Bacillus Mojavensis A21 Research Articles

Economic production and biocontrol efficiency of lipopeptide biosurfactants from Bacillus mojavenis A21.

In the wake of an increased attention on the eco-friendly biopesticidal products and the rising market requirements for organic agents, lipopeptides compounds have been described as biological control agents which improve the overall health growth and development of plants. Nevertheless, their high production cost constitue the major flaw in their wide use to control plant diseases. The present article aims to formulatean economic media for lipopeptides productionby Bacillus mojavensis A21 for application as natural fungicides for plant disease treatment. We herein demonstrated the suitability of the potato waste, as low cost substrate, for lipopeptides production. Moreover, sea water was found to be a good mineral salts sources. In the second part of this study, we investigate the inhibitory activity of A21 lipopeptides against the phtopathogenic Fusarium sp. The in vitro test showed a minimal inhibitory concentration of about 0.3mg/ml. The microscopic examination, of the treated Fusarium revealed an excessive lysis of the mycelia ultrastructure with destructed spores. The invivoantagonist activity wasconfirmedtowardsthe infectedpotato tubers. A21 lipopeptides are effective in decreasing by about 78.26% and 60.68% when applied as preventive and curative treatments, respectively, as compared to the untreated tubers.

Peptidomic analysis of bioactive peptides in zebra blenny (Salaria basilisca) muscle protein hydrolysate exhibiting antimicrobial activity obtained by fermentation with Bacillus mojavensis A21

Abstract The present study investigates the antibacterial activity of zebra blenny (Salaria basilisca) protein hydrolysates obtained by fermentation with a proteolytic bacterium, Bacillus mojavensis A21. The fermentative zebra blenny protein hydrolysate (FZPH), with a degree of hydrolysis (DH) of 17.35%, was fractionated by size exclusion chromatography on a Sephadex G-25 into six major fractions (F1-F6). Fraction F2, which exhibited antibacterial activity against several Gram-positive and Gram-negative bacteria, was further fractionated by reversed-phase high performance liquid chromatography (RP-HPLC). Fractions A and B from RP-HPLC exhibiting the highest antibacterial activity, were analysed using nano ESI-LC–MS/MS to identify the sequences of the peptides. A total of 28 and 41 peptides, containing from 8 to 31 amino acid residues, were identified in sub-fractions A and B, respectively. Further, identified bioactive peptides sharing sequences with previously identified peptides were reported. The results of this study suggest that FZPH is a good source of natural antimicrobial peptides and therefore, they could serve as a beneficial ingredient for nutraceuticals.

Optimization of proteins and minerals removal from shrimp shells to produce highly acetylated chitin

Chitin and derivatives used for biomedical or pharmaceutical applications require a high level of purity and quality that are difficult to achieve. In this study, we propose to optimize the extraction of chitin in order to obtain pure product keeping a structure as close as possible to the native form. Thus, demineralization step was firstly optimized using response surface methodology. In the optimized conditions predicted by the model, the obtained chitin has an acetylation degree (DA) and a demineralization degree (DM) equal to 99% and 100%, respectively.Then, different microbial and fish crude alkaline proteases were tested for their efficiency in deproteinization. Crude alkaline proteases giving the highest deproteinization degrees (DP), Bacillus mojavensis A21 and Scorpaena scrofa, were selected for chitin extraction. The obtained DP was 88±2% and 83±1%, respectively.At the end, effect of the use of mixed enzymatic treatment with the two selected crude enzymes and the order of demineralization/deproteinization steps were tested. The results demonstrated that two separated steps in enzymatic treatments realized on demineralized sample give the best DP (96%) preserving the DA (99%).

Acute and sub-chronic oral toxicity profiles of lipopeptides from Bacillus mojavensis A21 and evaluation of their in vitro anticoagulant activity

The aim of the present study was to evaluate the acute and sub-chronic toxicity of lipopeptides mixture produced by Bacillus mojavensis A21 as well as their in vitro anticoagulant activity. A21 lipopeptides was given to mice at single dose from 75mg to 1000mg/kg body weight (bw). The median lethal dose (LD50) of A21 lipopeptides was about 550mg/kgbw. Sub-chronic toxicity study for 28days was done by daily oral administration of A21 lipopeptides at doses of 40 and 400mg/kgbw in rats. Results showed that A21 lipopeptides did not cause any change in body weights and they did not produce any marked alterations in the hematological blood parameters including hematocrit concentration, hemoglobin level, white and red cells count. However, the platelets level decreased significantly compared to control value. Moreover, no significant differences in the serum biochemical characteristics were observed for rats treated by the lowest dose. In contrast, a little enhancement of alanine-aminotransferase (ALT) activity and decrease in total cholesterol were observed with the highest dose. A21 lipopeptides were also found to cause a prolongation of the thrombin time (TT), the prothrombin time (PT) and the activated partial thromboplastin time (APTT). Overall, A21 lipopeptides may be very promising compounds for therapeutic purposes.

