BAKTERI ENDOFIT PADA PURWOCENG (Pimpinella alpina Kds.) PENGHASIL KUMARIN

TRI WIDAYAT, Prof. Dra. A. Endang S. Soetarto, M.Sc., Ph.D.

2011 | Tesis | S2 Biologi

Purwoceng (Pimpinella alpina Kds.) merupakan tanaman obat langka penghasil metabolit sekunder yang berkhasiat sebagai afrodisiak. Tanaman yang mengandung kumarin tersebut berinteraksi dengan bakteri endofit. Penelitian ini bertujuan untuk mengetahui lokasi bakteri endofit pada organ tanaman purwoceng, mendapatkan bakteri tersebut dan menganalisis kemampuan tumbuh bakteri endofit dalam medium yang mengandung kumarin dan kemampuan bakteri tersebut dalam menghasilkan kumarin. Penelitian diawali dengan pengambilan sampel purwoceng di dataran tinggi Dieng kemudian dilanjutkan deteksi bakteri endofit in planta menggunakan methylene blue. Isolasi bakteri endofit dilakukan secara langsung dari organ tanaman yang mengandung bakteri. Seleksi bakteri endofit penghasil kumarin dilakukan berdasarkan kemampuan bertahan hidup pada medium cair ammonium salt sugar (ASS) yang mengandung infusa herba purwoceng. Kemampuan bakteri menghasilkan kumarin diuji melalui percobaan kultivasi isolat bakteri terpilih pada medium yang sama. Tiap interval waktu tertentu pertumbuhan dimonitor secara spektrofotometri dan plating pada medium nutrient agar (NA). Data yang diperoleh digunakan untuk menentukan waktu generasi (g) dan kecepatan pertumbuhan spesifik (µ). Kadar kumarin dalam kultur dideteksi menggunakan teknik kromatografi lapis tipis (KLT). Isolat bakteri yang menghasilkan kumarin diidentifikasi berdasarkan metode standar. Hasil pengamatan mikroskopi perbesaran 400x menunjukkan bahwa bakteri endofit terdapat mengelompok pada ruang antar sel jaringan parenkim korteks tanaman purwoceng. Sembilan isolat bakteri endofit berhasil diisolasi dari akar dan daun tanaman purwoceng secara langsung. Semua isolat mampu bertahan hidup pada medium basal yang diberi infusa herba purwoceng dengan waktu generasi (g) 2,7-5,07 jam dan kecepatan pertumbuhan spesifik (µ) 0,14-0,26/jam. Kultivasi isolat terpilih menunjukkan bahwa BAP5 menghasilkan senyawa dengan Rf 0,27 pada KLT yang diduga sebagai turunan kumarin. Bakteri BAP5 mampu tumbuh dengan kadar kumarin 1072 arbitrary unit (AU) dalam medium. Kumarin dalam kadar rendah diperlukan oleh isolat BAP5 untuk tumbuh dan menghasilkan senyawa turunan kumarin yang lain. Penelitian disimpulkan bahwa bakteri endofit terdapat pada semua organ tanaman purwoceng. Bakteri endofit dapat ditumbuhkan di laboratorium dan menghasilkan turunan kumarin.

ndophytic Purwoceng (Pimpinella alpina Kds.) is a an endangered medicinal plant which produces secondary metabolites with efficacy as an aphrodisiac. The plant contains coumarin and interacts with endophytic bacteria. The aims of this research were to determine the location of endophytic bacteria in the purwoceng organs, to isolate the bacteria and to analyze the growth of endophytic bacteria which were grown on coumarin containing medium and the ability of bacteria to produce coumarin. The research was begun with the purwoceng sampling in the Dieng plateau, then proceeded by in planta detection using methylene blue. E bacteria were isolated directly from plant organs containing bacteria. The selection of coumarin-producing endophytic bacteria was based on the ability to survive in ammonium salt sugar (ASS) liquid medium that contained purwoceng herbal infusion. The ability of bacteria to produce coumarin was assayed through experimental cultivation of selected bacterial isolate on the same medium. In every specific time interval the growth was monitored spectrophotometrically and plated on nutrient agar (NA). The obtained data were used to determine the generation time (g) and specific growth rate (μ). Coumarin level in the culture was detected by using thin layer chromatography (TLC). Bacterial isolate that produced coumarin was identified by standard methods. Observation under microscope with 400x magnification showed that endophytic bacteria resided and agglomerated in the intercellular space of parenchymal tissue of purwoceng cortex. Nine endophytic bacterial isolates were isolated from the roots and leaves of purwoceng directly. All isolates were able to survive on basal medium containing purwoceng herbal infusion with generation time (g) 2.7 to 5.07 hours and the specific growth rate (μ) 0.14 to 0.26/hour. Cultivation of selected isolate showed that BAP5 produced certain compound that was suspected as coumarin derivative with Rf 0.27 on TLC. BAP5 bacteria were able to grow on the medium with coumarin level of 1072 arbitrary units (AU). Coumarin in low level was required by the BAP5 isolate to grow and produce other coumarin derivative. From these results it could be concluded that endophytic bacteria were present in all purwoceng organs. Endophytic bacteria could be grown in the laboratory and produced coumarin derivative.

Kata Kunci : bakteri endofit, purwoceng (Pimpinella alpina Kds.), kumarin.