ABSTRACT
The most practical approach to reduce morbidity and mortality of coronary heart disease (CHD) is to delay the process of thrombus by usage of clot-dissolving agents. The necessities of such safer compounds are to be critically examined for thrombolytic activity especially, from marine sources. Thrombolytic agents have been investigated as a possible treatment for thrombus. The aim of this study was to investigate the in vitro thrombolytic potential of Streptomyces sp.VITJS4 (NCIM No. 5574); (ACC No: JQ234978.1) active compounds. The fibrin degradation revealed a clear transparent zone of clearance with 500µg/mL concentration showing 24mm hydrolysis. The thrombolytic effect of Streptomyces sp.VITJS4 compounds was also demonstrated in vitro clot lysis assay where The percent of thrombolysis by the crude extract showed 90±1.7% at the concentration of 1000µg/mL, whereas percent of thrombolysis by streptokinase was found 100± 00%%. The bioactive compounds were further studied for spectrophotometric analysis. The UV-VIS profile showed different peaks ranging from 400-700 nm with different absorption respectively. The data confirmed the presence of both analogues with absorption maxima at 210 and 310 nm. A sensitive method using LC-MS technique was optimized for the separation and identification of bioactive metabolites which was indicated by the fingerprints. The results of the LC-MS analysis provided different peaks determining the presence of compounds with different therapeutic activities. The current study refers the bioactive compound as impressive thrombolytic agent for further laboratory study. Further studies should be conducted to ensure the efficacy and safety of different concentration of bioactive compounds for drug development. Hence the results reported perhaps useful for the discovery of novel thrombolytic drugs from marine origin.
A abordagem mais prática para reduzir a morbidade e a mortalidade da doença arterial coronariana (CHD, do inglês coronary heart disease) consiste em retardar o processo de trombo através da utilização de agentes de dissolução de coágulos. As necessidades de tais compostos mais seguros devem ser criticamente examinadas para a atividade trombolítica, especialmente de fontes marinhas. Agentes trombolíticos tem sido estudados como um possível tratamento para o trombo. O objetivo deste estudo foi investigar o potencial trombolítico in vitro dos compostos ativos do Streptomyces sp.VITJS4 (NCIM No. 5574); (ACC No: JQ234978.1). A degradação da fibrina revelou um clara zona livre transparente com concentração de 500µg/mL mostrando uma hidrólise de 24mm. O efeito trombolítico dos compostos de Streptomyces sp.VITJS4 também foi demonstrado no ensaio in vitro de lise dos coágulos em que a percentagem de trombólise pelo extrato bruto mostrou 90±1.7% a uma concentração de 1000µg/mL, enquanto que a percentagem de trombólise pela estreptoquinase foi de 100± 00%. Os compostos bioativos foram estudados posteriormente através da análise espectrofotométrica. O perfil ultra violeta visível (UV-VIS profile, em inglês) mostrou diferentes picos variando entre 400-700 nm com diferentes absorções respectivamente. Os dados confirmaram a presença de ambos os análogos com absorção máxima em 210 e 300 nm. Um método sensível usando a técnica LC-MS (Liquid chromatographymass spectrometry) foi otimizado para a separação e identificação metabólitos bioativos que foram indicados pelas impressões digitais (?). Os resultados da análise LC-MS forneceram diferentes picos determinando a presença de compostos com diferentes atividades terapêuticas. O estudo atual refere-se ao composto bioativo como um agente trombolítico impressionante para futuros estudos em laboratório. Estudos futuros devem ser conduzidos para assegurar a eficácia e segurança de diferentes concentrações dos compostos bioativos para o desenvolvimento de drogas. Assim, os resultados reportados talvez sejam úteis para a descoberta de novas drogas trombolíticas de origem marinha.
Subject(s)
Streptomyces , Thrombosis , In Vitro Techniques , Actinobacteria , Fibrinolytic AgentsABSTRACT
In this present study, the bark extracts of Averrhoa bilimbi were subjected to the thrombolytic activities were assessed by using human erythrocyte and the results were compared with standard streptokinase (SK). On the other hand, bark extracts of A. bilimbi revealed moderate antibacterial activity against some microorganisms used in the screening. Preliminary phytochemical investigation suggested the presence of flavonoids, saponins and alkaloids.
ABSTRACT
Objective:To investigate the antioxidant, antimicrobial, cytotoxic and thrombolytic property of the fruits and leaves of Spondias dulcis (S. dulcis). Methods:Methanolic extracts of fruits and leaves of S. dulcis were partitioned with chloroform and dichloromethane. The antioxidant potential of the crude extract and partitioned fractions were evaluated in terms of total phenolic content, total flavonoid content, DPPH radical scavenging potential, reducing potential and total antioxidant capacity by specific standard procedures. The antimicrobial activity was evaluated using disc diffusion method. The cytotoxicity was evaluated by using brine shrimp lethality bioassay and compared with vincristine sulfate. The thrombolytic activity was compared with streptokinase. Results:The methanolic fruit extract exhibited the highest phenolic content, flavonoid content and antioxidant capacity, among the other extracts, with the highest DPPH radical scavenging activity at a concentration of 10 μg/mL (IC50: 1.91 μg/mL) and maximum reducing power at a concentration of 100 μg/mL (EC50: 3.58 μg/mL). Though all extract showed moderate antimicrobial activity against the bacterial strains, weak or no activity against fungus. The range of LC50 value of all extracts was 1.335-14.057 μg/mL which was far lower than the cut off index for cytotoxicity. All extracts exhibited statistically significant (P Conclusions:Our study suggested that S. dulcis exhibits antimicrobial activities against a wide variety of strains while it possesses significant antioxidant, cytotoxic and thrombolytic activity.
