scholarly journals Bioassay Guided Fractionation of Marine Streptomyces sp. GMY01 and Antiplasmodial Assay using Microscopic and Flow Cytometry Method

2020 ◽  
Author(s):  
Ema Damayanti ◽  
Jaka Widada ◽  
Puspa Dewi N. Lotulung ◽  
Achmad Dinoto ◽  
Mustofa Mustofa
Keyword(s):  
Flow Cytometry ◽  
Ethyl Acetate ◽  
Secondary Metabolite ◽  
Genome Mining ◽  
Vero Cells ◽  
Flash Chromatography ◽  
Main Constituent ◽  
Methanol Fraction ◽  
Streptomyces Sp ◽  
Bioassay Guided Fractionation

Genome mining study showed that marine-derived Streptomyces sp. GMY01 is a potential actinobacteria with abundant of secondary metabolite and anticancer activity. The study aimed to evaluate bioassay guided fractionation for antiplasmodial screening of GMY01 extract and to predict compound on active fractions. Ethyl acetate fraction was obtained from 11 days fermentation product of GMY01 and then it was fractionated using n-hexane and methanol solvent. Refractionated was applied using flash chromatography and column chromatography. Antiplasmodial assay was performed on Plasmodium falciparum FCR3 by microscopic method using thin blood smear + Giemsa stain, and flow cytometry method using SYBR Green I stain. Toxicity assay was performed on Vero cells line. Main constituent of active fraction was analyzed using LCMS/MS. The result of the study showed that ethyl acetate-methanol fraction has high antiplasmodial activity (IC50=3.96 µg/mL) with very low toxicity on Vero cells (IC50=30,072 µg/mL). Bioassay guided fractionation resulted F4.7 as highest Plasmodium inhibition (94.3% at 5 µg/mL) and was confirmed by microscopic and flow cytometry assay. Main constituent analysis showed C10H13NO (163.09971 Da) as mayor compound and predicted as nonribosomal polyketide synthetase (NRPS) secondary metabolite.

scholarly journals THE ANTIBACTERIAL COMPOUND COLLISMICIN A DERIVED FROM MARINE Streptomyces sp Q-629K

2010 ◽  
Vol 8 (3) ◽  
pp. 426-430
Author(s):  
Tutik Murniasih ◽  
Kyoko Adachi
Keyword(s):  
Ethyl Acetate ◽  
Secondary Metabolite ◽  
13C Nmr ◽  
Active Compound ◽  
Chemical Structure ◽  
Ethyl Acetate Extract ◽  
Water System ◽  
Antimicrobial Compound ◽  
Streptomyces Sp

In our course of screening for secondary metabolite derived from marine bacterial, we isolate the antimicrobial compound collysmicin A from the ethyl acetate extract of Streptomyces sp Q-629K. Separation of this compound was carried out by silica gel open column chromatography. Purification of an active compound was done using HPLC C18 with acetonitril-water system. Determination of chemical structure was done by 1H, 13C NMR and LC-MS analysis. Collysmicin A was contained in fraction 3, fraction 7.2 and fraction 8.7. The antimicrobial assayed from purified compound Fr.8.7 gave diameter inhibition approximately 13 mm against S. aureus and 12 mm against B. subtilis .   Keywords: antimicrobial, collismycin A and marine Streptomyces sp

scholarly journals CYTOTOXIC ACTIVITY AND APOPTOSIS INDUCTION OF T47D CELL LINES BY Turbinaria decurrens EXTRACT

2013 ◽  
Vol 8 (1) ◽  
pp. 23 ◽  
Author(s):  
Muhammad Nursid ◽  
Nurrahmi Dewi Fajarningsih ◽  
Ekowati Chasanah
Keyword(s):  
Flow Cytometry ◽  
Ethyl Acetate ◽  
Cytotoxic Activity ◽  
Cell Lines ◽  
Crude Extract ◽  
T47d Cell ◽  
Apoptosis Induction ◽  
Flow Cytometry Analysis ◽  
Methanol Fraction ◽  
Ic50 Value

