Ilham Harlan Amarullah
Dengue Study Group, Institute Of Tropical Disease, Universitas Airlangga

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RNA Isolation of Dengue Virus Type 2 with Different Precipitation Solvents : Methanol, Chloroform, and 2-Isopropanol. Yovilianda Maulitiva Untoro; Teguh Hari Sucipto; Harsasi Setyawati; Siti Churrotin; Ilham Harlan Amarullah; Puspa Wardhani; Aryati Aryati; Shuhai Ueda; Soegeng Soegijanto
Jurnal Kimia Riset Vol. 3 No. 1 (2018): Juni
Publisher : Universitas Airlangga

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (416.739 KB) | DOI: 10.20473/jkr.v3i1.7455

Abstract

Dengue virus distributed in tropical and subtropical regions in the world. DENV viruses are transmitted between humans primarily by Aedes aegypti and Aedes albopictus mosquitoes and are endemic in most areas in which the vectors occur. Four serotypes of dengue virus are DENV-1, DENV-2, DENV-3 and DENV-4. DENV-2 is comprised of six genotypes. Ethanol precipitation is a commonly used technique for concentrating and de-salting nucleic acids (DNA or RNA) preparations in aqueous solution. RNA isolation by combining Guanidinium thiocyanate and phenol reported has been reported. In this report, we investigated RNA isolation from DENV-2 using QIAamp Mini Kit with 2-Isopropanol, Methanol, Chloroform precipitation solvent. Electrophoregram showed DNA band as  the result of RNA isolation with methanol and 2-isopropanol are produced quite well. Dna band of the of RNA isolation with chloroform solvent has the lowest intensity than methanol and 2-isopropanol. This study showed that methanol and 2-isopropanol  can used as precipitation solvent for isolating RNA.
Precipitation Solvents for RNA Extraction of Dengue Virus Type 3: Dimethylformamide, Ethylenediamintetraacetic Acid, and Ultrapure H2O Rizqidhana Juliana Putri; Teguh Hari Sucipto; Harsasi Setyawati; Siti Churrotin; Ilham Harlan Amarullah; Puspa Wardhani; Aryati Aryati; Soegeng Soegijanto
Jurnal Kimia Riset Vol. 3 No. 2 (2018): Desember
Publisher : Universitas Airlangga

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (477.838 KB) | DOI: 10.20473/jkr.v3i2.9353

Abstract

Dengue is a disease caused by a virus from the family Flaviviradae, carried by a female mosquito of Aedes aegypti species. Dengue fever is widespread in the tropic areas. It caused by rainfall, temperature and unplanned urbanization. According to the ministry of health , almost all provinces in Indonesia are endemic areas of dengue fever. In 2014, up to mid-December Dengue Hemorrhagic Fever (DHF) patients in 34 provinces in Indonesia are 71,668 people and 641. This figure is lower than the previous year, 2013 with 112,511 people and 871 deaths . This disease consists of four types of serotypes, namely DENV-1, DENV-2, DENV-3, and DENV-4. This disease can be identified using a variety of methods, one of the method is Reverse Transcription - Polymerase Chain Reaction (RT-PCR) method. This study aims to determine the ability of Dimethylformamide (DMF), Ethylenediamintetraacetic Acid (EDTA), and Ultrapure H2O as the substitute of  Ethanol for precipitation in RNA extraction process. The sample used in this research obtained from Surabaya. RNA extraction itself can be done by using a special kit for RNA extraction. In Reverse Transcription - Polymerase Chain Reaction method, first RNA is extracted and then transcribed back (Reverse Transcription) which then form cDNA that later will be amplified by using PCR method. In this study used specific primers for dengue virus type 3 (DENV-3). The results of this study show that DMF, EDTA, and Ultrapure H2O can be used as the substitute of Ethanol for precipitation on RNA extraction. The result is evidenced by the formation of viral DNA bands on gel electrophoresis results.
In Vitro Study: Effect of Cobalt(II) Chloride Against Dengue Virus Type 1 in Vero Cells Teguh Hari Sucipto; Yovilianda Maulitiva Untoro; Harsasi Setyawati; Anisa Maharani; Novia Faridatus Sholihah; Siti Churrotin; Ilham Harlan Amarullah; Soegeng Soegijanto
Indonesian Journal of Pharmacy Vol 30 No 4, 2019
Publisher : Faculty of Pharmacy Universitas Gadjah Mada, Yogyakarta, Skip Utara, 55281, Indonesia