Chitin extraction from crab shells by Bacillus bacteria. Biological activities of fermented crab supernatants

Crab shells waste were fermented using six protease-producing Bacillus species (Bacillus subtilis A26, Bacillus mojavensis A21, Bacillus pumilus A1, Bacillus amyloliquefaciens An6, Bacillus licheniformis NH1 and Bacillus cereus BG1) for the production of chitin and fermented-crab supernatants (FCSs). In medium containing only crab shells, the highest demineralization DM was obtained with B. licheniformis NH1 (83±0.5%) and B. pumilus A1 (80±0.6%), while the highest deproteinization (DP) was achieved with A1 (94±1%) followed by NH1 (90±1.5%) strains. Cultures conducted in medium containing crab shells waste supplemented with 5% (w/v) glucose, were found to remarkably promote demineralization efficiency, and enhance slightly deproteinization rates. FTIR spectra of chitins showed the characteristics bands of α-chitin.FCSs showed varying degrees of antioxidant activities which were in a dose-dependent manner (p<0.01). In fact, FCS produced by B. amyloliquefaciens An6 exhibited the highest DPPH free radical-scavenging activity (92% at 4mg/ml), while the lowest hydroxyl radical-scavenging activity (60% at 4mg/ml) was obtained with B. subtilis A26 hydrolysates. However, the highest reducing power (OD700nm=2 at 0.5mg/ml) was obtained by B.amyloliquefaciens An6 hydrolysates. These results suggest that crab hydrolysates are good sources of natural antioxidants. Further, FCSs were found to exhibit antibacterial activity against Gram-positive and Gram-negative bacteria.

Wound healing and in vitro antioxidant activities of lipopeptides mixture produced by Bacillus mojavensis A21

The aim of the present study was to evaluate the in vitro antioxidant activities and the in vivo wound healing performance of lipopeptides produced by Bacillus mojavensis A21. The in vitro antioxidant activity was determined through four different assays: DPPH scavenging activity, reducing power, β-carotene bleaching by linoleic acid assay and lipid peroxidation inhibition activity. The scavenging effect of A21 lipopeptides on 1, 1-diphenyl-2-picrylhydrazyl (DPPH) radical at 1g/L was 65%. Moreover A21 lipopeptides showed good reducing power and significantly inhibited lipid peroxidation. The application of A21 lipopeptides gel on the wound site in a rat model accelerated significantly wound healing activity as compared to the control group, and a total closure was achieved after 13 days of wound induction. Further, histological study of biopsies treated with A21 lipopeptides showed fully re-epithelialized wound with a complete epidermal regeneration.

Identification and biochemical characteristics of lipopeptides from Bacillus mojavensis A21

This study reports the potential of a marine bacterium, Bacillus mojavensis A21, to produce lipopeptide biosurfactants. The crude lipopeptide mixture was found to be very effective in reducing surface tension to 31mNm−1. PCR experiments using degenerate primers revealed the presence of nonribosomal peptide synthetases genes implied in the biosyntheses of fengycin and surfactin. Matrix-Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF-MS) performed on whole cells of B. mojavensis A21 confirmed the presence of lipopeptides identified as members of surfactin and fengycin families. Further, a detailed analysis performed by MALDI-TOF-TOF revealed the presence of pumilacidin compounds. The crude lipopeptide mixture was tested for its inhibitory activity against Gram-positive and Gram-negative bacteria, and fungal strains. It was found to display significant antimicrobial activity. Strain A21 lipopeptide mixture was insensitive to proteolytic enzymes, stable between pH 3.0 and 11.0, and resistant to high temperature. Production of lipopeptides is a characteristic of several Bacillus species, but to our knowledge this is the first report involving identification of pumilacidin, surfactin and fengycin isoforms in a B. mojavensis strain.

