Amperometric determination of choline with enzyme immobilized in a hybrid mesoporous membrane

Choline sensor is successfully prepared by using immobilized enzyme, i.e., choline oxidase (ChOx) within a hybrid mesoporous membrane with 12 nm pore diameter (F127M). The measurement was based on the detection of hydrogen peroxide, which is the co-product of the enzymatic choline oxidation. The determination range and the response time are 5.0-800 μM and approximately 2 min, respectively. The sensor is very stable compared to the native enzyme sensor and 85% of the initial response was maintained even after storage for 80 days. These results indicate that ChOx is successfully immobilized and well stabilized, and at the same time, enzyme reaction proceeds efficiently. Such ability of hybrid mesoporous membrane F127M suggests great promise for effective immobilization of enzyme useful for electrochemical biosensors.     

Synthesis and characterization of spirobifluorene‐based polyimides

The synthesis and properties of organosoluble aromatic polyimides, containing spiro‐skeletal units in the polymer backbone on the basis of the spiro‐diamine monomer, 2,2′‐diamino‐9,9′‐spirobifluorene, are described. In the case of the spiro segment, the two fluorene rings are orthogonally arranged and connected through a tetrahedral bonding carbon atom, the spiro center. As a consequence, the polymer chain is periodically zigzagged with a 90° angle at each spiro center. This structural feature minimizes interchain interactions and restricts the close packing of the polymer chains, resulting in amorphous polyimides that have good solubility in organic solvents. Compared with their fluorene‐based cardo analogues, the spirobifluorene‐based polyimides have an improved solubility. Furthermore, the main‐chain rigidity of the polyimide appears to be preserved because of the presence of the spiro structure, which restricts the free segmental mobility. As a result, these polyimides exhibit a high glass‐transition temperature (T_g's) and good thermal stability. The T_g's of these polyimides were in the range of 287–374 °C, and the decomposition temperatures in nitrogen for a 10% weight loss occurred at temperatures above 570 °C. © 2002 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 40: 3615–3621, 2002     

Synthesis,characterization and memory properties of novel organosoluble polyimides

Two novel series of aromatic polyimides were prepared from 2,2′‐bis(4‐amino‐3‐trifluoromethylthylphenoxy) biphenyl, 2,2′‐bis(4‐amino‐3‐methoxyphenoxy)biphenyl with 3,3′,4,4′‐benzophenonetetracarboxylic dianhydride (BTDA), 3,3′,4,4′‐diphenylsulfonetetracarboxylic dianhydride (DSDA) via a one‐step procedure. The resulting polymers were fully characterized, and they exhibited excellent organosolubility. These polyimides are thermally stable with 5% weight loss over 430°C, and glass transition temperatures of the polyimides were found to be 225–262°C. Resistive switching devices with the configuration of Al/polymer/indium‐tin oxide were constructed from these polyimides by using conventional solution coating process. Devices with all polyimides exhibited nonvolatile and rewritable flash type memory characteristics with turn‐on voltage at ?1.1 to ?2.8 V. The ON/OFF current ratio of these devices was larger than 10⁴, and the retention times can be as long as 10⁴ s. The theoretical simulation based on the density functional theory suggested that greater distinct charge separation between the ground and charge transfer states led to a highly stable memory behavior. Copyright © 2016 John Wiley & Sons, Ltd.     

Synthesis and characterization of soluble fluorine‐containing polyimides based on 1,4‐bis(4‐amino‐2‐trifluoromethylphenoxy)benzene

The synthesis and properties of a class of soluble fluorine‐containing aromatic polyimides are described. Substituents of trifluoromethyl groups on the aromatic rings of paralinked aromatic ether diamine conferred the polymer prepared thereof with enhanced solubility, low‐moisture absorption, and low dielectric constants. The polyimides also exhibited exceptional thermal stability, good mechanical properties, and excellent hygrothermal resistance. These outstanding combined features ensure the polymers are desirable candidate materials for advanced microelectronics applications. © 2001 John Wiley & Sons, Inc. J Polym Sci Part A: Polym Chem 39: 2581–2590, 2001     

Synthesis and characterization of highly soluble aromatic polyimides

Two highly soluble aromatic polyimides were synthesized successfully from a diamine with two tert-butyl groups （MBTBA）and dianhydrides with a thioether or sulfone moiety（DTDA and DSDA）.Both of them showed excellent solubility in common solvents such as chloroform,tetrahydrofuran and dioxane at the room temperature.The number-average molecular weight was 6.0×10⁴ and 8.3×10⁴ according to gel permeation chromatography relative to a polystyrene standard,and the polydispersity index was 1.80 and 1.82 respectively.The glass-transition temperatures of them were 286℃and 314℃（or 315℃and 358℃）respectively,as measured by differential scanning calorimetry（or dynamic mechanical analysis）.The 5%weight loss temperature of both was near 490℃in N₂ by thermogravimetric analysis.These results indicated that the tert-butyl pendent groups reduced the interactions among polymer chains and the thioether or sulfone moiety was flexible which may improve their solubility in conventional organic solvents without the loss of thermal stability. Transparent and flexible films of the two polyimides were obtained via solution casting.The MBTBA-DTDA membrane had higher storage moduli than those of the MBTBA-DSDA membrane.     

