scholarly journals Thrombolytic Activity of Alkaline Protease Purified from a Mutant Strain Bacillus licheniformis MZK05M9

2018 ◽  
Vol 21 (1) ◽  
pp. 63-70
Author(s):  
Md Asad Uz Zaman ◽  
Taqiyah Akhtar ◽  
ATM Zafrul Azam ◽  
Md Arafat Al Mamun ◽  
Md Mozammel Hoq ◽  
...  
Keyword(s):  
Mutant Strain ◽  
Bacillus Licheniformis ◽  
Serine Proteases ◽  
Specific Activity ◽  
Pharmaceutical Journal ◽  
Crude Enzyme ◽  
Clot Lysis ◽  
Bacillus Species ◽  
Thrombolytic Activity

Investigations were performed to find out new microbial enzymes as thrombolytics having better efficacy and specificity. Mutant strain of Bacillus species, B. licheniformis MZK05M9 was cultured in modified urea-glucose media followed by purification using ammonium sulphate precipitation and ultrafiltration through centricon tube of specific MWCO value. The production method yielded 823.42 units/mg of the crude enzyme from mutant strain MZK05M9 and after purification 37695.64 units/mg. The molecular weight of the purified enzyme was estimated as 27.2 kDa and purification increased its specific activity to 16.5 fold with a recovery of 10%. The purified proteases were identified as serine proteases by irreversible inhibition of activity with phenylmethylsulfonyl fluoride (PMSF) and it exhibited 32.84% thrombolytic activity, by in vitro clot lysis assay. Stability studies showed that crude enzyme from mutant strain MZK05M9 remained stable up to a temperature of 45˚C and showed maximum stability at pH range 7.5 to 8.5. Our observation indicates that proteases produced by Bacillus licheniformis mutant have the potential to be developed as a viable thrombolytic agent.Bangladesh Pharmaceutical Journal 21(1): 63-70, 2018

scholarly journals Partial Purification of Alkaline Protease as Thrombolytic Agent from Mutant Strain Bacillus licheniformis EMS250-O-1

2017 ◽  
Vol 15 (2) ◽  
pp. 135-141 ◽  
Author(s):  
Md Asad Uz Zaman ◽  
Md Arafat Al Mamun ◽  
Shakila Nargis Khan ◽  
Md Mozammel Hoq ◽  
Md Abdul Mazid
Keyword(s):  
Mutant Strain ◽  
Bacillus Licheniformis ◽  
Specific Activity ◽  
Cardiovascular Complications ◽  
Clot Lysis ◽  
Partial Purification ◽  
Thrombolytic Activity ◽  
Ammonium Sulphate Precipitation ◽  
Maximum Stability

Thrombosis leads to myocardial infarction, stroke and other cardiovascular complications. Microbial thrombolytic agents such as urokinase, streptokinase etc. are used to treat complications related to thrombosis. To search for new microbial enzymes as thrombolytics having better efficacy and specificity, Bacillus licheniformis EMS-O-1 mutant strain was cultured in modified urea-molasses media followed by purification using ammonium sulphate precipitation and ultrafiltration through centricon tube of 100 MWCO value. The yield of crude enzyme was 11129.14 U/mg and after purification 40180.46 U/mg. Purification process increased the specific activity of purified enzyme to 12.28 fold with a recovery of 17.79%. The purified enzyme was a serine protease with molecular weight of 25.5 kDa as confirmed by irreversible inhibition of activity with phenylmethylsulfonyl fluoride (PMSF) followed by SDS-PAGE gel image and by LC-MS analyses. In vitro clot lysis assay of the purified enzyme exhibited 38.30% thrombolytic activity. The crude enzymes from the mutant strain EMS-O-1 were found to be stable up to 50oC and showed maximum stability between pH range 7.5 to 8.5. These findings signify that proteases produced by B. licheniformis mutant have the potential to be developed as a viable thrombolytic agent.Dhaka Univ. J. Pharm. Sci. 15(2): 135-141, 2016 (December)

scholarly journals In vitro Antioxidant, Membrane Stabilizing and Thrombolytic Activities of Glycosmis arborea

2012 ◽  
Vol 15 (2) ◽  
pp. 141-143 ◽  
Author(s):  
Sayed Mamur Hossain ◽  
Farhana Islam ◽  
Tasnuva Sharmin ◽  
Hasib Sheikh ◽  
AM Rashedul Hasan ◽  
...  
Keyword(s):  
Carbon Tetrachloride ◽  
Methanol Extract ◽  
Soluble Fraction ◽  
Pharmaceutical Journal ◽  
Clot Lysis ◽  
Hypotonic Solution ◽  
Rat Erythrocytes ◽  
Thrombolytic Activity ◽  
Negative Controls