Chitin extraction from shrimp shell using enzymatic treatment. Antitumor, antioxidant and antimicrobial activities of chitosan

Chitin was recovered through enzymatic deproteinization of the shrimp processing by-products. Different microbial and fish viscera proteases were tested for their deproteinization efficiency. High levels of protein removal of about 77±3% and 78±2% were recorded using Bacillus mojavensis A21 and Balistes capriscus proteases, respectively, after 3h of hydrolysis at 45°C using an enzyme/substrate ratio of 20U/mg. Therefore, these two crude proteases were used separately for chitin extraction and then chitosan preparation by N-deacetylation.Chitin and chitosan samples were then characterized by 13 Cross polarization magic angle spinning nuclear magnetic resonance (CP/MAS)-NMR spectroscopy and compared to samples prepared through chemical deproteinization. All chitins and chitosans showed identical spectra. Chitosans prepared through enzymatic deproteinization have practically the same acetylation degree but higher molecular weights compared to that obtained through chemical process. Antimicobial, antioxidant and antitumoral activitities of chitosan-M obtained by treatment with A21 proteases and chitosan-C obtained by alkaline treatment were investigated. Results showed that both chitosans inhibited the growth of most Gram-negative, Gram-positive bacteria and fungi tested. Furthermore, both chitosans exhibited antioxidant and antitumor activities which was dependent on the molecular weight.

Characterization and potential use of cuttlefish skin gelatin hydrolysates prepared by different microbial proteases.

Composition, functional properties, and in vitro antioxidant activities of gelatin hydrolysates prepared from cuttlefish skin were investigated. Cuttlefish skin gelatin hydrolysates (CSGHs) were obtained by treatment with crude enzyme preparations from Bacillus licheniformis NH1, Bacillus mojavensis A21, Bacillus subtilis A26, and commercial alcalase. All CSGHs had high protein contents, 74.3–78.3%, and showed excellent solubility (over 90%). CSGH obtained by alcalase demonstrated high antioxidant activities monitored by β-carotene bleaching, DPPH radical scavenging, lipid peroxidation inhibition, and reducing power activity. Its antioxidant activity remained stable or increased in a wide range of pH (1–9), during heating treatment (100°C for 240 min) and after gastrointestinal digestion simulation. In addition, alcalase-CSGH was incorporated into turkey meat sausage to determine its effect on lipid oxidation during 35 days of storage period. At 0.5 mg/g, alcalase-CSGH delayed lipid oxidation monitored by TBARS and conjugated diene up to 10 days compared to vitamin C. The results reveal that CSGHs could be used as food additives possessing both antioxidant activity and functional properties.

Characterization and stability of biosurfactant produced byBacillus mojavensisA21 and its application in enhancing solubility of hydrocarbon

BACKGROUND The present work aimed to characterize the physicochemical properties of the biosurfactant produced by Bacillus mojavensis A21 and to evaluate its potential use in diesel recovery. RESULTS The strain B. mojavensis A21 was found to be a potent producer of biosurfactant. Compared with SDS and Tween 80, biosurfactant A21 showed high physicochemical properties in terms of the surface activities and high emulsification index. The biosurfactant A21 reduced surface tension to 30 mN m-1. Biosurfactant A21 was also demonstrated to be stable in a wide range of pH, temperature and salinity. Moreover, it was found that it can solubilize diesel more effectively than SDS and Tween 80. It can be considered as an alternative to chemically synthesized surfactants since it shows high solubilization efficiency towards diesel oil (62% at 0.5 g L-1) in comparison with SDS and Tween 80. CONCLUSION Biosurfactant A21, owing its high emulsification capacity and its high tolerance to acidic and alkaline pH values and salinity, shows great potential for use in bioremediation processes to enhance the solubility of hydrophobic compounds. © 2013 Society of Chemical Industry

Effect of Degree of Hydrolysis and Protease Type on the Antioxidant Activity of Protein Hydrolysates From Cuttlefish (Sepia officinalis) By-Products