Marine algae is known to contain a wide variety of biomedical compounds having pharmaceutical applications. The aim of this research was to evaluate cytotoxic activity and apoptosis induction of Turbinaria decurrens extract on T47D cell lines.  Cytotoxic activity test was conducted by using MTT assay whereas detection of apoptosis was evaluated by DNA fragmentations and flow cytometry analysis. The MTT test showed that crude extract had medium cytotoxic activity to T47D, HepG2, and C28 cell lines with IC50 value of 172, againts 360 and 330 µg ml-1, respectively. After solvent partition of crude extract, the cytotoxic activity of n-hexane and ethyl acetate fractions T47D cell increased, the cytotoxic activity of n. hexane and ethyl acetate fractions T47D cell increased with  IC50  value of with IC50  43.1 and 51.9 µg ml-1, respectively, whereas IC50 value of methanol fraction was 383.0 µg ml-1. Analysis of DNA fragmentation of T47D cell showed that  both n-hexane and ethyl acetate fractions could not fragment DNA as a features of apoptosis. However, flow cytometry analysis by using annexin-V and propidium iodide staining revealed that n-hexane and ethyl acetate fractions could induce apoptosis in T47D cell. This research indicated that Turbinaria decurrens had potency to induce apoptosis in T47D cells.

Metabolite fingerprinting of novel Streptomyces UK-238 from the Himalayan forest

2020 ◽  
Vol 16 ◽  
Author(s):  
Nidhi Srivastava ◽  
Indira P. Sarethy
Keyword(s):  
Ethyl Acetate ◽  
16S Rdna ◽  
Retention Index ◽  
Ethyl Acetate Extract ◽  
Similarity Index ◽  
Antimicrobial Drug ◽  
16S Rdna Sequencing ◽  
Metabolite Fingerprinting ◽  
Streptomyces Sp ◽  
Rdna Sequencing

Aims: Characterization of antimicrobial metabolites of novel Streptomyces sp. UK-238. Background: Novel antimicrobial drug discovery is urgently needed due to emerging multi antimicrobial drug resistance among pathogens. Since many years, natural products have provided the basic skeletons for many therapeutic compounds including antibiotics. Bioprospection of un/under explored habitats and focussing on selective isolation of actinobacteria as major reservoir of bio and chemodiversity has yielded good results. Objective: The main objectives of the study were the identification of UK-238 by 16S rDNA sequencing and antimicrobial metabolite fingerprinting of culture extracts. Method: In the present study, a promising isolate, UK-238, has been screened for antimicrobial activity and metabolite fingerprinting from the Himalayan Thano Reserve forest. It was identified by 16S rDNA sequencing. Ethyl acetate extract was partially purified by column chromatography. The pooled active fractions were fingerprinted by GC-MS and compounds were tentatively identified by collated data analysis based on Similarity Index, observed Retention Index from Databases and calculated Retention Index. Results: UK-238 was identified as Streptomyces sp. with 98.4% similarity to S. niveiscabiei. It exhibited broad-spectrum antibacterial and antifungal activity. GC-MS analysis of active fractions of ethyl acetate extract showed the presence of eighteen novel antimicrobial compounds belonging to four major categories- alcohols, alkaloid, esters and peptide. Conclusion: The study confirms that bioprospection of underexplored habitats can elaborate novel bio and chemodiversity.

Identification of the kinanthraquinone biosynthetic gene cluster by expression of an atypical response regulator

2021 ◽  
Vol 85 (3) ◽  
pp. 714-721
Author(s):  
Risa Takao ◽  
Katsuyuki Sakai ◽  
Hiroyuki Koshino ◽  
Hiroyuki Osada ◽  
Shunji Takahashi
Keyword(s):  
Heterologous Expression ◽  
Gene Cluster ◽  
Secondary Metabolite ◽  
Response Regulator ◽  
Bac Library ◽  
Gene Clusters ◽  
Biosynthetic Gene Cluster ◽  
Gene Organization ◽  
Biosynthetic Gene ◽  
Streptomyces Sp

ABSTRACT Recent advances in genome sequencing have revealed a variety of secondary metabolite biosynthetic gene clusters in actinomycetes. Understanding the biosynthetic mechanism controlling secondary metabolite production is important for utilizing these gene clusters. In this study, we focused on the kinanthraquinone biosynthetic gene cluster, which has not been identified yet in Streptomyces sp. SN-593. Based on chemical structure, 5 type II polyketide synthase gene clusters were listed from the genome sequence of Streptomyces sp. SN-593. Among them, a candidate gene cluster was selected by comparing the gene organization with grincamycin, which is synthesized through an intermediate similar to kinanthraquinone. We initially utilized a BAC library for subcloning the kiq gene cluster, performed heterologous expression in Streptomyces lividans TK23, and identified the production of kinanthraquinone and kinanthraquinone B. We also found that heterologous expression of kiqA, which belongs to the DNA-binding response regulator OmpR family, dramatically enhanced the production of kinanthraquinones.