The methanol extract of leaves and stems of Glycosmis arborea and their pet-ether, carbon tetrachloride, chloroform and aqueous soluble partitionates were subjected to assays for antioxidant activity by Folin-Ciocalteau reagent, membrane stabilizing and thrombolytic activities. The carbon tetrachloride soluble fraction of stems of G. arborea demonstrated the presence of significant amount of phenolic compounds (36.95 ± 0.54 mg of GAE/g of extract). The extractives inhibited heat as well as hypotonic solution-induced haemolysis of rat erythrocytes in vitro. The pet-ether soluble fraction of leaves and stems of G. arborea showed 20.46 and 38.24% and 22.50 and 48.25%. inhibition of hemolysis of RBC caused by heat and hypotonic solution, as compared to 30.55 and 72.91% inhibition of hemolysis of RBC caused by the standard Acetyl salicylic acid at 0.01 mg/ml concentration, respectively. Among the four fractions, the chloroform soluble materials of the stems of G. arborea revealed highest thrombolytic activity with clot lysis value of 36.50% while standard streptokinase and water used as positive and negative controls, showed 64.25±0.26 and 2.35±0.35 % lysis of clot respectively DOI: http://dx.doi.org/10.3329/bpj.v15i2.12579 Bangladesh Pharmaceutical Journal 15(2): 141-143, 2012

scholarly journals Thrombolytic Activity of Methanolic Extracts of Desmodium paniculatum (L.) and Sarcochlamys pulcherrima (Roxb.)

2015 ◽  
Vol 17 (1) ◽  
pp. 67-69 ◽  
Author(s):  
Mohammed Aktar Sayeed ◽  
Humayun Kabir ◽  
Mohammad Mamun Ur Rashid ◽  
Md Farid Ahamad Bhuiyan ◽  
Mohammad A Rashid
Keyword(s):  
Pharmaceutical Journal ◽  
Positive Control ◽  
Negative Control ◽  
Clot Lysis ◽  
Herbal Extracts ◽  
Thrombolytic Activity ◽  
Methanolic Extracts ◽  
Desmodium Paniculatum

An in vitro thrombolytic model was used to check the clot lysis effect of two herbal extracts viz., Desmodium paniculatum (L.) and Sarchochlamys pulcherrima (Roxb.) by using Streptokinase as positive control and water as negative control. D. paniculatum and S. pulcherrima showed 31.92 ± 8.09% and 36.12 ± 6.81% clot lysis, respectively. From our study we found that D. paniculatum and S. pulcherrima showed significant % of clot lysis effect with reference to Streptokinase (72.54 ± 6.03%). and water (3.48 ± 0.84%). DOI: http://dx.doi.org/10.3329/bpj.v17i1.22318 Bangladesh Pharmaceutical Journal 17(1): 67-69, 2014

scholarly journals IN VITRO STUDY ON PROTEASE AND THROMBOLYTIC ACTIVITY OF AQUEOUS EXTRACT FROM LEUCAS ASPERA (L.) LEAVES

2021 ◽  
pp. 73-76
Author(s):  
NAGAMANI J. E. ◽  
USHA SAH ◽  
RAVINDRANATH H. A.
Keyword(s):  
Molecular Weight ◽  
Aqueous Extract ◽  
Lytic Activity ◽  
Crude Enzyme ◽  
Clot Lysis ◽  
Thrombolytic Activity ◽  
Reference Drug ◽  
Leucas Aspera ◽  
Caseinolytic Activity

Objective: The current study is an attempt to screen for the in vitro clot lysis and proteolytic activity of aqueous extract of Leucas aspera leaves. Methods: Thrombolytic activity and protease activity of the crude enzyme obtained by ammonium sulfate precipitation and dialysis were assayed using blood clot and casein as substrates respectively. Native PAGE and gel documentation studies were performed to calculate the molecular weight of the enzyme. Results: In the study, 40% salt fractioned crude enzyme sample exhibited significant thrombolytic and caseinolytic activity. Further dose-dependent increased activity was observed with the maximum lytic activity of 52.11±1.04 % at 1 mg/ml of the sample when compared to the reference drug streptokinase (71.39±0.32%). Also, 68.72±0.62 U/hr of caseinolytic activity was observed for 1 mg/ml of the sample fraction. Conclusion: The study highlights and validates the efficacy of Leucas aspera leaves extract for thrombolytic and proteolytic actions. Enzyme with an approximate molecular weight, 19.89 KDa could be responsible for the significant lytic activity.

