首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献,搜索用时 31 毫秒
1.
Covalent UV/vis-quantifiable bis-aryl hydrazone bond formation was investigated for the preparation of conjugates between α-poly-d-lysine (PDL) and either α-chymotrypsin (α-CT) or horseradish peroxidase (HRP). PDL and the enzymes were first modified via free amino groups with the linking reagents succinimidyl 6-hydrazinonicotinate acetone hydrazone (S-HyNic, at pH 7.6) and succinimidyl 4-formylbenzoate (S-4FB, at pH 7.2), respectively. The modified PDL and enzymes were then conjugated at pH 4.7, whereby polymer chains carrying several enzymes were obtained. Kinetics of the bis-aryl hydrazone bond formation was investigated spectrophotometrically at 354 nm. Retention of the enzymatic activity after conjugate formation was confirmed by using the substrates N-succinimidyl-l-Ala-l-Ala-l-Pro-l-Phe-p-nitroanilide (for α-CT) and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS, for HRP). Thus, not only a mild and efficient preparation and convenient quantification of a conjugate between the polycationic α-polylysine and enzymes could be shown, but also the complete preservation of the enzymatic activity.  相似文献   

2.
A biosensor based on the enzyme-catalysed dissolution of biodegradable polymer films has been developed. Three polymer-enzyme systems were investigated for use in the sensor: a poly(ester amide), which is degraded by the proteolytic enzyme alpha-chymotrypsin; a dextran hydrogel, which is degraded by dextranase; and poly(trimethylene) succinate, which is degraded by a lipase. Dissolution of the polymer films was monitored by Surface Plasmon Resonance (SPR). The rate of degradation was directly related to enzyme concentration for each polymer/enzyme couple. The poly(ester amide)/alpha-chymotrypsin couple proved to be the most sensitive over a concentration range from 4 x 10(-11) to 4 x 10(-7) mol l(-1) of enzyme. The rate of degradation was shown to be independent of the thickness of the poly(ester amide) films. The dextran hydrogel/dextranase couple was less sensitive than the poly(ester amide)/alpha-chymotrypsin couple but showed greater degradation rates at low enzyme concentrations. Enzyme concentrations as low as 2 x 10(-11) mol l(-1) were detected in less than 20 min. Potential fields of application of such a sensor system are the detection of enzyme concentrations and the construction of disposable enzyme based immunosensors, which employ the polymer-degrading enzyme as an enzyme label.  相似文献   

3.
4.
Oligo-tyrosine peptides such as Tyr-Tyr having angiotensin I-converting enzyme (ACE) inhibitory activity could be synthesized by α-chymotrypsin-catalyzed reaction with l-tyrosine ethyl ester in aqueous media. However, peptide yield in the reaction was below 10%. Since l-tyrosine amide showed highly nucleophilic activity for the deacylation of enzyme through which a new peptide bond was made, its application to the enzymatic peptide synthesis was evaluated in this study. Addition of tyrosine amide into the reaction produced Tyr-Tyr-NH2, of which yield exceeded 130% on the basis of tyrosine ethyl ester. Although purified Tyr-Tyr-NH2 did not inhibit ACE activity, α-chymotrypsin could act on the dipeptide amide and convert about 40% of it to Tyr-Tyr. The use of both ester and amide forms of tyrosine is expected to be a potent procedure for α-chymotrypsin-catalyzed synthesis of antihypertensive peptides.  相似文献   

5.
Microparticles continue to receive widespread attention from food professionals because of their potential use as carriers of bioactive substances. This study demonstrates a novel method to prepare casein microparticles, which co-assemble with α-tocopherol (αT) into emulsion droplets. When the particles were extracted from the pectin matrix via enzymatic degradation, they remained stable in a buffer solution for at least 3 weeks. Optical microscopy showed that the size distribution of the microparticles is between 5 μm and 50 μm, which is in accordance with previous observations in blend films. High-performance liquid chromatography revealed that the amounts of αS1- and κ-casein in the microparticles are significantly higher than those in native casein micelles. Confocal Raman microscopy showed that in the presence of α-tocopherol, the microparticles assemble into emulsion droplets, with phenol in the core and casein in the shell. Herein, we demonstrate a new method to form casein-based emulsion droplets for potential use as carriers of bioactive substances.  相似文献   