We have investigated the antioxidant activities of eight hydrolysates from cuttlefish by-products obtained by treatment with various gastrointestinal proteases (chymotrypsin, trypsin, and crude alkaline enzyme extracts from cuttlefish and sardinelle) and bacterial proteases (Alcalase and crude enzymes from Bacillus pumilus A1, Bacillus mojavensis A21, and Bacillus cereus BG1). The antioxidant activities of the cuttlefish by-products protein hydrolysates (CPHs) were evaluated using various in vitro antioxidant assays, such as 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical-scavenging activity, reducing power, and total antioxidant capacity. All hydrolysates showed different degrees of hydrolysis (DH) and varying degrees of antioxidant activity. Among the different hydrolysates, cuttlefish crude enzyme hydrolysate exhibited the highest antioxidant activity, followed by sardinelle crude enzyme and Alcalase hydrolysates. Further, CPHs with different degrees of hydrolysis were prepared by treatment with proteolytic enzymes from cuttlefish, sardinelle, and B. mojavensis A21. All hydrolysates showed a greater antioxidative activity as indicated by all the methods considered. In addition, antioxidant activity in hydrolysates was positively correlated with the increase of DH. The results of this study indicated that CPHs might be a good candidate for further investigation in developing new antioxidants.

English

The media containing new complex organic substrates suggested that&nbsp;Mirabilis jalapatuber powder (MJTP), shrimp wastes powder (SWP)&nbsp;and sardinella peptone could be used as an alternative source of cheaper carbon and nitrogen substrates for production of enzymes by&nbsp;Bacillus mojavensis&nbsp;A21. The use of 40 g/L MJTP as a sole carbon and nitrogen sources resulted in a proteases&nbsp;and &alpha;-amylases&nbsp;production of 730 and 3 U/ml, respectively. A two fold higher production of proteases by A21 strain was achieved with 4 g/L of sardinella peptones added in the MJTP medium. More interestingly, SWP supplementation&nbsp;in a medium containing of MJTP&nbsp;resulted in maximum growth and highest &alpha;-amylase production. &nbsp; Key words:&nbsp;Complex organic substrates, proteases,&nbsp;&alpha;-Amylases,&nbsp;Bacillus mojavensisA21.

Composition, functional properties and in vitro antioxidant activity of protein hydrolysates prepared from sardinelle (Sardinella aurita) muscle.

Composition, functional properties and in vitro antioxidative activities of protein hydrolysates prepared from muscle of sardinelle (Sardinella aurita) were investigated. Sardinelle protein hydrolysates (SPH) were obtained by treatment with crude enzyme preparations from Bacillus pumilus A1 (SPHA1), Bacillus mojavensis A21 (SPHA21) and crude enzyme extract from sardinelle (Sardinella aurita) viscera (SPHEE). The protein hydrolysates SPHA1, SPHA21 and SPHEE contained high protein content 79.1%, 78.25% and 74.37%, respectively. The protein hydrolysates had an excellent solubility and possessed interfacial properties, which were governed by their concentrations. The antioxidant activities of protein hydrolysates at different concentrations were evaluated using various in vitro antioxidant assays, including 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical method, reducing power assay, chelating activity, β-carotene bleaching and DNA nicking assay. All protein hydrolysates showed varying degrees of antioxidant activity. SPHA21 had the highest DPPH radical scavenging activity (89% at 6mg/ml) and higher ability to prevent bleaching of β-carotene than SPHA1 and SPHEE (p < 0.05). However, SPHEE exhibited the highest metal chelating activity (89% at 1mg/ml) and the strongest protection against hydroxyl radical induced DNA breakage (p < 0.05).

Effect of protein hydrolysates from sardinelle (Sardinella aurita) on the oxidative status and blood lipid profile of cholesterol-fed rats

This study was designed to test the hypolipidemic properties and antioxidative activities of sardinelle (Sardinella aurita) protein hydrolysates (SPHs) obtained by treatment with crude enzyme preparations from Bacillus pumilus A1(SPHA1), Bacillus mojavensis A21(SPHA21) and crude enzyme extract from sardinelle viscera (SPHEE).Wistar rats were fed during 7weeks a standard laboratory diet, a cholesterol-enriched diet (1%) or a cholesterol SPH-enriched diet. The hypercholesterolemic diet induced the elevation of total cholesterol (TC), triglycerides (TG), and low-density lipoprotein cholesterol (LDL-C). Supplementing cholesterol-enriched diet with SPHs or whole sardinelle protein (WSP) at a concentration of 5% (w/w) increased the serum level of high-density lipoprotein cholesterol (HDL-C) and HDL-C/TC ratio and decreased the serum levels of TC, TG, LDL-C and LDL-C/HDL-C ratio significantly.The thiobarbituric acid-reactive substances (TBARS) level, as an indicator of lipid peroxidation, and the activity of superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase (CAT) were examined. The hepatic antioxidant enzymes activities were significantly decreased and the malondialdehyde (MDA) level was increased in rats fed a cholesterol-enriched diet compared to those fed a standard diet. The treatment of hypercholesterolemic (HCD) diet rats with SPHs reduced the MDA concentration and increased the antioxidant enzyme activities.These results suggested that the hypolipidemic effect of SPHs might be due to their abilities to lower serum TC, TG, and LDL-C levels as well as to their antioxidant activities preventing the lipid peroxidation process.