scholarly journals In vitro Cytotoxic, Membrane Stabilizing and Thrombolytic Activities of Polygonum glabrum Willd

2015 ◽  
Vol 17 (2) ◽  
pp. 202-204 ◽  
Author(s):  
Mohammad Firoz Khan ◽  
Sikder Nahidul Islam Rabbi ◽  
Fahima Aktar ◽  
Md Hassan Kawsar
Keyword(s):  
Methanol Extract ◽  
Pharmaceutical Journal ◽  
The Other ◽  
Clot Lysis ◽  
Hypotonic Solution ◽  
Activity Assay ◽  
Thrombolytic Activity ◽  
Lc50 Value ◽  
Crude Methanol Extract

The crude methanol extract of leaves of Polygonum glabrum Willd and its Kupchan fractions were screened for cytotoxic, membrane stabilizing and thrombolytic activities. Among all fractions, the crude methanol extract showed significant cytotoxic activity having LC50 value 0.74 ± 0.045 ?g/ml. Moreover, in hypotonic solution- and heat- induced conditions, the crude methanol extract inhibited hemolysis of human erythrocyte by 79.21 ± 0.44% and 84.87±0.23%, respectively as compared to 71.9 ± 0.73% and 42.12 ± 0.37% demonstrated by the standard acetyl salicylic acid. On the other hand, in thrombolytic activity assay the methanol extract demonstrated highest clot lysis value of 35.17 ± 0.42%. DOI: http://dx.doi.org/10.3329/bpj.v17i2.22341 Bangladesh Pharmaceutical Journal 17(2): 202-204, 2014

scholarly journals SYNTHESIS OF BENTONITE NANOCLAY AND INCORPORATION OF CASSIA FISTULA LEAF EXTRACT TO FORM ORGANOBENTONITE: CHARACTERIZATION AND ITS BIOMEDICAL APPLICATIONS

2018 ◽  
Vol 11 (9) ◽  
pp. 381
Author(s):  
Anand Raj Lfa ◽  
Jeslin J
Keyword(s):  
Biomedical Applications ◽  
Leaf Extract ◽  
Spherical Shape ◽  
Xrd Analysis ◽  
Antioxidant Property ◽  
Clot Lysis ◽  
Cassia Fistula ◽  
Thrombolytic Activity ◽  
Lysis Activity

Objective: In this work, methanolic leaf extract from Cassia fistula (known as aragvadha) was incorporated into bentonite nanoclay to form organobentonite. This organobentonite of nanosize was further used for its effective biomedical applications since medicinal clay finds its own advantage over decades.Methods: The bentonite nanoclay was produced by energetic stirring followed by centrifugation and was characterized using scanning electron microscopy (SEM), X-ray diffraction (XRD), and Fourier transform infrared (FTIR). The organobentonite was produced using freeze and thaw method. Antioxidant property was studied using Molyneux method, and thrombolytic activity was analyzed using in vitro clot lysis method.Results: The nanosize of bentonite nanoclay between 57 and 82 nm with irregular to spherical shape was confirmed using SEM analysis. The sharp diffraction peak in XRD analysis shows the crystalline nature of bentonite nanoclay, and FTIR results revealed the successful incorporation of the methanolic extract within the bentonite nanoclay. The organobentonite exhibits 84.5% antioxidant property as well as 31% clot lysis activity when compared to the extract and the bentonite nanoclay.Conclusion: Thus, the non-toxic and inexpensive bentonite nanoclay have a high aspect ratio with multifarious applications in medicine, food, cosmetics, and health products. Through this study, the bentonite nanoclay modified using plant alkaloid (organobentonite) is found to possess good biomedical property.

scholarly journals Novel Fibrinolytic Protease Producing Streptomyces radiopugnans VITSD8 from Marine Sponges

Marine Drugs ◽  
2019 ◽  
Vol 17 (3) ◽  
pp. 164 ◽  
Author(s):  
Dhamodharan D ◽  
Jemimah S ◽  
Merlyn S ◽  
Subathra C
Keyword(s):  
Tamil Nadu ◽  
Specific Activity ◽  
Blood Clot ◽  
Exchange Chromatography ◽  
Marine Sponges ◽  
Protease Production ◽  
Clot Lysis ◽  
Carbon And Nitrogen ◽  
Lysis Activity