Synthesis of thianthrene‐5,5,10,10‐tetraoxide‐containing polyimides via Yamazaki–Higashi phosphorylation method and their pervaporation properties to aromatic/nonaromatic hydrocarbon mixtures

High molecular weight polyimide was successfully prepared from thianthrene‐2,3,7,8‐tetracarboxylic acid‐5,5,10,10‐tetraoxide (TADATO‐4A) and 3,7‐diamino‐2,8(6)‐dimethyldibenzothiophene sulfone (DDBT) via the Yamazaki–Higashi phosphorylation method in the presence of triphenyl phosphite (TPP) and pyridine (Py). The obtained polyimide showed about 3–4 times larger inherent viscosity than that prepared by the conventional two‐step method in which the anhydride form (TADATO) of TADATO‐4A was used. The combination of the conventional two‐step method and Yamazaki–Higashi phosphorylation method, in which a dianhydride monomer [3,3′,4,4′‐diphenylsulfonetetracarboxylic dianhydride (DSDA)] was allowed to react with excessive DDBT to form an amine end‐capped polyamic acid oligomer and subsequently the oligomer was further polymerized with TADATO‐4A in the presence of TPP and Py, succeeded in giving the high molecular weight copolyimide, TADATO/DSDA(1/1)‐DDBT. However, both TADATO‐DDBT and TADATO/DSDA(1/1)‐DDBT showed fairly poor thermal stability due to the highly rigid structures. The pervaporation (PV) properties of the prepared polyimide membranes for benzene/cyclohexane (Bz/Cx) and benzene/n‐hexane (Bz/n‐Hx) mixtures were investigated. TADATO‐DDBT showed similar PV performance to DSDA‐DDBT at 60 °C. The sorption measurement revealed that these two kinds of polyimide membranes had a similar sorption amount, solubility selectivity, and diffusivity selectivity. The PV performance of TADATO/DSDA(1/1)‐DDBT was also found similar to DSDA‐DDBT for Bz/Cx mixture. © 2000 John Wiley & Sons, Inc. J Polym Sci A: Polym Chem 38: 895–906, 2000     

Synthesis and characterization of novel sulfonated naphthalenic polyimides as proton conductive membrane for DMFC applications

To prepare proton conductive membrane for direct methanol fuel cells (DMFC), a novel sulfonated aromatic diamine monomer, 1,4-bis(4-amino-2-sulfonic acid-phenoxy)-benzene (DSBAPB) was synthesized and characterized by ¹H NMR and FT-IR. Then a series of sulfonated polyimides (SPIs) were prepared from DSBAPB with 1,4,5,8-naphthalene tetracarboxylic dianhydride (NTDA) and a non-sulfonated diamine, 4,4′-oxydianiline (ODA) via one-step high-temperature polymerization method. The sulfonation degree of the SPIs can be controlled by changing the mole ratio of sulfonated monomer to non-sulfonated monomer. The obtained SPI membranes exhibit desirable proton conductivity ranged from 7.9 × 10⁻³ to 7.2 × 10⁻² S cm⁻¹ and low methanol permeability of less than 2.85 × 10⁻⁷ cm² s⁻¹. Furthermore, the hydrolysis stability of the obtained SPIs is better than the BDSA based SPIs caused by the flexible structure.     

Synthesis and characterization of epoxy‐based semi‐interpenetrating polymer networks sulfonated polyimides proton‐exchange membranes for direct methanol fuel cell applications