Improvement of functional properties and antioxidant activities of cuttlefish ( Sepia officinalis) muscle proteins hydrolyzed by Bacillus mojavensis A21 proteases

Functional properties and antioxidant activities of cuttlefish ( Sepia officinalis) muscle protein hydrolysates, with different degrees of hydrolysis (DH from 7.3% to 18.8%), obtained by treatment with Bacillus mojavensis A21 alkaline proteases were investigated. Protein contents for all freeze-dried cuttlefish muscle protein hydrolysates (CMPHs) ranged from 80% to 86%. For the functional properties, hydrolysis by A21 proteases increased ( p < 0.05) protein solubility to above 78% over a wide pH range (2.0–11.0). However, the interfacial activities (emulsion activity index, emulsion stability index, foaming capacity and foaming stability) decreased with the increase of the DH. All CMPHs exhibited significant metal chelating activity and DPPH free radical-scavenging activity, and inhibited linoleic acid peroxidation. Antioxidant properties of protein hydrolysates increased with protein hydrolysis and the highest activities were obtained at DH of 16%. The IC 50 values for DPPH radical-scavenging and metal chelating activities were found to be 0.52 ± 0.01 mg/ml and 0.67 ± 0.13 mg/ml. The obtained results suggested that functional properties and antioxidant activities of cuttlefish muscle protein hydrolysates were influenced by the degree of hydrolysis. The composition of amino acids of undigested and hydrolyzed proteins was determined. CMPHs have a high percentage of essential amino acids such as arginine, lysine, histidine and leucine. They have a high nutritional value and could be used as supplement to poorly balanced dietary proteins.

A novel surfactant-stable alkaline serine-protease from a newly isolated Bacillus mojavensis A21. Purification and characterization

An extracellular bleach stable protease producing strain was isolated from marine water sample and identified as Bacillus mojavensis A21 on the basis of the 16S rRNA gene sequencing and biochemical properties. The A21 alkaline protease was purified from the culture supernatant to homogeneity using acetone precipitation, Sephadex G-75 gel filtration and CM-Sepharose ion exchange chromatography, with a 6.43-fold increase in specific activity and 16.56% recovery. The molecular weight of the purified enzyme was estimated to be 20 kDa by SDS-PAGE and gel filtration. The enzyme was highly active over a wide range of pH from 7.0 to 13.0, with an optimum at pH 8.5. The relative activities at pH 11.0 and 12.0 were about 80 and 71.7% of that obtained at pH 8.5. The enzyme was extremely stable in the pH range of 7.0–12.0. It exhibited maximal activity at 60 °C. The thermostability of the enzyme was significantly increased by the addition of CaCl 2. The activity of the enzyme was totally lost in the presence of PMSF, suggesting that the purified enzyme is a serine protease. The N-terminal amino acid sequence of the first 20 amino acids of the purified protease was DINGGGATLPQKLYQTSGVL. B. mojavensis A21 protease showed low homology with bacterial peptidases, suggesting that the enzyme is a new protease. The alkaline protease showed high stability towards anionic (0.1% SDS) and non-ionic (1 and 5% Tween 80 and 1% Triton X-100) surfactants. In addition, the enzyme was relatively stable towards oxidizing agents, retaining more than 79 and 70% of its initial activity after 1 h incubation in the presence of 1% H 2O 2 and 0.1% sodium perborate, respectively. The enzyme showed excellent stability with a wide range of commercial solid and liquid detergents at 30 and 40 °C. Considering its promising properties, B. mojavensis A21 may find potential application in laundry detergents.