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1.
A mixed-gel of κ-carrageenan and gelatin was used in l-phenylalanine production. The mixed-gel, containing 87.5% κ-carrageenan and 12.5% gelatin [the total gel concentration was 4 wt%], showed the best performance and was selected for further study with Escherichia coli EP8-10. The optimum pH and temperature were 8.5 and 37 °C, respectively. The effects of trehalose and Mg2+ were studied in the mixed-gel immobilization. Their optimum concentrations were 5 × 10?2 and 2 × 10?3 mol/L, respectively. Under the optimal conditions, 98.3% of the phenylpyruvic acid (PPA) was converted to l-phenylalanine. The activity recovery of the transaminase enzyme in the mixed-gel immobilization was higher than that in single κ-carrageenan immobilization, which was 93.6%. The total PPA conversion rate was over 80% in all 15 batches, suggesting great sustainability in the mixed-gel immobilization. The maximum reaction rate (rmax) was calculated to be 4.75 × 10?2 mol/(L g h).  相似文献   

2.
Laccase enzyme (L) from Trametes versicolor was entrapped in three hydrogel structures namely poly(acrylamide-N-isopropylacrylamide), P(AAm-NIPA), and semi-interpenetrating networks of poly(acrylamide)/alginate, P(AAm)/Alg, and poly(acrylamide-N-isopropylacrylamide)/alginate, P(AAm-NIPA)/Alg. The optimum temperatures for free and all immobilized systems were found to be 40 °C. For free and immobilized laccase systems of P(AAm-NIPA)-L, P(AAm)/Alg-L and P(AAm-NIPA)/Alg-L, Km values were found to be 6.7 × 10?3, 8.8 × 10?2, 5.5 × 10?2 and 1.8 × 10?2 mM; Vmax values were calculated as 1.8 × 10?3, 2.5 × 10?2, 1.5 × 10?2 and 6.1 × 10?3 mM min?1, respectively. For free and the same immobilized systems, the enzymes retained 42%, 91%, 79% and 86% of their initial activities at the end of 56 days of storage. After using the mentioned immobilized systems repeatedly 10 times, they retained 77%, 71% and 84% of their original activities, respectively. For free and the same immobilized systems, decolorization of Acid Orange 52 (AO52) in 6 h were found to be 63%, 50%, 48% and 66%, respectively. Addition of 2,2′-azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid), ABTS, into the assay medium increased these values up to 73%, 73%, 74% and 75%, respectively.  相似文献   

3.
Enzymes in the newly described rumen bacterium, Treponema zioleckii strain kT, capable of digesting Timothy grass fructan, inulin, and sucrose were identified and characterized. Two specific endolevanases and one non-specific β-fructofuranosidase were found in a cell-free extract. The molecular weight of the endolevanases were estimated to be 60 and 36 kDa, whereas that of β-fructofuranosidase, 87 kDa. The former of the specific enzymes was associated with the outer membrane, while the latter and the non-specific β-fructofuranosidase, with the periplasm or cytosol. The Km and Vmax for Timothy grass fructan degradation by endolevanase were 0.27% and 15.75 μM fructose equivalents × mg protein?1 × min?1, those for sucrose and inulin digestion by β-fructofuranosidase were 1.35 × 10?3 M and 1.73 μM hexoses × mg protein?1 × min?1 and 1.77% and 1.83 μM hexoses × mg protein?1 × min?1, respectively.  相似文献   

4.
New ionic liquid-modified silica sorbents were developed by the surface chemical modification of the commercial silica using synthesized ionic liquids. The obtained ionic liquid-modified particles were successfully used as a special sorbent in solid-phase extraction process to isolation of cryptotanshinone, tanshinone I and tanshinone IIA from Salvia Miltiorrhiza Bunge. Different washing and elution solvents such as water, methanol and methanol–acetic acid (90/10, v/v) were evaluated. A comparison of ionic liquid-modified silica cartridges and traditional silica cartridge show that higher recovery was observed using ionic liquid-modified silica sorbents. A quantitative analysis was conducted by high-performance liquid chromatography using a C18 column (5 μm, 150 mm × 4.6 mm) with methanol–water (78:22, v/v, and containing 0.5% acetic acid) as a mobile phase. Good linearity was obtained from 0.5 × 10?4 to 0.5 mg/mL (r2 > 0.999) with the relative standard deviations less than 4.8%.  相似文献   

