Improvement of phenolic profile and biological activities of wild mustard sprouts.

The current study aimed to assess the effect of the germination process of wild mustard seeds on the phenolic profile, antioxidant, antibacterial, and antidiabetic properties, and some relevant enzyme activities. The total phenolic and flavonoid contents increased 5- and 10-fold, respectively, and were maximized on 5-days sprouts. One new phenolic compound was identified on 5-days sprout extract using HPLC. The concentrations of the identified phenolic compounds increased 1.5-4.3 folds on 5-days sprouts compared with dry seeds. The total antioxidant activity multiplied 17- and 21-fold on 5-days sprouts using 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) assays, respectively. The activity of carbohydrate-cleaving, phenolic-synthesizing and antioxidant enzymes also increased during germination. On 5-days sprouts, there was a substantial correlation between the highest ß-glucosidase and peroxidase activities with highest phenolic and flavonoid levels and maximum antioxidant activity. The phenolic extract of 5-days sprouts exhibited antimicrobial activities against Escherichia coli and Staphylococcus aureus and showed potent antidiabetic activity established by its inhibitory effect against α-amylase and α-glucosidase compared to dry seeds.

Development of chia gum/alginate-polymer support for horseradish peroxidase immobilization and its application in phenolic removal.

Chia gum's molecular structure with distinctive properties as well as the alginate-based hydrogel's three-dimensionally cross-linked structure can provide a potent matrix for immobilization of enzyme. Herein, chia gum (CG)/alginate (A)-polymeric complex was synthesized and employed as a support material for the immobilization of horseradish peroxidase (HRP). HRP was successfully immobilized on the developed ACG-polymeric support, and the highest immobilization recovery (75%) was observed at 1.0% CG and 2% A, pH 7.0, and 50 units of the enzyme. The structure, morphology, and thermal properties of the prepared ACG-HRP were demonstrated using Fourier Transform Infrared (FTIR), Scanning Electron Microscope, and Thermogravimetric (TGA) analyses. ACG-HRP showed a good reusability (60%) over ten reuses. The immobilized ACG-HRP displayed an acidic pH optimum (6.0), a higher temperature optimum (50 °C), and improved thermal stability (30-50 °C) compared to the soluble HRP at pH 7.0, 40 °C and (30-40 °C), respectively. ACG-HRP has a lower affinity for hydrogen peroxide (H2O2) and guaiacol and a higher oxidizing affinity for a number of phenolic substrates. The ACG-HRP demonstrated greater resistance to heavy metals, isopropanol, urea, Triton X-100, and urea, as well as improved efficiency for eliminating phenol and p-chlorophenol. The developed ACG-polymeric support provided improved enzyme properties, allowed the reuse of the immobilized HRP in 10 cycles, and made it promising for several biotechnological applications.

Purification and characterization of cysteine protease of Sarcocystis fusiformis from infected Egyptian water buffaloes.

Sarcocystis spp. infects water buffaloes (Bubalus bubalis) causing sarcocystosis. In the present study, Sarcocystis fusiformis was recognized in Egyptian water buffaloes based on histological observation and molecular analysis of internal transcribed spacer 1 (ITS1), 18S ribosomal RNA (18S rRNA) and cytochrome c oxidase subunit I (COX-1) gene fragments. Chemotherapy and vaccines against Sarcocystis spp. could potentially target proteases because they may play a crucial role in the infection. Cysteine proteases are multifunctional enzymes involved in vital metabolic processes. However, the involvement of proteases in S. fusiform infection has not yet been characterized. Here, the purification and study on some biochemical properties of protease isolated from cysts of S. fusiform were carried out. Protease with a molecular weight of 100 kDa was purified. LC-MS/MS analyzed the protein sequence of purified protease and the data suggested that the enzyme might be related to the cysteine protease. The purified protease exhibited maximum activity at pH 6 and a temperature of 50 °C. The Michaelis-Menten constant (Km), the maximum velocity (Vmax), and the turnover number (Kcat) were determined. The complete inhibition effect of cysteine inhibitors indicated that the purified enzyme is a cysteine protease. The results suggested that S. fusiform proteolytic enzyme may be necessary for parasite survival in water buffaloes by digesting host tissues. Therefore, cysteine protease could be a suitable target for vaccinations.

