OPTIMASI TEKNIK WESTERN BLOT UNTUK DETEKSI EKSPRESI PROTEIN TANAMAN PADI (Oryza sativa L.)

Optimization of Western Blot Technique for Protein Expression of Rice Plant (Oryza sativa L.) Western blot (WB) technique has been widely used for analyzing protein expression and for identifying specific proteins derived from animals, plants, and microorganisms. During the use of WB, especially in agricultural studies, some difficulties are encountered such as unclear or unspecific protein bands, the presence of bubbles, and the absence of protein bands on membrane. This study aims to determine the WB conditions appropriate for the protein expression of rice plants (Oryza sativa L.). Protein from rice plant was extracted and the obtained protein lysate was then used for proteomic analysis using western blot with β-actin antibody. Our experiment showed that some optimized parameters like blocking buffers, the concentration of primary antibody and the ratio of secondary antibody determined the clarity of the results. β-actin was used as internal control that measured the success of the WB technique. Results showed that lysis process was important in determining good WB results in addition to the optimal blocking solution using a BSA of 0.2%, a primary antibody concentration of 1 μg mL–1, and a secondary antibody of 1:10,000. Optimizing techniques during extraction, incubation, and documentation facilitated good WB results.Keywords: β-actin; optimization; protein; rice plant; western blotABSTRAKTeknik western blot (WB) telah banyak digunakan untuk analisis ekspresi protein dan mengidentifikasi protein spesifik dari hewan, tumbuhan dan mikroorganisme. Dalam implementasi teknik WB, khususnya studi dalam bidang pertanian, beberapa kesulitan ditemui seperti pita protein tidak jelas, tidak spesifik, adanya gelembung, hingga tidak munculnya pita protein pada membran. Penelitian ini bertujuan untuk mengetahui kondisi WB yang tepat untuk deteksi protein tanaman padi (Oryza sativa L.). Protein tanaman padi diekstraksi, kemudian lysate protein yang didapat dianalisis dengan metode westernblot menggunakan antibody β-actin. Penelitian kami menunjukkan bahwa beberapa parameter yang dioptimasi seperti larutan blocking, konsentrasi antibodi primer dan rasio antibodi sekunder akan menentukan hasil yang jelas. β-actin digunakan sebagai kontrol internal yang menjadi tolok ukur keberhasilan teknik WB. Hasil menunjukkan bahwa proses lisis menjadi hal penting dalam menentukan hasil WB yang baik disamping larutan blocking yang optimal menggunakan BSA 0,2%, konsentrasi antibodi primer 1 µg mL–1 dan antibodi sekunder 1:10.000. Mengoptimalkan teknik selama ekstraksi, inkubasi dan dokumentasi membantu mendapatkan hasil WB yang baik

Senyawa Daun Rasamala (Altingia excelsa Nornha) sebagai Penghambat Proliferasi Sel Kanker Lidah Manusia In Vitro

Tongue cancer is a common malignant in the oral cavity and to now medically succes its no good so often treated of life. The aims of study were to identify the secondary matabolities of Rasamala leaves, to examine the compound of Rasamala leaves toward SP-C1 cell growth inhibition. True experimental laboratories design using SP- C1 oral tongue cancer cell line was used. The ethyl acetate extract was separated their secondary metabolites by various chromatographic techniques guided by antiproliferasi activity to yield five compounds, as 3,4-dihydroxy benzoic acid (1), gallic acid (2), and apigenin (3). Compounds 1-3 were tested on their antiproliferative activity against cancer cells tongue SP-C1 and showed IC 50 values of  100,  100, and  100 μg/mL, respectively. Data analysis using ANACOVA assay, Tukey HSD with the level of significance α=0.05. In conclusion, compound from Rasamala leaves have a strong anticancer activity in human oral tongue cancer SP-C1 through inhibit cell prolieration.