5.
A new analytical method for puerarin using capillary electrophoretic (CE) separation and chemiluminescence (CL) detection has been developed. The detection was based on the enhanced CL intensity of the reaction between luminol and potassium ferricyanide by puerarin in alkaline solution. A laboratory-built CE–CL apparatus was deployed for the puerarin detection. Under the optimal conditions, a linear range from 5.0 × 10?8 to 2.5 × 10?6 M and a detection limit of 1.0 × 10?8 M (S/N = 3) for puerarin were achieved. The determination of puerarin was achieved in less than 5 min, and the proposed method was applied to the determination of puerarin in pharmaceutical, human urine and human plasma samples.  相似文献   

6.
A new laccase from Shiraia sp.SUPER-H168 was purified by ion exchange column chromatography and gel permeation chromatography and the apparent molecular mass of this enzyme was 70.78 kDa, as determined by MALDI/TOF-MS. The optimum pH value of the purified laccase was 4, 6, 5.5 and 3 with 2,6-dimethoxyphenol (DMP), syringaldazine, guaiacol and 2,2′-azinobis-(3-ethylbenzthiazoline-6-sulphonate) (ABTS) as substrates, respectively. The optimum temperature of the purified laccase was 50 °C using DMP, syringaldazine and guaiacol as substrates, but 60 °C for ABTS. Inhibitors and metal ions of SDS, NaN3, Ag+ and Fe3+ showed inhibition on enzyme activity of 10.22%, 7.86%, 8.13% and 67.50%, respectively. Fe2+ completely inhibited the purified laccase. The Kcat/Km values of the purified laccase toward DMP, ABTS guaiacol and syringaldazine were 3.99 × 106, 3.74 × 107, 8.01 × 104 and 2.35 × 107 mol?1 L S?1, respectively. The N-terminal amino acid sequence of the purified laccase showed 36.4% similarity to Pleurotus ostrestus. Approximately 66% of the Acid Blue 129 (100 mg L?1) was decolorized by 2.5 U of the purified laccase after a 120 min incubation at 50 °C. Acid Red 1 (20 mg L?1) and Reactive Black 5 (50 mg L?1) were decolorized by the purified laccase after the addition of Acid Blue 129 (100 mg L?1).  相似文献   

7.
In the present work, a simple and high sensitive method based on hollow fiber liquid phase microextraction (HF-LPME) was developed followed by high performance liquid chromatography (HPLC) for determination of ultra-trace amounts of Se(IV) after derivatization in biological and natural water samples. Se(IV) was complexed with o-phenylenediamine to form piazselenol. The formed piazselenol was extracted into 20 μL of 1-octanol located in the lumen of a hollow fiber and the solution was injected into HPLC-UV for analysis. Using the Taguchi method, an orthogonal array design (OAD), OA16 (45) was employed to optimize the HF-LPME of piazselenol. The effect of five experimental factors (each factor at four levels) including the volume of the organic phase, extraction time, pH of the solution, stirring rate and ionic strength on the extraction efficiency of piazselenol was studied and optimized. The maximum extraction efficiency of piazselenol was obtained at 20 μL of 1-octanol as the extracting solvent, 30 min extraction time, pH 2, stirring rate of 500 rpm and 30% (w/v) NaCl. Under the optimum conditions, preconcentration factors up to 130 were achieved and the relative standard deviation (%RSD) of the method was <3.7% for different concentrations of Se(IV). The calibration curves were obtained in the ranges of 0.2–100 and 0.05–10 μg L?1 for the 11 and 50 mL of the sample volumes with reasonable linearity, respectively (r2 > 0.995). The limits of detection (LOD) were 0.1 and 0.02 μg L?1 for the 11 and 50 mL sample volumes, respectively (S/N = 3). Finally, the applicability of the proposed method was evaluated by the extraction and determination of Se(IV) in the plasma, urine and water samples.  相似文献   