Novel epoxy‐based semi‐interpenetrating polymer networks (semi‐IPNs) of aromatic polyimide, derived from 2,2‐benzidinedisulfonic acid (BDSA), were prepared through a thermal imidization reaction. Dynamic scanning calorimetry (DSC) and Fourier transform infrared spectroscopy (FTIR) were utilized to verify the synchronization of the imidization of sulfonated poly(amic acid) (SPAA) and the crosslinking reactions of epoxy. The semi‐IPNs of epoxy/sulfonated polyimides (SPI‐EPX) exhibit excellent film‐forming characteristics and mechanical integrity at room temperature. Conductivities at 100 °C of 0.0243 S cm^?1 (SPI‐EP30) and 0.0141 S cm^?1 (SPI‐EP50) were obtained, which are similar to that of the Nafion 117 (0.0287 S cm^?1). The increase in the conductivity of SPI‐EP(30,40) with temperature is more rapid than that of Nafion 117. The SPI‐EPX exhibited lower methanol permeability than did Nafion117. The hydrolytic stability of the SPI‐EPX was followed by FTIR spectroscopy at regular intervals. SPI‐EPX prepared using epoxy‐based semi‐IPNs of sulfonated polyimide, SPI‐EP(40,50), exhibited higher hydrolytic stability than the phthalic polyimides (five‐membered ring polyimides).The microstructure was analyzed using atomic force microscopy (AFM) in the tapping mode, which demonstrated that SPI‐EP50 exhibited a nanophase that was separated into an essentially reticulated and venous hydrophobic and hydrophilic domains. Transmission electron microscopy (TEM) confirmed widespread and well‐connected hydrophilic domains, proving the higher hydrolytic stability and strong proton‐transporting properties of the SPI‐EPX membrane. © 2008 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 46: 2262–2276, 2008     

Aldehyde oxidase assay by capillary electrophoresis: From off‐line,online up to immobilized enzyme reactor

Capillary electrophoresis is a modern separation technique characterized by many benefits, which qualify it also for enzyme assays and the study of enzyme kinetics during drug development. Homogeneous or heterogeneous approaches can be followed for the enzymatic incubation. In this study, an immobilization procedure of aldehyde oxidase on magnetic particles was developed considering their integration with capillary electrophoresis. A number of magnetic nano/microparticle types were tested for this purpose, showing that aldehyde oxidase was most active when immobilized on bare silica magnetic nanoparticles. Primarily, the reusability of the enzyme immobilized on bare silica nanoparticles was tested. Three consecutive incubations with substrate could be performed, but the activity considerably dropped after the first incubation. One reason could be an enzyme detachment from the nanoparticles, but no release was detected neither at 4°C nor at 37°C during 5 h. The drop in enzymatic activity observed in consecutive incubations, could also be due to inactivation of the enzyme over time at given temperature. For the immobilized enzyme stored at 4°C, the activity decreased to 83% after 5 h, in contrast with a steep decrease at 37°C to 37%.     

Synthesis and properties of novel sulfonated polyimides from 2,2′‐bis(4‐aminophenoxy)biphenyl‐5,5′‐disulfonic acid

Novel sulfonated polyimides (SPIs) were prepared from 1,4,5,8‐naphthalenetetracarboxylic dianhydride (NTDA), 2,2′‐bis(4‐aminophenoxy)biphenyl‐5,5′‐disulfonic acid (oBAPBDS) with nonlinear configuration, and common nonsulfonated diamines. Water uptake (WU) in liquid and vapor, water stability, and proton conductivity σ of the resulting SPI membranes were investigated. They were soluble in m‐cresol and dimethylsulfoxide, and their WUs in liquid were much larger than those of the SPIs from other sulfonated diamines with linear configuration such as 4,4′‐bis(4‐aminophenoxy)biphenyl‐3,3′‐disulfonic acid (BAPBDS). NTDA‐oBAPBDS membrane was soluble in water at room temperature, whereas all the oBAPBDS‐based copolyimide membranes were insoluble in water and maintained mechanical strength after being soaked in distilled water at 80 °C for 40–1000 h. This much improved water stability was due to the enhanced solubility stability of membrane toward water. The water vapor sorption isotherms were rather similar between the SPIs with the nonlinear and linear configurations of sulfonated diamine moieties. The present SPIs with IECs of 1.8–2.6 meq/g, including NTDA‐BAPBDS, showed reasonably high proton conductivities under the highly humid conditions and roughly fell on the same σ–WU relation line. © 2004 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 42: 1432–1440, 2004     

Synthesis and characterization of fluorinated polyimides derived from novel unsymmetrical diamines

Two kinds of aromatic, unsymmetrical diamines with ether-ketone group, 3-amino-4′-(4-amino-2-trifluoromethylphenoxy)-benzophenone and 4-amino-4′-(4-amino-2-trifluoromethylphenoxy)-benzophenone, were successfully synthesized with two different synthetic routes. Then, they were polymerized with 4,4′-oxydiphthalic anhydride, 3,3′,4,4′-benzophenone tetracarboxylic dianhydride, and 2,2′-bis(3,4-dicarboxyphenyl)-hexafluoropropane dianhydride to form a series of fluorinated polyimides via a conventional two-step thermal or chemical imidization method. The resulting polyimides were characterized by measuring their solubility, viscosity, mechanical properties, IR-FT, and thermal analysis. The results showed that the polyimides had inherent viscosities of 0.48-0.68 dl/g and were easily dissolved in bipolarity solvents and common, low-boiling point solvents. Meanwhile, the resulting strong and flexible polyimide films exhibited excellent thermal stability, e.g., decomposition temperatures (at 10% weight loss) are above 575 °C and glass-transition temperatures in the range of 218-242 °C. The polymer films also showed outstanding mechanical properties, such as tensile strengths of 86.5-132.8 MPa, elongations at break of 8-14%, and initial moduli of 1.32-1.97 GPa. These outstanding combined features ensure that the polymers are desirable candidate materials for advanced applications.     