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.14499/indonesianjpharm30iss4pp316

Abstract

Dengue virus (DENV) serotypes DENV-1 to DENV-4 are enveloped viruses that belong to the genus Flavivirus of the Flaviviridae. Dengue vaccine or antiviral has not yet been clinically approved for humans, even though there have been great efforts toward this end. Antiviral activity against DENV is needed to develop to be an alternative drug for DENV virus. Cobalt(II) chloride have been used in the treatment and prevention of diseases of humans since ancient times. The aim of this study is to investigate the antiviral effects and Cytotoxicity of Cobalt(II) chloride. This compound was further investigated for its inhibitory effect on the replication of DENV-1 in Vero cells. Antiviral activity and Cytotoxicity measured by WST-1 assay. The IC50 value of the Cobalt(II) chloride for DENV-1 was 0.38 μg/ml. The cytotoxicity of Cobalt(II) chloride to Vero cell suggest that the CC50 value was 2.91 µg/ml The results of this study demonstrate the anti-dengue serotype 1 inhibitory activity of Cobalt(II) chloride was a high toxic compound.
INHIBITORY ACTIVITY OF COBALT(II)–MORIN COMPLEX AGAINST THE REPLICATION OF DENGUE VIRUS TYPE 2 Sucipto, Teguh Hari; Churrotin, Siti; Setyawati, Harsasi Setyawati; Mulyatno, Kris Cahyo; Amarullah, Ilham Harlan; Ueda, Shuhai; Kotaki, Tomohiro; Sumarsih, Sri; Wardhani, Puspa; Bendryman, Sri Subekti; Aryati, Aryati; Soegijanto, Soegeng; Kameoka, Masanori
Indonesian Journal of Tropical and Infectious Disease Vol. 6 No. 6 (2017)
Publisher : Institute of Topical Disease Universitas Airlangga

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (402.971 KB) | DOI: 10.20473/ijtid.v6i6.6126

Abstract

Dengue virus (DENV) is a significant pathogen emerging worldwide as a cause of infectious disease. Antidengue treatments are urgently required to control the emergence of dengue. DENV is a mosquito-borne disease responsible for acute systemic diseases and serious health conditions. DENVs were distributed in the tropical and sub-tropical areas and transmitted to humans by Aedes agypty and Aedes albopictus. Dengue vaccine or antiviral has not yet been clinically approved for humans, even though there have been great efforts toward this end. Antiviral activity against DENV is an important alternative for the characterization and development of drugs. Metal–organic compounds were reported to exhibit fungicidal, bactericidal, and antiviral activities its inhibitory activity was not significant, at high concentration it was more toxic to replicating cells than to stationary cell monolayers of Vero cells. The aim of this study is to investigate the antiviral effects of Cobalt(II)–Morin complex. This compound was further investigated for its inhibitory effect on the replication of DENV-2 in Vero cells. The replication of DENV was measured by enzyme-linked immunosorbent assay and the value of selectivity index (SI). SI was determined as the ratio of the 50% cytotoxic concentration (CC50) to the 50% inhibitory concentration (IC50). The IC50 value of the Cobalt(II)–Morin complex for DENV-2 was 3.08 µg/ml, and the CC50 value of the complex for Vero cells was 3.36 µg/ml; thus, the SI value was 1.09. The results of this study demonstrate the antidengue serotype 2 inhibitory activity of Cobalt(II)–Morin complex and its high toxicity in Vero cells. Further studies are not required before Co(II)–Morin can be applied in the treatment of DENV-2 infections.
RNA ISOLATION OF DENGUE VIRUS TYPE 1 WITH DIFFERENT PRECIPITATION SOLVENTS: DIMETHYL SULFOXIDE, ACETONE, AND ETHANOL 70% Maharani, Anisa; Sucipto, Teguh Hari; Setyawati, Harsasi; Churrotin, Siti; Amarullah, Ilham Harlan; Wardhani, Puspa; Aryati, Aryati; Ueda, Shuhai; Soegijanto, Soegeng
Indonesian Journal of Tropical and Infectious Disease Vol. 7 No. 3 (2018)
Publisher : Institute of Topical Disease Universitas Airlangga