Statistical optimization, characterization, antioxidant and antibacterial properties of silver nanoparticle biosynthesized by saw palmetto seed phenolic extract.

On the global market, silver nanoparticles (Ag-NPs) are in high demand for their various applications in biomedicine, material engineering, and consumer products. This study highlighted the biosynthesis of the Ag-NPs using saw palmetto seed phenolic extract (SPS-phenolic extract), which contained vital antioxidant-phenolic compounds. Herein, central composite statistical design, response surface methodology, and sixteen runs were conducted to optimize Ag-NPs biosynthesis conditions for maximizing the production of Ag-NPs and their phenolic content. The best-produced SPS-Ag-NPs showed a surface plasmon resonance peak at 460 nm and nano-spherical sizes ranging from 11.17 to 38.32 nm using the UV spectrum analysis and TEM images, respectively. The produced SPS-Ag-NPs displayed a high negative zeta-potential value (- 32.8 mV) demonstrating their high stability. The FTIR analysis demonstrated that SPS-phenolic compounds were involved in sliver bio-reduction and in stabilizing, capping, and preventing Ag-NP aggregation. The thermogravimetric investigation revealed that the produced SPS-Ag-NPs have remarkable thermal stability. The produced SPS-Ag-NP exceeded total antioxidant activity (13.8 µmol Trolox equivalent) more than the SPS-phenolic extract (12.0 µmol Trolox equivalent). The biosynthesized SPS-Ag-NPs exhibited noticeably better antibacterial activity against multidrug-resistant Gram-negative E. coli and Gram-positive S. aureus compared to SPS-phenolic extract. Hence, the bio-synthesized SPS-Ag-NPs demonstrated great potential for use in biomedical and antimicrobial applications.

Antioxidant system of garden cress sprouts for using in bio-monitor of cadmium and lead contamination.

Based on garden cress significantly used for phytoremediation, the antioxidant system included antioxidant-phenolic compounds and antioxidant-enzymes of 6-day-garden cress sprouts (GCS) were assessed as potential bio-indicators for cadmium (Cd) and lead (Pb) contamination. Total phenolic and flavonoid contents of GCS germinated under Cd and Pb treatments (25-150 mg kg-1) gradually increased with increasing concentration of metals and peaked by 2.0, 2.6, and 2.5, 2.3 folds at 150 mg kg-1, respectively. By using DPPH, ABTS, and PMC antioxidant assays, the total antioxidant activity of phenolic compounds of GCS increased 6.1, 13.0, and 5.8-fold for Cd and 5.9, 14.6, and 8.2-fold for Pb at 150 mg kg-1, respectively. The antioxidant enzymes of GCS (POD, CAT, GR, and GST) were significantly activated in response to Cd and Pb stress, and two new electrophoretic POD bands were detected. GCS was absorbed 19.0% and 21.3% of Cd and Pb at 150 mg metal kg-1, respectively. In conclusion, the approaches of the antioxidant defense system of GSC could potentially be used as bio-indicator for monitoring Cd and Pb contamination in a short time of germination process.

Garden cress gum and maltodextrin as microencapsulation coats for entrapment of garden cress phenolic-rich extract: improved thermal stability, storage stability, antioxidant and antibacterial activities.

The obtained garden cress 6-day sprouts phenolic-rich extract (GCSP) contained efficient health-promoting antioxidant-phenolic compounds. To improve the stability, bioavailability, and functional properties of these valuable phenolic compounds, GCSP was encapsulated by freeze-drying technique using different ratios of garden cress gum (GG) and maltodextrin (M) in the absence and presence of sonication (S). The prepared S/GG-microcapsule retained the highest phenolic content (95%), antioxidant activity (141.6%), and encapsulation efficiency (98.2%). It displayed the highest bio-accessibility of GCSP-phenolic compounds in simulated intestine fluid (87%) and demonstrated the greatest storage-stability at 40 °C for 60 days. S/GG-microcapsule possessed better physical properties including moisture, solubility, swelling, and morphological structures using SEM. The main spectral features, crosslinking, and improved thermal stability were demonstrated for S/GG-microcapsule using FTIR and thermogravimetric analyses. S/GG-microcapsule demonstrated much greater antibacterial activity than GCSP against pathogenic bacteria. S/GG-microcapsule can be added to different food products to improve their antioxidant and antibacterial properties.