6.
An α-galactosidase fraction which hydrolyzes galactosyl(α1→4)galactosylceramide and 4-methylumbelliferyl-α-galactoside has been isolated from normal human plasma by affinity chromaography. It was partially separated into two enzymatically active proteins by isoelectric focusing or cellulose acetate electrophoresis. The protein fraction obtained by affinity chromatography of Fabry plasma also was divided into two proteins, but only the protein of slower electrophoretic mobility had detectable enzymatic activity. These results indicate that the accumulation of galactosyl(α1→4)galactosylceramide in certain organs in Fabry's disease is due to an alteration of a specific α-galactosidase.  相似文献   

7.
Carrageenans are sulfated polysaccharides that are found in the cell walls of red algae. These polysaccharides have gelling and texturizing properties that are widely appreciated in industrial applications. However, these functional properties depend strongly on the sulfation of the moieties of the carrabiose repetition unit. Here we aimed to monitor the sulfate composition of gelling carrageenan. To do so, we screened and purified from Pseudoalteromonas atlantica a 4S-iota carrageenan sulfatase that converts ι-carrabiose into α-carrabiose units. The sequence of this protein matched the annotated Q15XH3 (Uniprot databank) formylglycine-dependent sulfatase found in the P. atlantica genome. With pure enzyme, ι-carrageenan could be transformed into a hybrid ι-/α-carrageenan or pure α-carrageenan. Analysis of the distribution of the carrabiose moieties in hybrid carrageenan chain using enzymatic degradation with Alteromonas fortis ι-carrageenase, coupled with chromatography and NMR spectroscopy experiments, showed that the sulfatase has an endo mode of action. The endo-character and the specificity of the sulfatase made it possible to prepare hybrid κ-/ι-/α-carrageenan and κ-/α-carrageenan starting from κ-/ι-carrageenan.  相似文献   

8.
α-Galactosylceramide (αGalCer, KRN7000) has been identified as a modulator of immunological processes through its capacity to bind iNKT cells mediated by CD1d molecules. Some analogues in while the amide group in αGalCer is replaced with ester or ether groups were synthesized from d-arabinitol or l-ribose to evaluate their ability to activate iNKT cells. Ester analogues 30a, 31a, and 61 showed activity for IFNγ and IL-4 production of iNKT cells, while ether (31b) and 4-methoxy ester (76) analogues of α-galactosylceramide were not active for iNKT cells.  相似文献   

9.
High molar mass random poly(butylene succinate-co-butylene sebacate), P(BS-co-BSe), and poly(butylene succinate-co-butylene adipate), P(BS-co-BA), with different composition, were synthesized and subjected to enzymatic hydrolysis by Lipase from Mucor miehei or from Rhizopus arrhizus. The enzymatic hydrolysis of P(BS-co-BSe)s and P(BS-co-BA)s films produced a mixture of water-soluble monomers and co-oligomers that were separated and identified by on-line high performance liquid chromatography/electrospray ionization mass spectrometry (HPLC/ESI-MS). Optimization of the HPLC analysis allowed the separation of isobar co-oligomers, differing only for the co-monomers sequence. Oligomers with the same monomer composition and molar mass but different sequence were identified by HPLC/ESI-MS-MS on-line analysis. The results obtained show a preferential hydrolytic cleavage induced by the lipases used. In particular, these enzymes prefer cleaving sebacic ester bonds in P(BS-co-BSe) copolymers, whereas succinic ester bonds appear to be hydrolyzed faster than adipic ester bonds in P(BS-co-BA) copolyesters. 1H NMR analysis further substantiates these findings. The primary products generated by lipase hydrolysis of polyester films underwent further degradation at longer reaction times. The HPLC/ESI-MS analysis of these mixtures at various times provided the first evidence that lipase catalysis is active also in water solution, a hydrophobic effect induced by the aliphatic units of these polyesters.  相似文献   

