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L-asparaginase is an important enzyme as therapeutic agents used in combination with other drugs in the treatment of acute lymphoblastic leukemia. A newly isolated actinomycetes strain, Streptomyces sp. NEAE-82, was potentially producing extracellular L-asparaginase, it was identified as Streptomyces fradiae NEAE-82, sequencing product was deposited in the GenBank database under accession number KJ467538. L-asparaginase was purified from the crude enzyme using ammonium sulfate precipitation, dialysis and ion exchange chromatography using DEAE Sepharose CL-6B. Further the kinetic studies of purified enzyme were carried out. The optimum pH, temperature and incubation time for maximum L-asparaginase activity were found to be 8.5, 40 °C and 30 min, respectively. The optimum substrate concentration was found to be 0.06 M. The Km and Vmax of the enzyme were 0.01007 M and 95.08 Uml−1min−1, respectively. The half-life time (T1/2) was 184.91 min at 50 °С, while being 179.53 min at 60 °С. The molecular weight of the subunits of L-asparaginase was found to be approximately 53 kDa by SDS–PAGE analysis. The purified L-asparaginase showed a final specific activity of 30.636 U/mg protein and was purified 3.338-fold. The present work for the first time reported more information in the production, purification and characterization of L-asparaginase produced by newly isolated actinomycetes Streptomyces fradiae NEAE-82.

BackgroundL-asparaginase is a potential therapeutic enzyme widely used in the chemotherapy protocols of pediatric and adult patients with acute lymphoblastic leukemia. However, its use has been limited by a high rate of hypersensitivity in the long-term used. Hence, there is a continuing need to search for other L-asparaginase sources capable of producing an enzyme with less adverse effects.MethodsProduction of extracellular L-asparaginase by Streptomyces brollosae NEAE-115 was carried out using submerged fermentation. L-asparaginase was purified by ammonium sulphate precipitation and pure enzyme was reached using ion-exchange chromatography, followed by enzyme characterization. Anticancer activity towards Ehrlich Ascites Carcinoma (EAC) cells was investigated in female Swiss albino mice by determination of tumor size and the degree of tumor growth inhibition. The levels of anti-L-asparaginase IgG antibodies in mice sera were measured using ELISA method.ResultsThe purified L-asparaginase showed a total activity of 795.152 with specific activity of 76.671 U/mg protein and 7.835 − purification fold. The enzyme purity was confirmed by using SDS–PAGE separation which revealed only one distinctive band with a molecular weight of 67 KDa. The enzyme showed maximum activity at pH 8.5, optimum temperature of 37 °C, incubation time of 50 min and optimum substrate concentration of 7 mM. A Michaelis-Menten constant analysis showed a Km value of 2.139 × 10− 3 M with L-asparagine as substrate and Vmax of 152.6 UmL− 1 min− 1. The half-life time (T1/2) was 65.02 min at 50°С, while being 62.65 min at 60°С. Furthermore, mice treated with Streptomyces brollosae NEAE-115 L-asparaginase showed higher cytotoxic effect (79% tumor growth inhibition) when compared to commercial L-asparaginase group (67% tumor growth inhibition).ConclusionsThe study reveals the excellent property of this enzyme which makes it highly valuable for development of chemotherapeutic drug.

Zein constitutes about half of the endosperm proteins in corn. Recently, attempts have been made to utilize zein for food coatings and biodegradable materials, which require better physical properties, using chemical modification of zein. In this study, zein proteins were modified using citric acid, succinic anhydride, and eugenol as natural cross-linking agents in the wet state. The cross-linkers were added either separately or combined in increment concentrations (0.1, 0.2, 0.3, and 0.4%). The effects of those agents on the mechanical properties, microstructure, optical properties, infrared (IR) spectroscopy, and antibacterial activities of zein were investigated. The addition of cross-linking agents promoted changes in the arrangement of groups in zein film-forming particles. Regarding the film properties, incorporation of cross-linking agents into zein films prepared in ethanol resulted in two- to three-fold increases in tensile strength (TS) values. According to the Fourier-transform infrared (FTIR) spectra and Hunter parameters there were no remarkable changes in the structure and color of zein films. Transparency of zein films was decreased differentially according to the type and cross-linker concentration. The mechanical and optical properties of zein films were closely related to their microstructure. All cross-linked films showed remarkable antibacterial activities against Bacillus cereus ATCC 49064 and Salmonella enterica ATCC 25566. Food spoilage and pathogenic bacteria were affected in a film-dependent manner. Our experimental results show that even with partial cross-linking the mechanical properties and antipathogen activities of zein films were significantly improved, which would be useful for various industrial applications.

Lactobacillus acidophilus DSM 20079 is the producer of a novel bacteriocin termed acidocin D20079. In this paper, mode of action using three various concentrations of acidocin D20079 (2,048, 128 and 11.3 AU/ml) was determined against an indicator strain L. delbrueckii subsp. lactis DSM 20076. These concentrations all led to marked decreases in both the number of viable cells and in optical density, indicating that the activity of the acidocin D20079 was bactericidal with concomitant cell lysis. Moreover, the probiotic potential of L. acidophilus DSM 20079 was analyzed for its ability to survive and retain viability at conditions (acid and bile concentrations) mimicking the gastrointestinal (GI) tract, under which it survived exposure to pH 2.0 with a 1.2 log cycle reduction in viability and where 45% of the original population survived in a medium containing 0.3% bile for 3 h.

Pediococcus parvulus 2.6 (previously Pediococcus damnosus 2.6, here confirmed as P. parvulus by 16S DNA sequencing) displayed antibacterial activity toward several bacterial species, including isolates found as contaminants in oats, herein genetically identified as Bacillus cereus. No inhibition of Listeria monocytogenes was found under the conditions used. Antibacterial activity was retrieved after ammonium sulfate or acetone precipitation showed it to be peptide mediated. P. parvulus 2.6 has previously shown good technological properties in oat-based products. This, together with the currently found inhibition of food spoilage microorganisms like B. cereus, makes it suitable as a food protective culture. Survival trials of P. parvulus 2.6 at conditions mimicking the gastrointestinal tract were prompted by previously found cholesterol-lowering effects in humans after consumption of oat products cofermented by using P. parvulus 2.6 and Bifidobacterium spp. Viability was measured with in vitro, gutlike simulations at 37 degrees C. High survival was shown under two of three conditions (gastric juice, bile, and small intestine juice), defined as main obstacles of the gastrointestinal tract. The critical step was bile exposure. At a concentration of 20%, viability was low, but 0.3% bile (mean concentration in the intestine) did not have a major influence on growth. Viability of P. parvulus 2.6 was significantly decreased in gastric juice at pH 1.5 (with pepsin), but it was not significantly affected at pH 2.5, and was also improved at a lower pH in 20% oat milk. Viability was judged sufficient for colonization at gutlike conditions, qualifying the strain for further probiotic studies.