Fibrinolytic enzymes have received more attention due to their medicinal potential for thrombolytic diseases. The aim of this study is to characterize the in vitro fibrinolytic nature of purified protease producing Streptomyces radiopugnans VITSD8 from marine brown tube sponges Agelas conifera. Three varieties of sponge were collected from the Rameshwaram Sea coast, Tamil Nadu, India. The fibrinolytic activity of Streptomyces sp. was screened and determined by casein plasminogen plate and fibrin plate methods respectively. The crude caseinolytic protease was purified using ammonium sulfate fractionation, affinity and ion-exchange chromatography. Based on the morphological, biochemical, and molecular characterization, the isolate VITSD8 was confirmed as Streptomyces radiopugnans. Maltose and peptone were found to be the best carbon and nitrogen sources for the production of fibrinolytic protease. The carbon and nitrogen source peptone showed (781 U/mL) enzyme activity. The optimum pH and temperature for fibrinolytic protease production was found to be 7.0 and 33 °C respectively. The purified enzyme showed a maximum specific activity of 3891 U. The blood clot lysis activity was compared with the standard, and it was concluded that a minimum of 0.18 U (10 µL) of purified protease was required to dissolve the blood clot. This is the first report which exploits the fibrinolytic protease activity of Streptomyces radiopugnans VITSD8 extracted from a marine sponge. Hence the investigation suggests a potential benefit of purified fibrinolytic protease which will serve as an excellent clot buster alternative.

scholarly journals In vitro and in vivo evaluation of pharmacological potentials of Campsis radicans L

2019 ◽  
Vol 5 (1) ◽  
Author(s):  
Mirazul Islam ◽  
Tabassum Jannat ◽  
Md. Ruhul Kuddus ◽  
Mohammad Abdur Rashid ◽  
Mohammad Rashedul Haque
Keyword(s):  
Castor Oil ◽  
Methanol Extract ◽  
Radical Scavenging ◽  
Free Radical Scavenging ◽  
Clot Lysis ◽  
Thrombolytic Activity ◽  
Tail Immersion ◽  
Soluble Fractions

Abstract Background Campsis radicans L. is a flowering plant in Bangladesh, traditionally used for the treatment of several human diseases. In this study, in vitro antioxidant, thrombolytic, membrane stabilizing and in vivo analgesic, hypoglycemic, anti-diarrheal and CNS antidepressant activities of organic soluble fractions of crude methanol extract of C. radicans leaf were investigated using appropriate experimental models. Methods The leaves of C. radicans were collected, authenticated, dried and extracted with methanol at room temperature for 30 days. The concentrated methanol extract was partitioned to petroleum-ether (PESF), dichloromethane (DMSF) and ethyl acetate (EASF) soluble fractions. The antioxidant activity of these fractions was determined by DPPH free radical scavenging method. Total phenolic content was determined by the Folin-Ciocalteau’s spectrophotometric method. The thrombolytic activity was assessed by measuring clot lysis ability whereas the membrane stabilizing activity was evaluated by heat- and hypotonic solution-induced hemolysis assay. Tail immersion procedure and acetic acid- induced writhing model were used to measure the analgesic activity of C. radicans. The hypoglycemic, anti-diarrheal and CNS antidepressant activities were determined by oral glucose tolerance test, castor oil-induced diarrheal model and thiopental-sodium induced sleeping time test in mice, respectively. Results All the organic soluble fractions of C. radicans contained phenolic compounds varying from 6.38 to 60.13 mg of GAE/gm of extractive, while in DPPH assay, EASF showed the highest free radical scavenging activity with IC50 is 4.69 μg/ml. The PESF exhibited highest thrombolytic activity (57.14% clot lysis) and the DMSF showed maximum 53.95% inhibition of heat-induced hemolysis of human RBCs. In both tail immersion and acetic acid induced writhing models, the PESF, DMSF, EASF at the doses of 200 and 400 mg/kg body weight, induced a significant (P < 0.001) decrease in the painful sensation in mice. Substantial (P < 0.05) anti-hyperglycemic activity of test samples was found in mice loaded with glucose at the same doses mentioned earlier. Castor oil induced diarrheal test of the plant extractives has shown significant effect in comparison to control group. In CNS antidepressant activity assay, the test samples were able to reduce the duration of sleep in mice caused by thiopental administration. Conclusion All these findings revealed that C. radicans possess significant antioxidant, thrombolytic, membrane stabilizing, analgesic, hypoglycemic, anti-diarrheal and CNS antidepressant activities.