Flavonoids from Limau Peel (Citrus amblycarpa (Hassk.) Ochse) and Their Antioxidant Activity

Citrus amblycarpa (Hassk.) Ochse is an endemic Indonesian plant from West Java. This study aims to determine the chemical structure of the flavonoid compounds of C. amblycarpa peel ethanol extract and their antioxidant activity. Three flavonoid compounds, namely 5-hydroxy-3',4',6,7,8-pentamethoxyflavone (1), 8-hydroxy-3,3',4',5,6,7-hexamethoxyflavone (2), and 3',4',5,6,7,8-hexamethoxyflavone (3), were isolated for the first time from the ethanol extract of C. amblycarpa peel. Their chemical structure was determined by spectroscopic methods (MS, IR, 1H-NMR, 13C-NMR, and DEPT) and compared with previous reported spectral data. Furthermore, these compounds were evaluated for their antioxidant activity using the DPPH method. The results showed that 8-hydroxy-3,3',4',5,6,7-hexamethoxyflavone (2) has the highest antioxidant activity with IC50 value of 121.09 ± 0.24 ppm

Insecticidal Bufadienolides from The Leaves of Kalanchoe daigremontiana (Crassulaceae)

Kalanchoe is the biggest genera of Crassulaceae family and distributed in tropical and subtropical regions. This genera is found to be a rich source of biologically active natural products such as triterpenes, flavonoids and steroids. As a part of our continuing search for novel insecticidal compounds from Indonesian Kalanchoe plants, we examined Kalanchoe daigremontiana collected from Bandung region, West Java, Indonesia. The methanolic extract of the dried leaves of K. daigremontiana was concentrated and extracted with methylene chloride The methylene chloride extract exhibited an insecticidal activity toward silkworms. The methylene chloride extract was partitioned between n-hexane and methanol containing 10% water. The active lower layer was extracted with ethyl acetate. By using the insecticidal activity to follow the separations, the ethyl acetate fraction was separated by combination of column chromatography on Kieselgel 60 and ODS to afford two insecticidal bufadienolides 1 and 2. The structures of these compounds were elucidated based on spectroscopic analysis (UV, IR, NMR, 2D-NMR) and comparison with those related data previously reported. In this paper, the isolation, structural elucidation, and insecticidal activities against the third instar larvae of silkworm will be described

ISOLASI DAN MOLECULAR DOCKING SENYAWA 6,7-DIHIDRO-17-HIDROKSIERISOTRIN DARI DAUN DADAP BELENDUNG (Erythrina poeppigiana) TERHADAP AKTIVITAS SITOTOKSIK ANTIKANKER PAYUDARA MCF-7

Erythrina (Leguminose) memiliki jumlah alkaloid paling banyak jika dibandingkan dengan senyawa metabolit sekunder lainnya, sehingga tumbuhan Erythrina dikenal dengan sebutan alkaloid eritrina. Erythrina yang paling banyak tumbuh di Indonesia adalah E. poeppigiana. Tujuan penelitian ini adalah untuk mengungkapkan senyawa aktif antikanker payudara MCF-7 secara in silico yang terdapat di dalam daun E. poeppigiana. Alkaloid dipisahkan dan dimurnikan melalui tahapan ekstraksi, fraksinasi, pemisahanan dan pemurnian yang dipandu dengan uji Dragendorff, senyawa alkaloid yang diperoleh diuji antikanker secara in silico terhadap reseptor EGFR2 yang bekerja pada sel kanker payudara MCF-7 menggunakan program ArgusLab dengan pembanding canertinib dan parameter yang diukur adalah energi bebas Gibbs ikatan (ΔG) dan konstanta inhibisi (Ki). Hasil dari penelitian ini diperoleh senyawa alkaloid golongan isokuinolin yakni 6,7-dihidro-17-hidroksierisotrin. Senyawa 6,7-dihidro-17-hidroksierisotrin diprediksi mempunyai aktivitas sitotoksik terhadap sel kanker payudara MCF-7 dengan nilai ΔG (Kcal/mol) dan Ki (nM) masing-masing sebesar -8,11457 dan 1,12.10-6. Sehingga dapat disimpulkan bahwa tumbuhan E. poeppigiana mempunyai potensi sebagai bahan dasar obat herbal antikanker payudara