Biotechnology approach using watermelon rind for optimization of α-amylase enzyme production from Trichoderma virens using response surface methodology under solid-state fermentation.

Production of amylases by fungi under solid-state fermentation is considered the best methodology for commercial scaling that addresses the ever-escalating needs of the worldwide enzyme market. Here response surface methodology (RSM) was used for the optimization of process variables for α-amylase enzyme production from Trichoderma virens using watermelon rinds (WMR) under solid-state fermentation (SSF). The statistical model included four variables, each detected at two levels, followed by model development with partial purification and characterization of α-amylase. The partially purified α-amylase was characterized with regard to optimum pH, temperature, kinetic constant, and substrate specificity. The results indicated that both pH and moisture content had a significant effect (P < 0.05) on α-amylase production (880 U/g) under optimized process conditions at a 3-day incubation time, moisture content of 50%, 30 °C, and pH 6.98. Statistical optimization using RSM showed R2 values of 0.9934, demonstrating the validity of the model. Five α-amylases were separated by using DEAE-Sepharose and characterized with a wide range of optimized pH values (pH 4.5-9.0), temperature optima (40-60 °C), low Km values (2.27-3.3 mg/mL), and high substrate specificity toward large substrates. In conclusion, this study presents an efficient and green approach for utilization of agro-waste for production of the valuable α-amylase enzyme using RSM under SSF. RSM was particularly beneficial for the optimization and analysis of the effective process parameters.

Egyptian chia seeds (Salvia hispanica L.) during germination: Upgrading of phenolic profile, antioxidant, antibacterial properties and relevant enzymes activities.

Little studies on chia sprouts were not deeply address the polyphenols profiles and their functional properties during long period of germination. This study aims to evaluate the impact of germination process on the phenolic profile, antioxidant and antibacterial properties and relevant enzymes activities of Egyptian chia seeds. The total phenolic and flavonoid contents of chia sprouts increased several times during ten days of germination and maximized on 7-day sprouts (6.4 and 11.5 folds, respectively). In HPLC analysis, seventeen phenolic compounds were detected on 7-day sprouts compared to fifteen in dry seeds, where two new phenolic compounds (p-coumaric acid and kaempferol) were detected. The concentrations of all the identified phenolic compounds increased several folds (1.8-27) on 7-day sprouts. The total antioxidant activity increased 10, 17, and 29 folds on 7-day sprouts using DPPH, ABTS and PMC antioxidant methods, respectively compared to the dry seeds. Both antioxidant and carbohydrate-cleaving enzymes increased in chia sprouts and correlated with their phenolic content and antioxidant activity. The phenolic content of 7-day sprouts showed a potent antibacterial activity against some human enteric pathogenic bacteria including Escherichia coli O157-H7, Salmonella typhi, Pseudomonas aeruginosa and Staphylococcus aureus with lower MIC values compared to the raw seeds.

Immobilization of horseradish peroxidase on cationic microporous starch: Physico-bio-chemical characterization and removal of phenolic compounds.

In the present study, two different modified starches; microporous starch (MPS) and cationic microporous starch (CMPS) were synthesized. The granules of MPS that distributed regularly were destroyed after the etherification reaction. The data depicted that the immobilization of horseradish peroxidase (HRP) on CMPS revealed highest immobilization efficiency (86%) at 100 mg of CMPS at pH = 6.0 and 100 units of enzyme. After 10 reuses of the CMPS-HRP, it retained 66% of initial activity. The soluble HRP showed broad pH optimum of 6.0-7.0, which changed to sharp pH = 6.0 for CMPS-HRP. Soluble-HRP and CMPS-HRP showed temperature optima at 30 °C and 40 °C, respectively. The CMPS-HRP showed high thermal stability up to 50 °C compared to the soluble HRP (40 °C). The Km values of soluble HRP and CMPS-HRP were 6.6 and 10.8 mM for H2O2 and 34 and 41.6 mM for guaiacol, respectively. CMPS-HRP showed higher affinity toward various substrates than the soluble-HRP. CMPS-HRP showed more resistance against heavy metals, urea, isopropanol, Triton X-100 and trypsin than soluble enzyme. The CMPS-HRP showed higher ability to remove phenol and p-chlorophenol compared to soluble-HRP.