Effect of Standard Heparin and a Low Molecular Weight Heparin on Thrombolytic and Fibrinolytic Activity of Single-Chain Urokinase Plasminogen Activator In Vitro

1989 ◽  
Vol 62 (03) ◽  
pp. 923-926 ◽  
Author(s):  
D M Lourenço ◽  
A M Dosne ◽  
A Kher ◽  
M Samama
Keyword(s):  
Molecular Weight ◽  
Low Molecular Weight Heparin ◽  
Clot Lysis ◽  
Low Molecular Weight ◽  
Single Chain ◽  
Plasma Clot ◽  
Thrombolytic Activity ◽  
Significant Difference ◽  
Plasma Factor

SummaryThe effect of unfractioned heparin (UH) and low molecular weight heparin (LMWH) (Kabi 2165 - Fragmin®) on in vitro scu-PA thrombolytic and fibrinogenolytic activity was investigated. Thrombolytic activity was evaluated by following lysis of radiolabeled plasma clot immersed in plasma in presence of scu-PA alone or with either form of heparin. A 200 IU/ml scu-PA concentration produced clot lysis within 7 hr. UH or LMWH led to a slightly faster clot lysis which was statistically significant only at the 2nd and 3rd hour. No significant difference could be evidenced between UH and LMWH effect. During clot lysis, plasmin, generated within the clot led to a gradual transformation of scu-PA to tcu-PA, specially after a 4-hr incubation. Appearance of tcu-PA activity in the plasma surrounding the clot was significantly inhibited by either form of heparin. This finding contrasts with results observed in purified systems and suggests the presence of heparin-dependent plasma factor(s) inhibiting tcu-PA formation or its activity. Possible candidates might be anti-thrombin III and PAI-3.No fibrinogen breakdown was observed when plasma was incubated for 7 hr at 37° C in presence of scu-PA alone (200 IU/ ml) or with either form of heparin. However, in presence of a plasma clot, an important fibrinogen breakdown was observed during clot lysis reflecting the action of plasmin and/or tcu-PA generated within the clot, in the surrounding plasma. Fibrinogenolysis was less pronounced in the presence of both heparin preparations possibly as a consequence of the reduction in the tcu-PA level. These results underline the importance of plasma factors in the interaction of heparin with plasminogen activators such as scu-PA.

scholarly journals In silico and antithrombotic studies of 1,3,4-oxadiazoles derived from benzimidazole

2015 ◽  
Vol 11 (1) ◽  
pp. 67 ◽  
Author(s):  
B. Vishwanathan ◽  
B. M. Gurupadayya ◽  
K. Venkata Sairam
Keyword(s):  
In Silico ◽  
Factor Xa ◽  
Docking Studies ◽  
Negative Control ◽  
Clot Lysis ◽  
Drug Candidate ◽  
Thrombolytic Activity ◽  
Reference Drug ◽  
Oxadiazole Derivatives

<p class="Abstract">In the present study, a series of 1,3,4-oxadiazole derivatives (4a-4k) derived from benzimidazole were docked onto factor Xa (PDB: 1NFY) protein using SYBYLX 2.1. and also evaluated for <em>in vitro</em> clot lysis for thrombolytic activity. The synthesized molecules were also screened for in silico ADME studies. The molecular docking studies highlighted that the molecules showed high affinity towards 1NFY with higher docking score and the <em>in silico</em> ADME results were promising and indicated that the molecules holds great potential as a drug candidate. The thrombolytic evaluation was performed for decrease in solid clot weight by the clot lysis study at a concentration of 6.25, 12.5 and 25 µM strengths, respectively. The results of in vitro clot lysis for thrombolytic evaluation revealed that the tested compounds 4a-4k exhibited significant clot lysis with respect to negative control phosphate buffered saline and in comparison to the reference drug streptokinase (30,000 IU). Among all the tested compounds, compound 4j, 4d and 4g exhibited potent thrombolytic activity with EC<sub>50</sub> value of 16.2, 18.1 and 23.7 µM, respectively. The thrombolytic efficacy investigation highlights that the synthesized compound 4j could be considered for further clinical studies to ascertain its possible hit as thrombolytic agents.</p><p> </p>

Sign in / Sign up

Export Citation Format

Share Document