10.
Poly(lactide-co-glycolide) (PLGA) is extensively used in pharmaceutical applications, for example, in targeted drug delivery, because of biocompatibility and degradation rate, which is easily tuned by the copolymer composition. Nevertheless, synthesis of sugar-labeled amphiphilic copolymers with a PLGA backbone is quite a challenge because of high sensitivity to hydrolytic degradation. This Article reports on the synthesis of a new amphiphilic copolymer of PLGA grafted by mannosylated poly(ethylene oxide) (PEO). A novel building block, that is, α-methoxy-ω-alkyne PEO-clip-N-hydroxysuccinimide (NHS) ester, was prepared on purpose by photoreaction of a diazirine containing molecular clip. This PEO block was mannosylated by reaction of the NHS ester groups with an aminated sugar, that is, 2-aminoethyl-α-d-mannopyroside. Then, the alkyne ω-end-group of PEO was involved in a copper alkyne- azide coupling (CuAAC) with the pendent azides of the aliphatic copolyester. The targeted mannose-labeled poly(lactide-co-glycolide-co-ε-caprolactone)-graft-poly(ethylene oxide) copolymer was accordingly formed. Copolymerization of d,l-lactide and glycolide with α-chloro-ε-caprolactone, followed by substitution of chlorides by azides provided the azido-functional PLGA backbone. Finally, micelles of the amphiphilic mannosylated graft copolymer were prepared in water, and their interaction with Concanavalin A (ConA), a glyco-receptor protein, was studied by quartz crystal microbalance. This study concluded to the prospect of using this novel bioconjugate in targeted drug delivery.  相似文献   

11.
《Organogenesis》2013,9(4):212-216
Biodegradable polymers with high mechanical strength, flexibility and optical transparency, optimal degradation properties and biocompatibility are critical to the success of tissue engineered devices and drug delivery systems. Most biodegradable polymers suffer from a short half life due to rapid degradation upon implantation, exceedingly high stiffness, and limited ability to functionalize the surface with chemical moieties. This work describes the fabrication of microfluidic networks from poly(ester amide), poly(1,3-diamino-2-hydroxypropane-co-polyol sebacate) (APS), a recently developed biodegradable elastomeric poly(ester amide). Microfluidic scaffolds constructed from APS exhibit a much lower Young’s Modulus and a significantly longer degradation half-life than those of previously reported systems. The device is fabricated using a modified replica-molding technique, which is rapid, inexpensive, reproducible, and scalable, making the approach ideal for both rapid prototyping and manufacturing of tissue engineering scaffolds.  相似文献   

12.
The purpose of this study was to synthesize and study the in vitro enzymatic and non-enzymatic hydrolysis of indomethacin-TEG ester and amide prodrugs. It was found that the ester conjugate 10 was comparatively stable between pH 3 and 6 (half-life>90h), with a half-life equal to 5.2h in 80% buffered plasma. In contrast, the amide conjugate 12 appeared to be stable over the entire pH range studied with the only observed degradation being cleavage of the indolic N-4-chlorobenzoyl moiety.  相似文献   

13.
In this study, the transition process of the helix‐sense inversion of poly(β‐phenethyl‐L‐aspartate) was investigated by Raman scattering and 2‐dimensional correlation spectroscopy. Temperature‐dependent Raman spectra were obtained during the helix‐sense inversion. The results of 2‐dimensional correlation analysis in the spectral regions of 1600‐1800 and 3200‐3400 cm?1 showed that the intensity changes of the side‐chain ester C═O stretching bands occurred prior to those of amide A and amide I bands in the unwinding process of αR‐helix on heating. The sequential order of the intensity changes for amide A, amide I, and the side‐chain ester C═O stretching bands during the inversion process was determined. It was found that the conformation change of the side chain occurred prior to that of the main chain for the αR‐helix on heating. Thus, we concluded that the transformation of the backbone chain from right‐handed to left‐handed is triggered by the conformational change of the side chains.  相似文献   