8.
An LC–MS method for the determination of metoclopramide in human plasma was developed and validated. Sample preparation involved extraction with ethyl acetate. Chromatographic separation was performed on a Thermo Hypersil-Hypurity C18 (150 mm × 2.1 mm, 5 μm) with the mobile phase consisting of 40 mM ammonium acetate–methanol–acetonitrile. A single-quadrupole mass spectrometer with an electrospray interface was operated in the selected-ion monitoring mode to detect the [M+H]+ ions at m/z 300 for metoclopramide and at m/z 384 for the internal standard (prazosin). The method was validated over 0.78–50.00 ng mL?1 for metoclopramide. The recovery was 67.8–83.1%, and the limit of quantitation (LOQ) detection was 0.78 ng mL?1 for metoclopramide. The intra- and inter-day precision of the method at three concentrations was 5.0–13.6% with accuracy of 99.2–104.0%. Stability of compounds was established in a battery of stability studies. The method was successfully applied to bioequivalence studies of metoclopramide hydrochloride tablets to obtain the pharmacokinetic parameters.  相似文献   

9.
Recent evidences suggested a possible relationship between zinc deficiency and leptin levels in pathogenesis of anorexia in chronic kidney disease. The present study addressed the relationship between zinc and leptin in hemodialysis (HD) patients.MethodsFifty HD patients (54.3 ± 12.7 years old, 62% men) were studied and compared to 21 healthy volunteers (50.7 ± 15.7 years old, 43% men). Biochemical data, serum zinc, plasma leptin, IL-6, TNF-α and C-Reactive Protein levels were determined. Anthropometric parameters, food intake and appetite score were also assessed.ResultsThe leptin levels were higher in HD patients (16.1 μg/mL (0.21–118.25) vs 6.0 μg/mL (0.50–23.10)) in healthy volunteers (p = 0.04), whereas serum zinc levels were lower (54.5 ± 16.3 μg/dL) compared to healthy volunteers (78.4 ± 9.4 μg/dL) (p = 0.0001). The plasma leptin was correlated negatively with plasma zinc (r = ?0.33; p = 0.007), energy (r = ?0.38; p = 0.002) and protein intake (r = ?0.34; p = 0.006) and, positively correlated with BMI (r = 0.54; p = 0.0001), % body fat (r = 0.70; p = 0.0001) and conicity index (r = 0.46; p = 0.001). Plasma zinc was associated with hemoglobin (r = 0.30; p = 0.04) and negatively associated with TNF-α (r = ?0.37; p = 0.002) and C-Reactive Protein (r = ?0.37; p = 0.004). There was no correlation among Zn, leptin and appetite score in these patients.ConclusionThis study showed that low plasma zinc levels are negatively associated with high leptin levels in HD patients.  相似文献   

10.
New pre-concentration technique, triple phase suspended droplet microextraction (SD-LPME) and liquid chromatography-photodiode array detection was applied to determine ecstasy, MDMA (3,4-methylendioxy-N-methylamphetamine) in hair samples. In this research MDMA in hair was digested and after treatment extracted. The effective parameters were investigated and method was evaluated. Under the optimal conditions, the MDMA was enriched by factor 98.11. Linearity (r = 0.9921), was obtained in the range of 10–15,000 ng mL?1 and detection limit was 0.1 ng mL?1.  相似文献   