ABSTRACT
Cassia senna leaves belonging to the family Fabaceae have been investigated for the presence of its secondary metabolites and evaluation of biological activities of the crude extracts with special emphasis to the antimicrobial activity, cytotoxic activity and thrombolytic activity. The antimicrobial activities of n-hexane, chloroform, ethyl acetate & methanolic extracts of C. senna leaves were screened against five gram(+) bacteria, eight gram(-) bacteria and three fungi by ‘disc diffusion method’. The methanol extract possesses no antimicrobial activity but chloroform and n-hexane fractions exhibited moderate to less activity against some organisms tested compared with the standard antibiotic Kanamycin. Brine shrimp lethality bio-assay was done using brine shrimp Nauplii and dimethyl sulfoxide as a solvent for the methanol extracts of C. senna. The LC50 value (1.5625) of methanol extract of the plant indicated that the cytotoxicity was very significant. The percentages found in thrombolytic tests are 41.46%, 53.22%, 33.33%, 4.08% and standard 92.85%. So, in comparison with standard, C. senna can be further use as mild thrombolytic agent.
ABSTRACT
The crude methanolic extract of the bark of Sarcolobus globosus (Family-apocynaceae) and its different organic soluble Kupchan fractions were screened for total phenol content (TPC), cytotoxic, membrane stabilizing and thrombolytic activities. The polyphenol content was determined colorimetrically using Folin-Ciocalteu method and expressed in gallic acid equivalent. The chloroform soluble Kupchan fraction (CSF) exhibited higher level of Total Polyphenol Contents (TPC, 54.21 gm of GAE/100 gm of dried extract). In the brine shrimp lethality bioassay, the crude methanolic extract (MEBP) exhibited significant cytotoxicity. The membrane stabilizing activity was assessed by using erythrocyte in hypotonic solution and was compared with acetyl salicylic acid. The hexane soluble Kupchan fraction (HSF) produced 52.73 percent inhibition of hemolysis of RBC as compared to 65.38 percent revealed by acetyl salicylic acid (0.10 mg/mL). In thrombolytic study screening, the crude methanolic extract demonstrated significant thrombolytic activity in human blood specimen.
El extracto crudo metanólico de la corteza de Sarcolobus globosus (Familia-apocynaceae) y sus diferentes fracciones solubles Kupchan fueron identificadas para contenido total de fenoles (CTF), actividades citotóxicas, estabilizantes de membrana y trombolíticas. El contenido de polifenoles fue determinado colorimétricamente usando el método Folin-Ciocalteu y expresados en equivalentes a ácido gálico. La fracción Kupchan soluble en cloroformo (FSC) exhibió los mayores niveles de Contenido Total de Polifenoles (CFT, 54,21 gm of GAE/100gm de extracto seco). En el bioensayo de letalidad (Artemia salina), el extracto metanólico crudo (EMC) exhibió una siginificativa citotoxicidad. La actividad estabilizadora de membrana fue estimada usando eritrocitos en un medio hipotónico y fue comparado con el ácido acetil salicílico. La fracción Kupchan soluble en hexano (FSH) produjo un 52,73 por ciento de inhibición de la hemólisis de los glóbulos rojos comparado con un 65,38 por ciento revelado por el ácido acetil salicílico (0,1 mg/mL). En las determinaciones trombolíticas, el extracto metanólico crudo demostró una significativa actividad trombolítica en una muestra de sangre humana.
Subject(s)
Apocynaceae/chemistry , Plant Extracts/pharmacology , Plant Extracts/chemistry , Phenols/analysis , Fibrinolytic Agents/pharmacology , Apocynaceae/chemistry , Cell Membrane , Polyphenols/analysisABSTRACT
Objective To express and purify recombinant protease with thrombolysis activity of Perinereis aibuhitensis Grube and study on its characterization.Methods pMAL-PPA was in- troduced into E.coli DH5?to construct engineering bacteria and overexpression of the prote- ase of fused with maltose binding protein(MBP-PPA)was achieved with IPTG induction. The fusion protein was purified by affinity chromatography on amylose-resin column fol- lowed by chromatography on a DEAE-sepharose FF column.PPA cut with Factor Xa was assayed using casein plates supplied with plasminogen.Results Engineering bacteria express- ing maltose binding protein-thrombolytic protease of P.aibuhitensis were constructed and overexpression of MBP-PPA was achieved with IPTG induction.A recombinant fusion pro- tein of 51kD was purified,and PPA cut down from the fusion protein had a plasminogen-acti- rating activity.The protease showed a good thermal stability with an optimal pH of 8.0. This enzyme was also relatively stable in a pH range of 6.0～9.0 and still active after stored in organic solvents for 20d.Conclusion PPA was verified as a plasminogen activator,and might be a new thrombolytic medicine in the future.