Purification and characterization of peroxidases from garden cress sprouts and their roles in lignification and removal of phenol and p-chlorophenol.

The study aims to evaluate the relation between peroxidases of day-6 garden cress sprouts and phenolic compounds. Three cationic, three anionic, and two unbounded peroxidases were separated from day-6 garden cress sprouts. Cationic (GCP1) and anionic (GCP2) peroxidases were purified with molecular masses of 25 and 40 kDa, respectively. The Km values of GCP1 toward H2 O2 and guaiacol were lower than GCP2. The anionic GCP2 exhibited high affinity toward some lignin monomers, sinapyl alcohol, coniferyl alcohol, cinnamic and ferulic acids. Therefore, GCP2 is considered as a lignin peroxidase and contributed in lignin synthesis. The activity of GCP1 and GCP2 was stable at a wide pH range 5.5-8.0 and 6.0-7.5, respectively. Both peroxidases showed the same thermal stability range 20-50°C. GCP2 was more resistant against the effect of metal ions than GCP1. GCP2 showed high ability to remove of phenol and p-chlorophenol from effluent compared to GCP1. PRACTICAL APPLICATIONS: Generally, garden cress is used as a test plant to conduct biomonitoring of pollution in urban soil on a wide scale because of its simplicity, sensitivity, and cost-effectiveness. Peroxidase is an important antioxidant enzyme, which elevated when plant subjected to pollution. Recently, we reported that the increase of peroxidase activity was strongly correlated with high phenolic content and antioxidant activity during the germination of garden cress. In the present study, anionic peroxidase GCP2 may play an important role in lignification process and removal of phenol and p-chlorophenol from polluted soil/wastewater as well as resisted the harmful effect of heavy metals. Cationic peroxidase GCP1, as a natural scavenger, had high affinity toward H2 O2 coupled to oxidation of some plant phenolic compounds suggesting its role in consuming of excess H2 O2 .

Diabetic complications and oxidative stress: The role of phenolic-rich extracts of saw palmetto and date palm seeds.

Recently, we reported that the date palm seed (DP) and saw palmetto seed (SP) extracts possessed a great amount of phenolic contents with potent antioxidant, antimicrobial, and anti-inflammatory activities. Therefore, this study aimed to assess the role of DP and SP phenolic-rich extracts in modulating diabetic complications and oxidative stress in the STZ- diabetic rat. DP and SP extracts significantly inhibited both microbial and pancreatic α-amylases. The STZ-induced diabetic rat groups treated with DP and SP extracts exhibited reversed hyperglycemia (40% and 54%, p < .001-.01) and body weight (70%, p < .001) alteration close to normal. Moreover, DP and SP extracts modulated serious damages in the structures of the pancreas, kidney, and liver of diabetic rats. DP and SP extracts improved the decline of the activities of antioxidant enzymes: Catalase, glutathione-S-transferase, glutathione reductase, and glutathione peroxidase in liver, kidney, and pancreas of the diabetic rats. PRACTICAL APPLICATIONS: Generally, date seed is a rich source of dietary fibers, polyphenols, and antioxidants and used in foods and pharmaceuticals. Our study reported that date palm seed (DP) and saw palmetto seed (SP) phenolic-rich extracts attenuated diabetes and its complications, probably tissue regeneration and normalizing the oxidative stress in the STZ-induced diabetic rat.

Bioremediation of potentially toxic metal and reactive dye-contaminated water by pristine and modified Chlorella vulgaris.