11.
Biodegradation rate and the high molecular weight hydrocarbons are among the important concerns for bioremediation of crude oil. Inoculation of a non-oil-degrading bacterium as supplementary bacteria increased oil biodegradation from 57.1% to 63.0% after 10 days of incubation. Both the oil-degrading bacteria and the non-oil-degrading bacteria were isolated from Malaysian marine environment. Based on the 16S rDNA sequences, the oil-degrading bacteria was identified as Pseudomonas pseudoalcaligenes (99% similarity) while the non-oil-degrading bacterium was Erythrobacter citreus (99% similarity). E. citreus does not grow on crude oil enriched medium under present experimental condition but it withstands 5000 mg kg?1 Tapis blended crude oil in sediment. Under optimal condition, the oil-degrading bacterium; P. pseudoalcaligenes, alone utilized 583.3 ± 3.8 mg kg?1 (57.1%) at the rate of 3.97 × 10?10 mg kg?1 cell?1 day?1 Tapis blended crude oil from 1000 mg kg?1 oil-contaminated sediment. Inoculation of E. citreus as the supplementary bacteria to P. pseudoalcaligenes enhanced biodegradation. The bacterial consortium degraded 675.8 ± 18.5 mg kg?1 (63.0%) Tapis blended crude oil from the 1000 mg kg?1 oil-contaminated sediment. Biodegradation rate of the bacterial consortium increased significantly to 4.59 × 10?10 mg kg?1 cell?1 day?1 (p = 0.02). Improvement of the oil degradation by the bacterial consortium was due to the synergetic reaction among the bacterial inoculants. There are two implications: (1) E. citreus may have a role in removing self-growth-inhibiting compounds of P. pseudoalcaligens. (2) P. pseudoalcaligenes degraded Tapis blended crude oil while E. citreus competes for the partially degraded hydrocarbons by P. pseudoalcaligenes. P. pseudoalcaligenes forced to breakdown more hydrocarbons to sustain its metabolic requirement. The bacterial consortium degraded 78.7% of (C12–C34) total aliphatic hydrocarbons (TAHs) and 74.1% of the 16 USEPA prioritized polycyclic aromatic hydrocarbons.  相似文献   

12.
The aim of this research was to identify a low cost medium based on commercial products and by-products that provided maximum Bacillus subtilis CPA-8 growth and maintained biocontrol efficacy. Low cost media combining economical nitrogen and carbon sources such as yeast extract, peptone, soy products, sucrose, maltose and molasses were tested. Tests were carried out in 250-ml flasks containing 50 ml of each tested medium. Maximum cell growth (>3 × 109 CFU ml?1) was obtained in defatted soy flour 44% combined with sucrose or molasses media. Second, CPA-8 production was scaled up in a 5-l fermenter and CPA-8 population dynamics, pH and oxygen consumption in the optimized medium (defatted soy flour 44% – molasses) was recorded. In these tests, there was a 5-h lag phase before growth, after which exponential growth occurred and maximum production was 3 × 109 CFU ml?1 after 20 h. Fruit trials with cells and cell free supernatants from CPA-8 grown in optimized medium maintained biocontrol efficacy against Monilinia fructicola on peaches, resulting in disease reductions up to 95%. CPA-8 populations survived in wounds on inoculated peaches, regardless of the culture media used. The results show that B. subtilis CPA-8 can be produced in a low cost medium combining inexpensive nitrogen and carbon sources (40 g l?1 defatted soy flour 44%, 5 g l?1 molasses plus mineral trace supplements) in shake flasks and a laboratory fermenter (5 l). The results could be used to provide a reliable basis for scaling up the fermentation process to an industrial level.  相似文献   

13.
Low-molecular-mass trypsin inhibitor (clTI-1; chicken liver Trypsin Inhibitor-1) was purified from chicken liver by extraction with perchloric acid, ammonium sulfate precipitation, a combination of ethanol-acetone fractionation followed by gel filtration, ion-exchange chromatography and RP-HPLC on a C18 column. The inhibitor occurs in two isoforms with molecular masses of 5938.56 and 6026.29 Da (determined by MALDI TOFF mass spectrometry). The complete amino acid sequences of both isoforms were determined (UniProtKB/Swiss-Prot P85000; ISK1L_CHICK). The inhibitor shows a high homology to Kazal-type family inhibitors, especially to trypsin/acrosin inhibitors and pancreatic secretory trypsin inhibitors. clTI-1 inhibits both bovine and porcine trypsin (Ka = 1.1 × 109 M?1 and 2.5 × 109 M?1, respectively). Significant differences were shown in the inhibition of the anionic and cationic forms of chicken trypsin (Ka = 4.5 × 108 M?1 and 1.2 × 1010 M?1). Weak interaction with human plasmin (Ka = 1.2 × 107 M?1) was also revealed.  相似文献   