scholarly journals Chitin Degradation Machinery and Secondary Metabolite Profiles in the Marine Bacterium Pseudoalteromonas rubra S4059

Marine Drugs ◽  
2021 ◽  
Vol 19 (2) ◽  
pp. 108
Author(s):  
Xiyan Wang ◽  
Thomas Isbrandt ◽  
Mikael Lenz Strube ◽  
Sara Skøtt Paulsen ◽  
Maike Wennekers Nielsen ◽  
...  
Keyword(s):  
Secondary Metabolite ◽  
Genetic Manipulation ◽  
Genome Mining ◽  
Hydrolytic Enzymes ◽  
Metabolite Profile ◽  
Chitinolytic Enzyme ◽  
Chitin Degradation ◽  
Metabolite Profiles ◽  
Bioactive Potential ◽  
Pseudoalteromonas Rubra

Genome mining of pigmented Pseudoalteromonas has revealed a large potential for the production of bioactive compounds and hydrolytic enzymes. The purpose of the present study was to explore this bioactivity potential in a potent antibiotic and enzyme producer, Pseudoalteromonas rubra strain S4059. Proteomic analyses (data are available via ProteomeXchange with identifier PXD023249) indicated that a highly efficient chitin degradation machinery was present in the red-pigmented P. rubra S4059 when grown on chitin. Four GH18 chitinases and two GH20 hexosaminidases were significantly upregulated under these conditions. GH19 chitinases, which are not common in bacteria, are consistently found in pigmented Pseudoalteromonas, and in S4059, GH19 was only detected when the bacterium was grown on chitin. To explore the possible role of GH19 in pigmented Pseudoalteromonas, we developed a protocol for genetic manipulation of S4059 and deleted the GH19 chitinase, and compared phenotypes of the mutant and wild type. However, none of the chitin degrading ability, secondary metabolite profile, or biofilm-forming capacity was affected by GH19 deletion. In conclusion, we developed a genetic manipulation protocol that can be used to unravel the bioactive potential of pigmented pseudoalteromonads. An efficient chitinolytic enzyme cocktail was identified in S4059, suggesting that this strain could be a candidate with industrial potential.

scholarly journals Draft Genome Sequences of Fungus Aspergillus calidoustus

2016 ◽  
Vol 4 (2) ◽  
Author(s):  
Fabian Horn ◽  
Jörg Linde ◽  
Derek J. Mattern ◽  
Grit Walther ◽  
Reinhard Guthke ◽  
...  
Keyword(s):  
Secondary Metabolite ◽  
Genome Sequence ◽  
Functional Annotation ◽  
Draft Genome ◽  
Genome Mining ◽  
Gene Clusters ◽  
Draft Genome Sequence ◽  
Genome Sequences

Here, we report the draft genome sequence of Aspergillus calidoustus (strain SF006504) . The functional annotation of A. calidoustus predicts a relatively large number of secondary metabolite gene clusters. The presented genome sequence builds the basis for further genome mining.

scholarly journals Antioxidant activity test of ethyl acetate fraction of binjai (Mangifera Caesia) leaf ethanol extract

2018 ◽  
Vol 51 (4) ◽  
pp. 164
Author(s):  
K. Khairiah ◽  
Irham Taufiqurrahman ◽  
Deby Kania Tri Putri
Keyword(s):  
Antioxidant Activity ◽  
Ascorbic Acid ◽  
Ethyl Acetate ◽  
Secondary Metabolite ◽  
Antioxidant Properties ◽  
Linear Connection ◽  
Sampling Technique ◽  
Ethanol Extract ◽  
Ethyl Acetate Fraction ◽  
Control Group