This is the first study on the biosorption of Cu2+ and reactive yellow 145 (RY145) dye by citric acid (CA), NaOH, and heat-treated Chlorella vulgaris (Cv). Influence of contact time, initial adsorptive concentration, and biomass dosage on the biosorption process was explored. The biosorption kinetics and isotherm were comprehensively investigated as well. The Fourier transform infrared analysis proved the successful insertion of carbonyl groups on Cv surface by CA modification and the intensification of all Cv functional groups by heat treatment. CA modified Cv was the best biosorbent for RY145, 0.5 g/L removes 97% of 10 mg/L solution (pHi 2) in 40 min. The biosorption was favorable, occurred via the formation of a monolayer of RY145 on the homogenous surface of CA-modified Cv and followed the pseudo-second-order kinetics. On the other hand, heat-treated Cv was the best biosorbent for Cu2+, 0.5 g/L removes 92% of 10 mg/L solution (pHi 5) in 5 min. The biosorption of Cu2+ on heat-treated Cv was complex and involves more than one mechanism. The Langmuir theoretical monolayer saturation capacity of RY145 on CA-modified Cv was comparable to other biosorbents, while that of Cu2+ on heat-treated Cv was drastically superior.

A hemorrhagic metalloprotease of Egyptian Cerastes vipera venom: Biochemical and immunological properties.

A hemorrhagic metalloprotease (CVHT1) was isolated from Cerastes vipera (CV) viper venom and characterized in a set of biochemical and immunological assays. A simple two-step purification procedure included gel filtration and ion-exchange increase the purity of enzyme 39-fold with specific activity of 20,200 Umg-1 compared to 520 Umg-1 for crude venom. CVHT1 is a dimer enzyme with two subunits of ~60 kDa. The LC-MS/MS analysis of CVHT1 revealed that the identified peptides show high homology to other P-III snake venom zinc-metalloproteases. The activity of CVHT1 showed stability at pH (6.5-8.5) and temperature (30-60 °C) with optima at pH 8.5 and 60 °C. Activators for CVHT1 included Mg+2, Zn+2, Ca+2, K+, Ba+2 and Na+, while the full inhibition was given by other tested ions, SH-group reagents and metalloproteinase inhibitors. The CVHT1 potentially digested gelatin, fibrinogen, fibronectin and inhibited the platelet aggregation. The hemorrhagic and proteolytic activities of medically important Egyptian viper venoms were highly cross-neutralized by anti-CVHT1. The anti-CVHT1 increased the survival time of mice injected with lethal dose of CV venom to 23 ±â€¯2.5 h compared to the mice injected with venom alone 0.52 ±â€¯0. 05 h. This study could be useful for production of safer and more efficient therapeutic anti-venom.

Ficus sycomorus latex: An efficient alternative Egyptian source for horseradish peroxidase in labeling with antibodies for immunodiagnostic kits.

AIM: In view of various peroxidase applications, the searching for new sources of unique peroxidase properties is highly desirable. The present study aims to evaluate the efficiency of the peroxidase of locally grown sycamore latex (POL) for conjugation with antibodies and to study the conjugate optimal conditions, storage stability, and affinity toward different substrates as compared with commercial horseradish peroxidase (HRP). MATERIALS AND METHODS: Anti-mouse antibodies were prepared in rabbits and purified by protein A sepharose affinity column chromatography. The POL and HRP conjugates were prepared by one-step glutaraldehyde coupling method. The reactivity of the prepared conjugates was examined using the enzyme-linked immunosorbent assay (ELISA). The optimal enzymatic conditions, storage stability, and affinity toward substrates were also determined for both the conjugates. RESULTS: The POL showed higher percent recovery (98%) than HRP (78%) over the initial activity after conjugation process. The POL and HRP conjugates showed ELISA titers of 1:120 and 1:80, respectively, demonstrating high binding affinity of POL-conjugate. The POL-conjugate showed high thermal stability up to 70°C compared with HRP-conjugate up to 40°C. After conjugation, POL had wide pH stability (5.0-8.0) compared with HPR (4.5-6.0). Both of the prepared conjugates had a high affinity toward the substrates used in immunoassays with lower Km values. The POL-conjugate showed high storage stability for its enzymatic activity and ELISA titer compared with HRP-conjugate after 1 year at -20°C. CONCLUSION: The POL of Ficus sycomorus latex is an efficient source for labeling antibodies and could be utilized in immunodiagnostic kits.