14.
Twelve four-month old Suffolk × Small-tail-Han male sheep (live weight 21–26 kg), fitted with rumen and abomasum fistulas and nourished by total intragastric infusions, were used to study the relationship between the volatile fatty acids (VFA) supply and the nitrogen (N) retention in sheep. The animals were randomly divided into four groups and four levels of mixed VFA energy (the molar proportion of acetic acid, propionic acid and butyric acid was 65:25:10), i.e. 333, 378, 423 and 468 kJ kg?1 W0.75 d?1, were infused into the rumen, as treatments I, II, III and IV, respectively. The results showed that the N retention was significantly increased (P < 0.05) with the VFA infusion level. Significant regression relationship was found between the VFA supply (x, g d?1) and the N retention (y, mg d?1): y = 2.75x ? 403, r2 = 0.86, n = 12, P < 0.01. It was concluded that to get efficient utilization of dietary N and high N retention in sheep, it is necessary to supply enough dietary energy.  相似文献   

15.
Five new α-aminophosphonates are synthesized and characterized by EA, FT-IR, 1H NMR, 13C NMR, 31P NMR, ESI-MS and X-ray crystallography. The X-ray analyses reveal that the crystal structures of 1–5 are monoclinic or triclinic system with the space group P 21/c, P  1, P  1, P2(1)/c and P  1, respectively. All P atoms of 1–5 have tetrahedral geometries involving two O-ethyl groups, one Cα atom, and a double bond O atom. The binding interaction of five new α-aminophosphonate N-derivatives (1–5) with calf thymus(CT)-DNA have been investigated by UV–visible and fluorescence emission spectrometry. The apparent binding constant (Kapp) values follows the order: 1 (3.38 × 105 M−1) > 2 (3.04 × 105 M−1) > 4 (2.52 × 105 M−1) > 5 (2.32 × 105 M−1) > 3 (2.10 × 105 M−1), suggesting moderate intercalative binding mode between the compounds and DNA. In addition, fluorescence spectrometry of bovine serum albumin (BSA) with the compounds 1–5 showed that the quenching mechanism might be a static quenching procedure. For the compounds 1–5, the number of binding sites were about one for BSA and the binding constants follow the order: 1 (2.72 × 104 M−1) > 2 (2.27 × 104 M−1) > 4 (2.08 × 104 M−1) > 5 (1.79 × 104 M−1) > 3 (1.17 × 104 M−1). Moreover, the DNA cleavage abilities of 1 exhibit remarkable changes and the in vitro cytotoxicity of 1 on tumor cells lines (MCF-7, HepG2 and HT29) have been examined by MTT and shown antitumor effect on the tested cells.  相似文献   

16.
A rapid and selective method for simultaneous determination of cyclophosphamide and its metabolite carboxyethylphosphoramide mustard (CEPM) was developed using online sample preparation and separation with tandem mass spectrometric detection. Diluted plasma was injected onto an extraction column (Cyclone MAX 0.5 mm × 50 mm, >30 μm), the sample matrix was washed with an aqueous solution, and retained analytes were transferred to an analytical column (Gemini 3 μm C18 110A, 100 mm × 2.0 mm) using a gradient mobile phase prior to detection by MS/MS. Analytes were detected in an API-3000 LC-MS/MS system using positive multiple-reaction monitoring mode (m/z 261/140 and 293/221 for CTX and CEPM, respectively). Online extraction recoveries were 76% and 72% for cyclophosphamide and CEPM. Within-day and between-day variabilities were <3.0%, and accuracies were between ?6.9% and 5.2%. This method has been used to measure plasma cyclophosphamide and CEPM concentrations in an ongoing Phase II study in children with newly diagnosed medulloblastoma.  相似文献   