Background: Binjai (Mangifera caesia) is a herb derived from South Kalimantan possessing antioxidant properties which promote wound healing inhibiting oxidation radicals. The natural antioxidants present in binjai leaves can be extracted by fractionation. Purpose: This study aimed to analyze the antioxidant activity of ethyl acetate fraction in 96% ethanol extract of binjai leaf. Methods: The study constituted a pure experimental study incorporating a post-test design with only random sampling technique consisting of two groups, namely; an ethyl acetate fraction as the treatment group and ascorbic acid as the positive control group. The leaves were treated in accordance with the soxhlet method and subsequently fractionated to extract ethyl acetate fraction. This was used to measure antioxidant activity with DPPH radical damping method using a UV-Vis spectrophotometer. A linear regression calculation was performed with a standard curve to quantify the IC50 value, before the ethyl acetate fraction underwent a qualitative test of secondary metabolite. Results: An independent t-test indicated significant differences between groups, an average value of IC50 in ascorbic acid of 13.812 ppm with 0.996 linearity and a fraction of ethyl acetate 38.526 ppm with a linearity of 0.999. In contrast, at this linearity value ascorbic acid and ethyl fraction acetate demonstrate a very high linear connection between concentration and inhibition. A secondary metabolite test conducted on the ethyl acetate fraction produced positive results for flavonoid, tannins, and phenol. Conclusion: Based on the IC50 parameters, the fraction of ethyl acetate in 96% ethanol extract of binjai leaf produces very strong antioxidant activity in the content of the compounds in the fraction, namely: flavonoid, tannins and phenol.

scholarly journals Isolation of sea cucumbers’s (Holothuria atra) secondary metabolite using column-chromatography technique

2021 ◽  
Vol 869 (1) ◽  
pp. 012010
Author(s):  
S Agustina ◽  
S Bella ◽  
S Karina ◽  
I Irwan ◽  
M Ulfah
Keyword(s):  
Secondary Metabolites ◽  
Ethyl Acetate ◽  
Secondary Metabolite ◽  
Column Chromatography ◽  
Extraction Method ◽  
Sea Cucumber ◽  
Sea Cucumbers ◽  
Marine Chemistry ◽  
Holothuria Atra ◽  
Metabolite Content

Abstract Identification of sea cucumbers from Benteng Inong Balee, Aceh Besar and their phytochemistry screening were conducted in December 2020 to January 2021 at Laboratory of Marine Chemistry and Fisheries Biotechnology, Universitas Syiah Kuala. The purpose of this study was to identify the species of sea cucumbers and its secondary metabolite content using phytochemistry screening and column chromatography. The species of sea cucumbers that were identified was Holothuria atra. The extraction method used in sea cucumber extraction was maceration method, while the separation of secondary metabolites used column-chromatography with eluent of n-hexane : ethyl acetate (8:4). The results showed that secondary metabolites obtained from phytochemical tests were flavonoids, saponins and triterpenoids.

scholarly journals Antagonistic activity of terrestrial Streptomyces sp. VITNK9 against Gram negative bacterial pathogens affecting the fish and shellfish in aquaculture

2018 ◽  
Vol 53 (2) ◽  
pp. 171 ◽  
Author(s):  
Mohammed Ishaque Nabila ◽  
Krishnan Kannabiran
Keyword(s):  
Antibacterial Activity ◽  
Ethyl Acetate ◽  
Vibrio Harveyi ◽  
Bacterial Pathogens ◽  
Vibrio Anguillarum ◽  
Antagonistic Activity ◽  
Edwardsiella Tarda ◽  
Streptomyces Sp ◽  
Aeromonas Caviae ◽  
Fish And Shellfish

A total of 72 morphologically different actinomycetes isolates were isolated from samples collected at different regions of Vellore, Tamil Nadu, India and screened for its antibacterial activity against fish and shellfish pathogens. All actinomycetes isolates were screened for antibacterial activity by cross streak method against the selected fish and shellfish bacterial pathogens including Aeromonas caviae, Aeromonas hydroplila, Edwardsiella tarda, Vibrio anguillarum and Vibrio harveyi. Secondary screening of antagonistic isolates by well diffusion method leads to the identification of potential isolate. Culture conditions for the potential isolate were optimized for maximal growth and yield of the ethyl acetate (EA) crude extract. The potential isolate was characterized by molecular taxonomy and phylogeny and identified as Streptomyces species and named as Streptomyces sp. VITNK9. The 16S rDNA nucleotide sequence was searched through the GenBank database and showed 83% similarity to Streptomyces vinaceusdrappus. The EA extract prepared from Streptomyces sp. VITNK9 showed moderate antagonistic activity accessed by the formation of zone of growth inhibition against, Aeromonas caviae (15.33 mm), Aeromonas hydrophila (17.66 mm), Edwardsiella tarda (18.33 mm), Vibrio anguillarum (14.33 mm) and Vibrio harveyi (14.33 mm). The MIC value of EA extract was ranged between 0.03-0.125 mg mL-1. The GC-MS spectrum of the ethyl acetate extract revealed the presence of two major compounds, pyrrolo [1,2-A] pyrazine-1,4-Dione (56.67%) and Hexahydro-3-(2-Methylpropyl) (27.91%), respectively. The results of the study suggest that Streptomyces sp. VITNK9 is a potential source for antagonistic secondary metabolites against fish and shellfish bacterial pathogens.