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (585.948 KB) | DOI: 10.20473/ijtid.v7i3.6748

Abstract

Dengue Hemorrhagic Fever (DHF) is caused by dengue viruses that belong to Flaviviridae. The disease is known to be caused by 4 types of dengue viruses, namely DENV-1, DENV-2, DENV-3, and DENV-4 associated with antigenic. Dengue virus is a virus RNA that causes illness with clinical manifestations of Dengue Fever, Dengue Hemorrhagic Fever and Dengue Shock Syndrome. The aim of research was to determine the effectiveness of dimethyl sulfoxide, acetone, and ethanol 70% as precipitation solvent in the process of RNA isolation. The method used was Reverse Transcription - Polymerase Chain Reaction (RT-PCR) and Polymerase Chain Reaction (PCR) with specific primers for dengue virus type 1 (DENV-1). RNA isolation can be done easily using an RNA Isolation Kit. Use of RNA Isolation Kit results in a purer RNA isolate from contaminants and from RNA degradation. In generally the isolation is using cold ethanol / alcohol with concentration 90-95%. Ethanol / Alcohol does not dissolve RNA and light density of alcohol lighter than water makes RNA rise and hover on the surface. In RNA isolation solvent precipitation that used are acetone, ethanol 70%, and DMSO. In qualitative RNA measurements using agarose gel electrophoresis and was examined under the UV light-illuminator and quantitative RNA measurements using Nanodrop spectrophotometry with absorbance ratio at 260/280 and 260/230 showed a good result indicated by the appearance of the band on electrophoresis results in PCR. While the measurement quantitatively is showed that there was still protein contamination but the results are quite good because it does not much different from the ratio set in the reference. Acetone, ethanol 70%, and DMSO can be used as a substitute of 96% ethanol in the process of RNA isolation in DENV-1 virus and can also be applied to other dengue virus because the structure of the 4th antigen serotype is very similar one with the other and no effect.
ANTI-DENGUE TYPE 2 VIRUS ACTIVITIES OF ZINC (II) COMPLEX COMPOUNDS WITH 2-(2,4 -DIHYDROXYPHENYL)-3,5,7-TRIHYDROXYCROMEN-4-ONE LIGANDS IN VERO CELLS Sucipto, Teguh Hari; Setyawati, Harsasi; Churrotin, Siti; Amarullah, Ilham Harlan; Sumarsih, Sri; Wardhani, Puspa; Aryati, Aryati; Soegijanto, Soegeng
Indonesian Journal of Tropical and Infectious Disease Vol. 7 No. 5 (2019)
Publisher : Institute of Topical Disease Universitas Airlangga

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (777.006 KB) | DOI: 10.20473/ijtid.v7i5.10851

