Streptokinase (SK) is an extracellular enzyme secreted by various strains of β-hemolytic Streptococci. The main focus of the current study is to evaluate the in vitro thrombolytic activity of purified SK extracted from Streptococcus equinus VIT_VB2 (Accession no. JX406835) isolated from milk sample. The growth rate of S. equinus VIT_VB2 strain was studied with pH and biomass content which has positive significant effect on enzyme yield. A temperature of 10 °C and pH of 6 was found to be optimum for maximum SK activity. The specific activity of the purified SK produced by VIT_VB2 strain was found to be 6,585 IU mg(-1). The molecular mass of the enzyme was determined as 47 kDa by SDS-PAGE. In vitro thrombolytic activity of purified SK was determined using synthetic chromogenic substrate S-2251, the activity of the purified enzyme was found to be 6,330 ± 2.2 IU. The purity of SK was compared with standard SK by HPLC. This is the first report which reveals the SK activity of S. equinus isolated from milk sample.

Babu S

Department of Biotechnology

School of Bio Sciences and Technology

Vellore Campus

Subathradevi C

Vellore Institute of Technology (VIT) is a private university located in&nbsp;Tamil Nadu, India. Founded in 1984, as Vellore Engineering College, the institution offers 20 undergraduate, 34 postgraduate, four integrated and four research programs. It has campuses in Vellore, Amravati, Bhopal and Chennai.

VIT is one of the top ranked private universities in India according to NIRF, THE and QS Rankings.&nbsp;Govt. of India has recognized&nbsp;VIT, Vellore as an&nbsp;Institution of Eminence. This has allowed VIT to take independent quality initiatives and move up in world ranking.

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VIT University

Journal of Thrombosis and Thrombolysis

Streptokinase purified from Streptococcus equinus VIT_VB2 isolated from bovine milk sample was immobilized in various solid supports namely entrapment in agarose gel, calcium alginate beads and gelatin gel by cross-linking with formaldehyde. Immobilization of streptokinase in calcium alginate beads showed maximum efficiency (81.8 ± 1.06%) when compared with entrapment with agarose gel (55.6 ± 2.17%) and cross-linked gelatin formaldehyde gel (71.0 ± 1.54%). The purified SK activity was expressed maximum in calcium alginate (1%) and gelatin gel (0.25%) with 1292.68 ± 1.33 and 1121.9 ± 1.2 U mL−1, respectively. Similarly, SK entrapped in gelatin gel and calcium alginate showed maximum in vitro blood clot lysis activity with 77.67 ± 2.64% and 76.16 ± 2.72%, respectively. The immobilized SK in gelatin gel showed complete clot lysis within 15 min; hence, this application of the study could be used in the treatment of superficial thrombophlebitis, phlebitis, and venous thrombosis. These beads were used for three repeated cycles to check the conversion of substrates into their products, and we concluded that SK can be immobilized in the suitable matrices. Therefore, this helps in the drug-delivery strategies in highly efficient way, moreover, economically competent process in the pharmaceutics. © 2017 Taylor &amp; Francis.

Preparative Biochemistry and Biotechnology

Potential application of immobilized streptokinase extracted from Streptococcus equinus VIT_VB2

Herbal preparations have been used since ancient times for the treatment of several diseases and constitute a major share of all the officially recognized systems of health in India. The aim of the present study was to assess the in vitro thrombolytic activity of leaf extracts from Leucas aspera, Cynodon dactylon and Murraya koenigii in combination with clot buster enzymes (i.e., Nattokinase-NK and Staphylokinase-SaK). Methanolic leaf extract of L. aspera, C. dactylon and aqueous leaf extract of M. koenigii were prepared and assessed for their thrombolytic activity by clot lysis assay. The average clot lysis activity of L. aspera, C. dactylon and M. koenigii were found to be 45.32±0.53%, 38.56±0.42% and 30.32±0.25%. Combination of clot buster enzymes and leaf extracts showed an maximum clot lysis activity at the proportional ratio 1:1 of NK:L. aspera (47.61±0.35%), NK:C. dactylon (55.88±0.52%), SaK:L. aspera (59.04±0.54%) and SaK:C. dactylon (50.75±0.56%). GC–MS analysis of L. aspera, C. dactylon and M. koenigii extracts revealed the presence of thrombolytic compounds. This is the first report on combined thrombolytic activity of clot buster enzymes and selected herbal extracts. Hence this study demonstrates the combinational properties of the herbal extracts and clot buster enzymes which could be one of the best sources of herbal drugs for thrombotic disorders with minimal side effects. Advances in this research could lead to the development of new combination of thrombolytics. © 2016 Elsevier Ltd

Biocatalysis and Agricultural Biotechnology

Combination of clot buster enzymes and herbal extracts: A new alternative for thrombolytic drugs

In vitro thrombolytic activity of purified streptokinase extracted from Streptococcus equinus VIT_VB2 isolated from bovine milk

The aim of this study was to isolate and characterize streptokinase-producing β-hemolytic Streptococcus sp. from bovine milk. A total of 50 milk samples were collected randomly from different breeds of cow and goat (Vellore, Tamil Nadu, India). The samples were characterized and screened for streptokinase-producing isolates using microbial and biochemical analysis. About 97 colonies were isolated from milk samples showing hemolytic patterns of α (19.6%), β (24.7%) and γ (55.6%). Out of 20β-hemolytic isolates, only 6 colonies (VB2, VB3, VB8, VB14, VB16, and VB17) were identified as β-hemolytic Streptococci as potent producers of streptokinase. VB2 and VB14 showed the greatest streptokinase activities of 265 U mL-1 and 225 U mL-1, respectively. Based on biochemical and molecular characterization, the potent isolates VB2 and VB14 were identified and confirmed as S. equinus and S. agalactiae, respectively. The identified strains were named Streptococcus equinus VIT_VB2 (GenBank accession no. JX406835) and Streptococcus agalactiae VITVS5 (GenBank accession No. KF186620) The strains isolated from bovine milk provide a variance in the fibrinolytic activity on blood clots. The current study has demonstrated that the isolation of streptokinase producers from bovine milk, and the production of streptokinase from novel strain, enhanced the fibrinolytic activity. This study is the first to report that Streptococcus equinus produces streptokinase. © 2015 Applied Microbiology, Molecular and Cellular Biosciences Research Foundation.

Fulltext

The Journal of General and Applied Microbiology

Exploring the in vitro thrombolytic potential of streptokinase-producing β-hemolytic Streptococci isolated from bovine milk

In the present study, Saccharopolyspora erythraea MTCC 1103 was used for the enhanced production of erythromycin. To enhance the yield of erythromycin, effects of various parameters such as bagasse concentration, organic nitrogen source, inorganic nitrogen source, pH and temperature were analysed. It was found that bagasse can be used as an alternate carbon source in erythromycin production medium. Erythromycin production in the new formulation of bagasse based medium was found to be 512 mg/L which was 28 % higher than glucose based medium. Strain improvement was done by random UV-mutagenesis. When compared to wild type strain, mutant strain showed 40 % higher yield in production medium. Erythromycin potency assay and HPLC analysis were performed to confirm the presence of erythromycin in the partially purified samples. These optimized conditions could be used for the commercial production of this unique antibiotic which gave significant industrial perspectives. © 2013, The Author(s).

3 Biotech

Strain improvement and optimization studies for enhanced production of erythromycin in bagasse based medium using Saccharopolyspora erythraea MTCC 1103

Journal	Data powered by TypesetJournal of Thrombosis and Thrombolysis
Publisher	Data powered by TypesetSpringer Science and Business Media LLC
ISSN	0929-5305
Open Access	No