Rosemary leaves extract: Anti-snake action against Egyptian Cerastes cerastes venom.

The morbidity caused by viper bites is very dangerous and the anti-venom therapy couldn't treat the local injures such as hemorrhage, edema, necrosis and inflammation of bitten tissues. Searching for safe and effective anti-venom compounds from natural sources is very important. This study was designed to explore the neutralizing ability of Rosmarinus officinalis L. leaves aqueous extract (RMAE) against Egyptian Cerastes cerastes (Cc) viper venom toxicity. The RMAE contained a considerable amount of phenolic and flavonoid contents with 3,300 and 800 mg/100 g dry weight, respectively. The RMAE showed a considerable variation of phenolic acids by using HPLC technique. Rosmarinic acid is the major component of the RMAE which recorded 400 mg/100 g dry weight and 64% of all the identified compounds. In vitro, the RMAE neutralized the enzymatic activities of proteases, l-amino acid oxidases, and phospholipases A2 of the Cc venom dose-dependently. In addition, the RMAE effectively neutralized the gelatinolytic, fibrinogenolytic, hemolytic and procoagulant activities of Cc venom. In vivo, the RMAE markedly reduced lethality, hemorrhage, edema, muscle and liver toxicities induced by Cc venom. In conclusion, the venom neutralizing property of the RMAE gives a new prospect for efficient treatment of the lethal viper bites.

Phenolic-antioxidant capacity of mango seed kernels: therapeutic effect against viper venoms

Abstract In this study, mango seed kernels extract contained a considerable amount of phenolics and flavonoids (17,400 and 3325 mg/100 g seed, respectively). The HPLC profiling revealed that hesperidin was the major phenolic compound of the mango seed kernels extract. This is the first report find hesperidin in mango extracts. The phenolic compounds of mango seed kernels extract were effective in scavenging free radicals of DPPH and ABTS with IC50 values of 47.3 and 7.9 µg/ml, respectively. The total antioxidant activity of mango seed kernels extract based on the reduction of molybdenum was also measured. The phenolic compounds of mango seed kernels extract potentially inhibited the protease, fibrinogenase, phospholipase A2, l-amino acid oxidase, hyaluronidase, and hemolytic activities of the most dangerous Cerastes cerastes and Echis coloratus viper venoms. The phenolic compounds of mango seed kernels extract could completely neutralize the hemorrhage and lethality of both venoms in experimental animals. It could be concluded that the mango seed kernels extract phenolic compounds with potential antioxidant activity are considered as a new avenue in the viper bite treatment.

Purification and characterization of a new thermophilic collagenase from Nocardiopsis dassonvillei NRC2aza and its application in wound healing.

A thermostable metallo-collagenase enzyme (150 kDa), recently identified in a newly isolated actinomycestes strain (Nocardiopsis dassonvillei NRC2aza), has been purified from natural source, characterized to have application in wound healing. A simple 3 step purification procedure gave an increase of purity by 6.23 fold with a specific activity of 387.2 U mg-1. The enzyme activity showed stability across a range of pH (7.0-8.5) and temperature (40-55 °C) with optima at pH 8.0 and 60 °C, respectively. Activators include Mg+2, Ca+2, Zn+2, Na+, K+ and Ba+2, while Mn+2, Co+2, Ni+2and Ag+ ions gave partial inhibition. Full inhibition was given by other tested ions and metalloproteinase inhibitors. Broad substrate specificity was demonstrated including activity against a native collagen. The Km and Vmax of the enzyme using azocollagen were 5.5 mg/ml and 1280 U, respectively. The purified collagenase enhanced wound closure in vitro and in vivo and the repair process was dose dependent. Topical application of the purified collagenase (either of 25 or 50 U) to cutaneous wounds significantly accelerated the rate of wound healing and the formation of granulation tissue. Hence, the purified collagenase has a great potential as a therapeutic agent in wound care and collagen related diseases.