14.
Maltosyl-α-cyclodextrin (6-α-maltosylcyclomaltohexaose, M-CD) was prepared from maltose and α-cyclodextrin by the reverse action of Bacillus pullulanase, and the action of α-amylases on this dextrin was examined. Among α-amylases tested, Thermoactinomyces vulgaris α-amylase (TVA) and Taka-amylase A (TAA) were found to attack the M-CD. Their action pattern on M-CD was studied. These α-amylases cleaved, first the cyclodextrin ring of M-CD, and the branched octasaccharides formed were immediately degraded to form glucose, branched tetraose, or pentaose, though the action pattern was different for TVA and TAA. In addition, TAA also split M-CD into glucose and glucosyl-α-cyclodextrin. Fission products at various stages of the reaction were separated and analyzed by paper chromatography and high performance liquid chromatography, their structures were analyzed, and the degradation pattern of M-CD was found.  相似文献   

15.
16.
H Cui  Y Liu  M Deng  X Pang  P Zhang  X Wang  X Chen  Y Wei 《Biomacromolecules》2012,13(9):2881-2889
Biodegradable poly(ester amide)s have recently been used as biomaterials due to their desirable chemical and biological characteristics as well as their mechanical properties, which are amendable for material processing. In this study, electroactive tetraaniline (TA) grafted poly(ester amide)s were successfully synthesized and characterized. The poly(ester amide)s-graft-tetraaniline copolymers (PEA-g-TA) exhibited good electroactivity, mechanical properties, and biodegradability. The biocompatibility of the PEA-g-TA copolymers in vitro was systematically studied, which demonstrated that they were nontoxic and led to favorable adhesion and proliferation of mouse preosteoblastic MC3T3-E1 cells. Moreover, the PEA-g-TA copolymers stimulated by pulsed electrical signal could serve to promote the differentiation of MC3T3-E1 cells compared with TCPs. Hence, the biodegradable and electroactive PEA-g-TA copolymers possessed the properties in favor of the long-time potential application in vivo (electrical stimulation directly to the desired area) as bone repair scaffold materials in tissue engineering.  相似文献   

17.
The aim of this work was to purify and characterize the extra-cellular leucine amino peptidase (LAP) from Streptomyces gedanensis and also study its applications for protein hydrolysis. The enzyme was purified to homogeneity by ammonium sulfate fractionation and sequential chromatography steps. LAP appeared to be a monomeric enzyme with a molecular weight of ~75 kDa determined by sodium dodecyl sulfate poly acryl amide gel electrophoresis (SDS-PAGE). The enzyme preferentially hydrolyzed leucine p-nitroanilide followed by Met, Phe, Lys and Arg derivatives. Kinetic studies on the purified enzyme confirmed that it can hydrolyze peptide as well as ester substrates at comparable rates. This amino peptidase was highly resistant to different concentrations of various organic solvents. The characteristics of this amino peptidase, including thermo stability, organic solvent resistance, its activity against various substrates, and also it showed esterase and peptidase activity at comparable rates; identified this amino peptidase as a novel one. The specificity towards aromatic and hydrophobic amino acid residues, the solvent-resistance and thermo stability make this amino peptidase could offer interesting possibilities for various industrial applications including debittering of protein hydrolysates, peptide and ester synthesis.  相似文献   