AKTIVITAS ANTIBAKTERI DAN ANALISIS KLT-BIOAUTOGRAFI DARI FRAKSI DAUN MANGGIS (Garcinia mangostana L.)

PHARMACON ◽  
2019 ◽  
Vol 8 (3) ◽  
pp. 525
Author(s):  
Ayu Natasya Paputungan ◽  
Widya Astuty Lolo ◽  
Imam Jayanto
Keyword(s):  
Escherichia Coli ◽  
Staphylococcus Aureus ◽  
Antibacterial Activity ◽  
Ethyl Acetate ◽  
Ethyl Acetate Fraction ◽  
Diffusion Method ◽  
Garcinia Mangostana ◽  
Methanol Fraction ◽  
Tlc Bioautography ◽  
Fractionation Method

Mangosteen leaves have flavonoid compounds, tannins, and saponins that can be efficacious as antibacterial. The aim of this study was to determine the fraction of mangosteen leaves having an antibacterial effect and knowing the class of compounds identified as having antibacterial activity after TLC- Bioautography testing was carried out. The samples were extracted using 96% maceratarion method and fractioned using liquid-liquid fractionation method with methanol, n-hexane and ethyl acetate solvents, antibacterial activity using agar diffusion method (Kirby and Bauer) with 3 concetrations namely 10%, 20% and 30%. Thin Layer Chromatography (TLC) uses n-hexane and chloroform solvens. TLC-Bioautography uses contact bioautography methods. The resultd showed that mangosteen leaves in methanol fraction with a concentration of 30% had a very large inhibitory activity again Staphylococcus aureus and ethyl acetate fraction with a concentration of 30% had the gratest antibacterial activity against  Escherichia coli. The results of the TLC- Bioautography study showed that the flavonoids compounds after spraying with AlCl3 and the mangosteen leaf Biosutography test had inhibitory zone activity against the bacteria Staphylococcus aureus dan Escherichia coli. Keywords: Mangosteen Leaves. Antibacterial, TLC Bioautography.  ABSTRAK Daun manggis mempunyai senyawa flavonoid, tanin, dan saponin yang dapat berkhasiat sebagai antibakteri. Penelitian ini bertujuan untuk mengetahui fraksi daun manggis memiliki efek antibakteri dan mengetahui golongan senyawa yang teridentifikasi memiliki aktivitas antibakteri setelah dilakukan pengujian KLT Bioautografi. Sampel diektrak dengan metode maserasi dengan pelarut 96% dan difraksinasi dengan metode  fraksinasi cair-cair dengan pelarut metanol, n-heksan dan etil asetat, aktivitas antibakteri menggunakan metode difusi agar (Kirby and Bauer) dengan 3 kosentrasi yaitu 10%, 20% dan 30%. Kromatografi Lapis Tipis (KLT) menggunakan pelarut n-heksan dan klorofom. KLT-Bioautografi menggunakan metode bioautografi kontak. Hasil penelitian menunjukan daun manggis pada fraksi metanol  dengan kosentrasi 30% memiliki aktivitas zona hambat ppaling besar terhadapat bakteri Staphylococcus aureus dan fraksi etil asetat dengan kosentrasi 30% memiliki aktivitas antibakteri paling besar terhadap  Escherichia coli. Hasil penelitian KLT-Bioautografi menunjukan golongan senyawa flavonoid setelah disemprotkan dengan AlCl3 dan uji Bioautografi daun manggis memiliki aktivitas zona hambat terhadap bakteri Staphylococcus aureus dan Escherichia coli. Kata kunci : Daun Manggis, Antibakteri, KLT- Bioautografi.

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