Immobilization of Trichoderma harzianum α-amylase on PPyAgNp/Fe3O4-nanocomposite: chemical and physical properties.

In this study, a new support has been developed by immobilization of α-amylase onto modified magnetic Fe3O4-nanoparticles. The characterization of soluble and immobilized α-amylases with regards to kinetic parameters, pH, thermal stability and reusability was studied. The effect of polypyrrole/silver nanocomposite (PPyAgNp) percentage on weight of Fe3O4 and pH on the immobilization of α-amylase was studied. The highest immobilization efficiency (75%) was detected at 10% PPyAgNp/Fe3O4-nanocomposite and pH 7.0. Immobilization of α-amylase on PPyAgNp/Fe3O4-nanocomposite was characterized by FT-IR spectroscopy and scanning electron microscopy. The reusability of the immobilized enzyme activity was 80% of its initial activity after 10 reuses. The immobilized enzyme was more stable towards pH, temperature and metal ions compared with soluble enzyme. The kinetic study appeared higher affinity of immobilized enzyme (Km 2.5 mg starch) compared with soluble enzyme (Km 3.5 mg starch). In conclusion, the immobilization of α-amylase on PPyAgNp/Fe3O4-nanocomposite could successfully be used in industrial and medical applications.

Detection of three herbicide, and one metabolite, residues in brown rice and rice straw using various versions of the QuEChERS method and liquid chromatography-tandem mass spectrometry.

A single-run analytical method was developed to analyze the three herbicides azimsulfuron, bensulfuron-methyl, and mesotrione and its metabolite (4-methylsulfonyl-2-nitrobenzoic acid (MNBA)) in brown rice and rice straw using liquid chromatography-tandem mass spectrometry (LC/MS/MS). Samples extracted using various versions of Quick, Easy, Cheap, Effective, Rugged, and Safe "QuEChERS" (original unbuffered, acetate (AOAC), and citrate (EN) buffered) methods gave poor recoveries of all the tested analytes in both matrices. The extraction efficiency was improved when primary-secondary amine (PSA) sorbent was removed from the purification step, with the best recovery being achieved for EN-QuEChERS, which was subsequently used throughout the study. Overall, a determination coefficients (R(2))⩾0.995 was achieved at matrix-matched calibration curves at various concentration ranges. The recovery rates at three fortification levels (limit of quantification (LOQ), 1/2 maximum residue limit (1/2MRL), and MRL) ranged from 78 to 114.5, with relative standard deviations (RSDs)<18% for all the tested analytes in both matrices. The LOQs for all herbicides were lower than the MRL set by the Ministry of Food and Drug Safety (MFDS), Republic of Korea. Field trials with the recommended, or double the recommended dose, revealed that the herbicides can safely be applied to rice, as no residues were detected in the harvested samples at 110days.

Purification and partial characterization of serine-metallokeratinase from a newly isolated Bacillus pumilus NRC21.

A serine metallokeratinase enzyme (30 kDa) produced by a newly isolated Bacillus strain (Bacillus pumilus NRC21) cultivated under optimized conditions in medium containing chicken feather meal was purified and characterized in a set of biochemical assays. The purification was carried out using two successive chromatographic steps; cation exchange chromatography on CM-cellulose and gel filtration on sephadex G-100 columns. The purified enzyme showed a specific activity of 2000 units/mg protein against 170 units/mg protein for crude extract with 12 fold purification. The enzymatic activity of the keratinase stimulated by (Na(+), K(+), Mg(2+)), Hg(+2) had no effect, and inhibited by entire tested cations, serine and metalloproteinase inhibitors, therefore it can be considered as a serine metalloenzyme. The optimum pH and temperature for the purified enzyme were (7.5, 8.5) and (50, 45 °C) when using keratin azure and azocasein as substrates, respectively. The purified enzyme was highly stable at broad pH and temperature ranged (5-10) and (20-60 °C), respectively and its thermoactivity and thermostability were enhanced in the presence of 5 mM Mg(+2). These results suggest that the purified keratinase may be used in several industrial applications.