Abstract

Dengue virus (DENV) is a disease that is transmitted through Aedes aegypti and Aedes albopictus mosquitoes, and is spread in tropical and sub-tropical regions. Now, dengue or antiviral vaccines for humans do not yet exist, but there are great efforts to achieve this goal. Complex compounds are reported to fungicidal, bactericidal and antiviral activity. Antiviral activity against DENV is an important alternative to the characterization and development of drugs candidate. The purpose of this study was to study zinc(II) compounds with 2-(2,4-dihydroxyphenyl)-3,5,7-trihydroxycromen-4-one ligand on DENV-2 replication in Vero cells. Vero cell lines (African green monkey kidney) was used in this study, maintained and propagated in Minimum Essential Eagle Medium containing 10% fetal bovine serum at 37°C in 5% CO2. The activity of dengue virus was carried out by enzyme-immunosorbent assay (ELISA) method and CellTiter96® Non-Radioactive Proliferation. The value of activity inhibition (IC50) of complex compounds with variations of mol metal: ligand 1:2, 1:3, and 1:4 against dengue virus type 2 (DENV2) was 2.44 μg/ml, 2.75 μg/ml, respectively and 2.00 μg/ml, also the toxicity value (CC50) of complex compounds with variation mol metal: ligand 1:4 for Vero cells is 3.59 μg/ml. The results of this study were indicate that these properties have been shown to inhibit anti-dengue type 2 virus (DENV-2), but are also toxic in Vero cells. Including previous study about complex compound interaction with dengue virus type 2 activity, Zn(II) more reactive compound then Cu(II), and Co(II). The comparison with Cu(II) complex compound, it has been revealed that Co(II) and Zn(II) is more toxic, was found to be nontoxic to human erythrocyte cells even at a concentration of 500 μg/ml.
Effect of Zinc(II)-2,4,5-triphenyl-1H-imidazole Complex Against Replication DENV-2 in Vero Cell Sucipto, Teguh Hari; Wibrianto, Aswandi; Martak, Fahimah; Churrotin, Siti; Amarullah, Ilham Harlan; Setyawati, Harsasi; Wardhani, Puspa; Aryati, Aryati; Soegijanto, Soegeng
Indonesian Journal of Tropical and Infectious Disease Vol. 8 No. 3 (2020)
Publisher : Institute of Topical Disease Universitas Airlangga

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.20473/ijtid.v8i3.11776

Abstract

Dengue virus (DENV) is a significant pathogen emerging worldwide as a cause of infectious disease. DENVs are transmitted to humans through female mosquitoes from Aedes aegypti and Aedes albopictus species. Indonesia is one of the largest countries in the world in dengue endemic regions worldwide. Dengue fever was occurred for the first time as an outbreak in Surabaya and Jakarta in 1968. Many efforts have been made to prevent and treat DENV infections, and clinical trials of a number of vaccines are currently underway. Antiviral testing of DENV is an important alternative for drug characterization and development. Complex compounds are formed as a result of metal and organic complex reactions. Complex compounds can be used as an anti-inflammatory, antimicrobial antifungal, antibacterial, antivirus. The Zn2+ ion can be used as an antiviral candidate. The purpose of this project was investigated Zinc(II)-2,4,5-triphenyl-1H-imidazole antiviral compound to be further tested for inhibitory effect on the replication of DENV-2 in cell culture. DENV replication was measured by antiviral activity assay and cytotoxicity assay. The inhibitory activity of Zinc(II)-2,4,5-triphenyl-1H-imidazole complex compound was determined by Viral ToxGloTM Assay. The cytotoxicity of Zinc(II)-2,4,5-triphenyl-1H-imidazole complex compound was determined by CellTiter96® AQuoeus assay. The inhibitory concentration (IC50) of Zinc(II)-2,4,5-triphenyl- 1H-imidazole against dengue virus type-2 was 34.42 μg/ml. The cytotoxic concentration (CC50) of compound against Vero cell was <100 μg/ml. The results of this study demonstrate the antidengue serotype 2 inhibitory activity of investigated Zinc(II)-2,4,5-triphenyl-1H-imidazole complex and its high toxicity in Vero cells. Further studies are not required before investigated Zinc(II)-2,4,5-triphenylimidazole can be applied in the treatment of DENV-2 infections