Judul: Inhibiton and Staphylococcus aureus Cell Membrane Leakage by Papaya Latex.
Abstrak: Dangke is a traditional milk curd product, made by coagulation of milk using fresh papaya latex. This product is usually kept at room temperature until consumption and it can lasting until 24 hours. Dried papaya latex in dangke production was considered not only as coagulant but also act as antibacterial agent. In this study, the antibacterial activity of dried papaya latex toward S. aureus was determined by direct contact in broth media and dangke. Fresh papaya latex was dried using vacuum oven at 50-55 for 22 hours. As much as 0.0277 g/100 mL in concentration of dried papaya latex could reduce approximately 1 log CFU/mL of S. aureus and 0.0011 g/100 mL of papain showed inhibition effect instead of reducing effect against S. aureus in broth after 24 hours. Dried papaya latex and papain also showed ability to inhibit S. aureus growth in dangke within 24 hours storage at room temperature. The antibacterial activity of non-proteolytic compound of papaya latex, i.e ethanolic extract of papaya latex was determined by macrodilution method, resulted in the MIC90 of 8 mg/mL. Antibacterial agent could inhibit bacterial growth by some mechanisms, one of them is cell membrane damage, protein synthesize and DNA synthesize disruption. Papaya latex which contains proteolytic enzyme as bioactive compound was considered can cause the cell membrane leakage. The antibacterial activity toward cell membrane caused the leakage of 260 nm absorbing material from cell. In this study, the cell membrane leakage after exposure was detected by measuring the optical density of bacterial supernatant at 260 nm. The result showed that exposure to increasing antibacterial concentration resulted in increasing of optical density of S. aureus supernatant, indicating that the antibacterial caused the S. aureus membrane leakage. Imaging by fluorescence microscopy showed that S. aureus exposure to antibacterial caused the membrane leakage thus gave Propidium Iodide (PI) chance to penetrate onto the cell, which indicated by the changing of fluorescence color from green fluorescence to red fluorescence.
Keyword: cell membrane leakage, dangke, dried papaya latex

Judul: Aktivitas ekstrak kasar alkaloid daun pepaya terhadap Staphylococcus aureus dan ekspresi gen Staphylococcal enterotoxina
Abstrak: Papaya plant is widely grown in Indonesia. Papaya leaves are used by people as vegetable, meat tenderizer and traditional herbs. Extracts of papaya leaves have been reported to have antimicrobial activity against both Gram positive and Gram negative bacteria. This antimicrobial activity was related to phytochemical content of papaya leaves. Tannins and terpenoids can make cell membrane disruption, whereas alkaloids have been shown to intercalate into DNA and inhibit DNA synthesis. Therefore, besides as meat tenderizer, papaya leaves can be an alternative natural preservatives in food. Staphylococcus aureus is known pathogen causing intoxication by producing staphylococcal enterotoxins (SEs) in food. Staphylococcal enterotoxin A (SEA) is one of SEs that is commonly implicated in staphylococcal food poisoning. SEA is synthesized from sea gene and expressed from the midexponential phase of growth. In addition to the ability to inhibit S. aureus growth, there is a potential action of papaya leaves extract, i.e the ability to inhibit SEA production through the alkaloids activity. This study was conducted to investigate the ability of crude alkaloid extract from papaya leaves to inhibit SEA expression. Crude alkaloid extract was extracted from grounded dried papaya leaves using ultrasonic-assisted extraction. S. aureus was isolated from ready to eat food and raw cow milk using modified Bacteriological Analytical Manual (BAM) method. Isolates with typical colony of S. aureus on Baird-Parker Agar (BPA) and Mannitol Salt Agar (MSA) and produced coagulase were identified using API Staph test and PCR of 16S rRNA gene, followed by sequencing. Those 16S rRNA gene sequence were analyzed using BLAST. Minimum Inhibitory Concentration (MIC) of crude alkaloid extract were determined by broth macro-dilution method. Furthermore, S. aureus isolate was exposed to crude alkaloid extract at 0, 1, and 2 folds of MIC, and subsequently the expression of sea gene was analyzed using a quantitative reverse transcription PCR (qRT-PCR). The results demonstrated that the yield of crude alkaloid extracts was 0.48% to 1.82% per dry weight of papaya leaves. SEA-producing S. aureus were obtained from some foods, i.e raw milk, fried egg in chilli sauce, sauteed chicken intestine, and chicken liver satay. The MIC of crude alkaloid extract to S. aureus was 0.25 mg/ml. After exposure to the crude alkaloid extract at 0.25 mg/ml and 0.5 mg/ml for 2 h, significant increase in cycle threshold (CT) values of sea gene was observed. These results indicated a decrease of initial amount of sea cDNA which meant lesser mRNA was synthesized from the sea gene. The sea gene was expressed 29 and 41 times less when S. aureus was exposed to crude alkaloid extract at 1 and 2 folds MIC, respectively, determined relatively to the calibrator sample and an internal control using comparative CT method. The crude alkaloid extract of papaya leaves possessed antibacterial activity against S. aureus which was not only inhibited the bacteria but also the toxin formation by suppressing the expression of sea gene.
Keyword: crude alkaloid extract, papaya leaves, Staphylococcus aureus, staphylococcal enterotoxin A

Judul: Cycle Hamilton pada Graf Lengkap, Graf Regular, dan Graf 2-Connected 4-Regular Berorder Kurang dari Sepuluh
Abstrak: Suatu graf disebut graf Hamilton apabila graf tersebut memuat cycle Hamilton, yaitu cycle yang melewati setiap vertex tepat satu kali. Menentukan keberadaan cycle Hamilton pada suatu graf dapat menggunakan syarat cukup yang berupa teorema-teorema yang telah diperkenalkan sejak tahun 1950-an. Beberapa syarat cukup yang dibahas dalam karya ilmiah ini adalah syarat cukup yang berhubungan dengan graf lengkap dan graf regular. Suatu graf lengkap berorder 𝑛������ ≥ 3 dan graf r-regular berorder 𝑛������ ≤ 2𝑟������ + 1 merupakan graf Hamilton, Karena kedua graf tersebut memuat cycle Hamilton. Selanjutnya, syarat cukup tersebut digunakan untuk menentukan keberadaan cycle Hamilton pada graf 2-connected 4- regular berorder kurang dari 10.
Keyword: graf Hamilton, graf regular, graf lengkap, graf 2-connected 4-regular