksidase serta mengakibatkan kerusakan masif pada sel dan jaringan saluran pencernaan larva Ae. aegypti.
Kata kunci: β-asaron, ekstrak rimpang jeringau/dringo (Acorus calamus L. ), larva Aedes aegypti, enzim detoksifikasi, histopatologi midgut



Introduction: Dengue hemorrhagic fever remains a prominent public health concern in Indonesia. This problem is aggravated with global insecticide resistance issue of dengue vector Aedes aegypti. Alternative vector control methods such as utilization of plant extracts are potential solution for the growing problem of dengue transmission, yet there are still not many studies regarding the matter. Sweet flag (Acorus calamus L. ) rhizome, with β-asarone as its main phytochemical content, is known to have neuroprotective and antioxidant properties in traditional medication. However, its larvicidal potential and mechanism of toxicity towards Ae. aegypti are yet to be explored. Therefore, the objective of this study is to evaluate larvicidal activity of β-asarone and sweet flag (Acorus calamus L.) rhizome extract against Ae. aegypti larvae with mechanism of detoxifying enzymes and midgut histopathology. Method: In this study, Ae. aegypti larvae instar III-IV were exposed to two different treatments, β-asarone and sweet flag rhizome extract, with concentrations of 0.25, 1.25, 6.25, and 24.25 ppm. Larval mortality was observed 24 h, 48 h, and 72 h post-treatment using WHO guideline. Acetylcholinesterase (AChE), glutathione-S-transferase (GST), dan oxidase enzyme activities were analyzed with biochemistry method using CDC guideline. Midgut histopathological changes were evaluated using H&E staining and light microscope. Result: β-asarone and sweet flag rhizome extract produced 66.4% and 52% larval mortality at 24.25 ppm respectively 72h post-treatment. β-asarone exhibited higher toxicity (LC50 = 6,768 ppm, LC90 = 14,408 ppm) than sweet flag rhizome extract LC50 = 54,566 ppm, LC90 = 130,884 ppm). Both β-asarone and sweet flag rhizome extract inhibited AChE and oxidase activities but enhanced GST activities. Both treatments caused massive damage on larval midgut histology: lysis of food bolus (FB), loss of peritrophic membrane (PM) integrity, ruptured epithelium layer (EL) on separate parts of midgut, metaplasia of epithelial cells (EC), and atrophy of microvilli (MV). Conclusion: This study proved that both β-asaron and sweet flag rhizome extract are toxic towards Ae. aegypti larvae and were able to cause > 50% larval mortality even with low concentration (24.25 ppm). β-asaron exhibited higher larvicidal activity than sweet flag rhizome extract with mechanism of AChE and oxidase enzymes inhibition and caused massive injuries on Ae. aegypti larval midgut cells and tissues.
Keywords: β-asarone, sweet flag / jeringau (Acorus calamus L. ) rhizome extract, Aedes aegypti larvae, detoxification enzymes, midgut histopathology