18.
Li X  Loh XJ  Wang K  He C  Li J 《Biomacromolecules》2005,6(5):2740-2747
Poly(ester urethane)s with poly[(R)-3-hydroxybutyrate] (PHB) as the hard and hydrophobic segment and poly(ethylene glycol) (PEG) as the soft and hydrophilic segment were synthesized from telechelic hydroxylated PHB (PHB-diol) and PEG using 1,6-hexamethylene diisocyanate as a nontoxic coupling reagent. Their chemical structures and molecular characteristics were studied by gel permeation chromatography, 1H NMR, and Fourier transform infrared spectroscopy. Results of differential scanning calorimetry and X-ray diffraction indicated that the PHB segment and PEG segment in the poly(ester urethane)s formed separate crystalline phases with lower crystallinity and a lower melting point than those of their corresponding precursors, except no PHB crystalline phase was observed in those with a relatively low PHB fraction. Thermogravimetric analysis showed that the poly(ester urethane)s had better thermal stability than their precursors. The segment compositions were calculated from the two-step thermal decomposition profiles, which were in good agreement with those obtained from 1H NMR. Water contact angle measurement and water swelling analysis revealed that both surface hydrophilicity and bulk hydrophilicity of the poly(ester urethane)s were enhanced by incorporating the PEG segment into PHB polymer chains. The mechanical properties of the poly(ester urethane)s were also assessed by tensile strength measurement. It was found that the poly(ester urethane)s were ductile, while natural source PHB is brittle. Young's modulus and the stress at break increased with increasing PHB segment length or PEG segment length, whereas the strain at break increased with increasing PEG segment length or decreasing PHB segment length.  相似文献   

19.
Guo K  Chu CC 《Biomacromolecules》2007,8(9):2851-2861
A new family of novel biodegradable poly(ether ester amide)s (PEEAs) consisting of three building blocks (L-phenylalanine, oligoethylene glycol, and aliphatic acid dichloride) were synthesized by solution polycondensation. Using N,N-dimethylacetamide as the solvent, these PEEA polymers were obtained with fairly good yields with reduced viscosity (eta(red)) ranging from 0.13 to 0.61 dL/g. The chemical structures of the PEEAs were confirmed by IR, NMR spectra, and elemental analysis. The PEEAs had Tg values lower than that of the saturated poly(ester amide)s (PEAs) of similar structures due to the incorporation of ether bonds in the backbones. An increase in the number of ether bonds in PEEA resulted in a lower Tg value. The solubility of the PEEA polymers in a wide range of common organic solvents was significantly improved when compared with unsaturated PEAs. The preliminary in vitro biodegradation behaviors of PEEA polymers were investigated in both pure PBS buffer and alpha-chymotrypsin solution of different concentrations. The polymers showed a significantly faster weight loss in an enzyme solution (alpha-chymotrypsin) but a very slow biodegradation rate in pure PBS buffer. The enzymatic hydrolysis rates of PEEAs (in terms of weight loss) were found to be much faster than those of saturated and unsaturated polyesteramides reported in previous studies. The zero-order-like biodegradation kinetics and molecular weight data also suggested surface erosion biodegradation mechanisms for these PEEAs.  相似文献   

20.
The design of new generation cardiovascular biomaterials focuses on biomimetic properties that are capable of eliciting specific cellular responses and directing new tissue formation. Synthetic poly(ester amide)s (PEAs) containing α-amino acid residues have the potential to elicit favorable cellular responses. Furthermore, they are biodegradable owing to the incorporation of naturally occurring amino acids. In this study, a family of PEAs was synthesized from selected α-amino acids using both solution and interfacial polymerization approaches to optimize their properties for vascular tissue engineering applications. By careful selection of the monomers and the polymerization approach, high-molecular-weight PEAs with low glass-transition temperatures were obtained. Human coronary artery smooth muscle cells (HCASMCs) cultured directly on bare PEA films attached and spread well up to 7 days of culture. Moreover, cell viability was significantly enhanced on all nonfunctional PEAs compared with tissue culture polystyrene controls. The trifluoroacetic acid salt of the lysine-containing functional PEAs was found to retard cell growth but still supported cell viability up to 5 days of culture. Immunostaining of HCASMCs revealed strong vinculin expression, suggesting that the HCASMCs initiated cellular processes for focal adhesion contacts with all PEA surfaces. Conversely, smooth muscle α-actin expression was not abundant on the PEA surfaces, suggesting a proliferative smooth muscle cell phenotype. Altogether, our results indicate that these PEAs are promising materials for vascular tissue engineering scaffolds.  相似文献   

设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号