17.
Polyhydroxyalkanoates (PHAs) have been recognized as good substitutes for the non-biodegradable petrochemically produced polymers. However, their high (real or estimated) current production cost limits their industrial applications. This work exploits two strategies to enhance PHAs substitution potential: the increase in PHA volumetric productivity in high density cultures and the use of waste glycerol (GRP), a by-product from the biodiesel industry, as primary carbon source for cell growth and polymer synthesis. Cupriavidus necator DSM 545 was used to accumulate poly(3-hydroxybutyrate) (P(3HB)) from GRP and from commercial glycerol (PG) as control substrate. On PG, productivities between 0.6 gPHB L?1 h?1 and 1.5 gPHB L?1 h?1 were attained. The maximum cell DW was 82.5 gDW L?1, the P(3HB) content being 62%. When GRP was used, 68.8 gDW L?1 with a P(3HB) accumulation of 38% resulting in a final productivity of 0.84 gPHB L?1 h?1 was obtained. By decreasing the biomass concentration at which accumulation was triggered, a productivity of 1.1 gPHB L?1 h?1 (50% P(3HB), w/w) was attained using GRP. P(3HB) molecular weights (Mw) ranged from 7.9 × 105 to 9.6 × 105 Da.  相似文献   

18.
A sensitive liquid chromatography–mass spectrometric method was validated for the quantification of RU38486 (mifepristone) in human and murine plasma. The analyte and internal standard (alfaxolone) were extracted by liquid–liquid extraction with diethyl ether, resolved on a C18 column using gradient elution with methanol and ammonium acetate and detected after positive electrospray ionization (m/z 430  372; m/z 333  297, respectively). Quantification was linear over the range 0.5–500 ng (r2 > 0.997), precise and accurate (intra-assay RSD  7.2%, RME  8.2%; inter-assay RSD  15.7% RME  10.2%). The limit of quantification (LOQ) was 50 pg injected on column, permitting reproducible analysis of RU38486 in small volumes of plasma.  相似文献   

19.
Constructed wetlands have a good potential for wastewater treatment in developing countries due to the simple operation and low implementation costs. Ornamental plants like Canna and Heliconia are used in the wetlands to increase their aesthetic value and these two species were compared in this study. Six pilot scale horizontal subsurface flow constructed wetland units were constructed at the Asian Institute of Technology (AIT) campus in Bangkok, Thailand, of which three were planted with Heliconia psittacorum L.f. × H. Spathocircinata (Aristeguieta) and three with Canna × generalis L. Bailey. The beds were loaded with domestic wastewater in four trials with hydraulic loading rates ranging from 55 to 440 mm d?1 corresponding to nominal detention times between 12 h and 4 days. Both plant species grew well in the systems and especially Canna had high growth rates (3100 ± 470 g DW m?2 yr?1) compared to Heliconia (550 ± 90 g DW m?2 yr?1). TSS mass removal rates were very high with efficiencies >88% even at hydraulic loading rates of 440 mm d?1. COD mass removal rates varied between 42 and 83% depending on the loading rates. The removal rate constants for COD as fitted by the first-order kC* model were estimated to be 0.283 and 0.271 m d?1 for Canna and Heliconia beds, respectively (C* = 28.1 and 26.7 mg l?1). Removals of nitrogen (N) and phosphorus (P) were low compared to the loading rates, but removal of total-N was higher in the beds planted with Canna than in beds with Heliconia because of the higher growth rate of Canna. It is concluded that ornamental species like Canna and Heliconia can be used to enhance the aesthetic appearance and hence the public acceptance of wastewater treatment systems in tropical climates. Canna is the preferred species from a treatment perspective because of its more vigorous growth, but since Heliconia has an economic potential as cut flowers may be preferred in many cases.  相似文献   

20.
A series of potent and subtype selective H3 receptor antagonists containing a novel tetrazole core and diamine motif is reported. A one-pot multi-component Ugi reaction was utilised to rapidly develop the structure–activity relationships (SAR) of these compounds. Optimisation for liver microsome stability (t1/2 >60 min), minimal CYP inhibition (IC50 >50 μM) and high cell permeability (Caco-2 Papp >20 × 10?6 cm/s) identified several compounds with drug-like properties.  相似文献   

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