IDENTIFICATION OF FLAVONOIDS FROM ACALYPHA INDICA L. (EUPHORBIACEAE) AS CASPASE-3 ACTIVATORS USING MOLECULAR DOCKING AND MOLECULAR DYNAMICS

Objective: The purpose of this study was to determine the structural-based molecular interactions between flavonoids contained in Acalypha indica L. and caspase-3 by molecular docking and molecular dynamics (MD) simulations. Methods: In a computer simulation, ten flavonoids contained in A. indica L. were evaluated for caspase-3 using the X-ray crystal structure of human caspase-3 (PDB ID 1NME). The AutoDock 4.2 software was used to study molecular docking, and MD simulations were done with GROMACS v2018. Results: The results of molecular docking identified the top four compounds, namely nicotiflorin, naringenin, hesperetin, and kaempferol, with docking scores of-6.81,-6.45,-6.33, and-6.10 kcal/mol, respectively. According to the MD simulation results, nicotiflorin was most effective in stabilizing the complex with caspase-3, with a total energy (ΔGbind, MM-PBSA) of-96.315 kcal/mol. Conclusion: This study showed that nicotiflorin was the flavonoid in A. indica L. that activated caspase-3 the best

Lignan (+)-Piperitol-γ,γ-Dimethylallylether from Stem Bark of Zanthoxylum rhetsa (Roxb.) DC (Rutaceae)

Lignans are a group of phenylpropanoid dimers in which the phenylpropane units are linked by their propyl side chains' central carbon (C8). Lignans have various biological activities, including antiviral, anticancer, cancer preventive, and cytotoxic. Zanthoxylum rhetsa (Roxb.) DC is a tree with prickly branchlets belonging to the Zanthoxylum genus, commonly known as panggal buaya in Indonesia. Asian tribes have used this plant as traditional medicine. In this study, (+)-piperitol-γ,γ-dimethylallylether, a furofuran lignan, was successfully isolated. The chemical structure of compound 1 was determined based on spectroscopic data, including 1D- and 2D-NMR, mass spectroscopy, and by comparing with previous spectral data. In addition, compound 1 was tested for its cytotoxic activity against MCF-7 breast cancer cell lines in vitro and showed weak activity with the IC50 value of 261.37 µg/mL

Kuersetin dan Kuersetin-3-O-Glukosida dari Kulit Batang Sonneratia Alba (Lythraceae)

Two flavonoid compounds, quercetin (1) and quercetin-3-O-glucoside (2) have been isolated from the bark of Sonneratia alba (Lythraceae). Chemical structure of both compounds were determined on the basis of spectroscopic data and comparison with those spectra data previously reported. Compound 1 and 2 exhibited antibacterial activity against Gram-positive bacteria, Staphylococcus aureus and Streptococcus mutans with MIC values of 51.2; 48.8; 72.5; dan 100.7 µg/mL respectively.DOI :http://dx.doi.org/10.15408/jkv.v0i0.315

Isolation and Structure Determination of Stigmaterol from the Stem Bark of Lansium domesticum Corr. Cv. Kokossan

Indonesia is a tropical country with biodiversity and a source of natural compounds such as terpenoids, phenolic, flavonoids, and alkaloid groups. The steroid is one of the large groups of terpenoid compounds with various structures and bioactivities. The steroid was also commonly found in the Meliaceae family. One species of this family is Lansium domesticum Corr. which has been studied to have various activities from isolated compounds (limonoids, sesquiterpenoids, onoceranoids, cycloartanoids, and steroids) as well as from their extracts such as antidiarrheal, antimalarial, antimicrobial, antiinsecticide, antifeedant and were used as traditional medicine such as eye drops and scorpion stings. The n-hexane fraction from the bark of L. domesticum Cor. cv. Kokosan produced steroid compounds whose structure is determined by IR, 1H-NMR, 13C-NMR, and DEPT spectroscopy methods and compared with the literature. It was concluded that the steroid isolated was (22E)-stigmasta-5,22-dien-3ol and was first isolated from this genus. This compound was inactive against MCF-7 breast cancer cells with an IC50 value of 832.14 µM