Synthesis and characterization of novel perfluorinated polyimides with 3D‐controlled structures for photonic applications

We have designed and synthesized novel perfluorinated polyimides with 3D controlled structure. At first, we successfully developed the new synthetic routes to diamines with pendant bulky perfluorinated aromatic units in a multi‐step synthetic procedure. Novel perfluorinated polyimides were prepared in a two‐step reaction in N‐methyl‐2‐pyrrolidinone (NMP) solution: The first step was for the synthesis of polyamic acids (PAAs) and the second reaction was for the imidization of PAA. The polymer yield was over 89% and the inherent viscosity of PAAs was in the range of 0.24–0.36 dL/g. The thin films were prepared by spin‐coating the PAA solution in NMP onto various substrates such as a Si wafer or a KBr pellet, dried at 80 °C and further cured at 230 °C. The resultant polyimides are thermally stable over 400 °C. The refractive index and birefringence of the resultant polyimides are 1.5858–1.6452 and 0.01–0.005, respectively. The refractive index of polyimide decreases with increasing the fluorine content. The copolymerization and the ether linkages into the backbone reduce the birefringence of polyimides. Surprisingly, the pendant ether linkage is not a crucial factor in reducing the polyimide birefringence. Their birefringence is comparatively very low, compared with that of previous polyimides. © 2005 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 44: 1326–1342, 2006     

Synthesis and characterization of soluble polyimides based on trifluoromethylated aromatic dianhydride and substitutional diaminetriphenylmethanes

A series of novel and soluble polyimides containing fluorine were synthesized by the polycondensation of fluorine-containing aromatic dianhydride TFDA with substitutional diaminetriphenylmethanes TDPT, FTDPT and 3FTDPT. The composition, structure and properties of the resulting polyimides were studied by means of FT-IR, DSC, TGA and elemental analysis methods, as well as general characterization methods that test solubility or viscosity. The results show that all the novel polyimides were obtained in quantitative yields with inherent viscosities of 0.70-0.76 dL/g, and showed excellent solubility in common organic solvents, such as NMP, DMAc, DMF, DMSO, THF, m-cresol, chloroform and 4-butyrolactone. Meanwhile, their T_g values from DSC are in the range of 265-293 °C, the temperature of 5 and 10% weight loss from TGA are in the range of 460-465 and 513-524 °C in N₂, respectively.     

An optical glucose biosensor based on glucose oxidase immobilized on a swim bladder membrane

An optical glucose biosensor using a swim bladder membrane as an enzyme immobilization platform and an oxygen-sensitive membrane as an optical oxygen transducer has been developed. During the enzymatic reaction, glucose is oxidized by glucose oxidase with a concomitant consumption of dissolved oxygen resulting in an increase in the fluorescence intensity of the optical oxygen transducer. The fluorescence intensity is directly related to the glucose concentration. The effects of pH, temperature, buffer concentration, and selectivity have been studied in detail. The immobilized enzyme retained 80% of its initial activity after being kept for more than 10 months at 4°C. The glucose biosensor has been successfully applied to the determination of glucose content in human blood serum and urine samples. Martin M.F. Choi was on sabbatical leave at The University of North Carolina at Chapel Hill from July 2004 to July 2005.     

Synthesis and FET characterization of novel ambipolar and low‐bandgap naphthalene‐diimide‐based semiconducting polymers

A novel series of naphthalene‐diimide‐based semiconducting polymers ( P1–P4 ) containing benzodithiophene or dithienopyrrole were successfully synthesized for ambipolar semiconducting materials showing near infrared absorptions. The incorporation of a 3‐hexylthiophene (3HT) spacer extended the intramolecular charge‐transfer (ICT) peak from λ_onset = 739 nm ( P1 ) to 785 nm ( P3 ). Moreover, about 250 nm red‐shift of the ICT peaks was observed in P2 and P4 compared to P1 and P3 due to the increased high‐lying HOMO energy levels. The grazing incidence X‐ray scattering of the P3 and P4 films proved the slightly improved crystalline order in the π?π stacking direction, indicating that the planar backbone is probably due to the introduced 3HT. The P1–P4 ‐based field‐effect transistor showed n‐type dominant ambipolar characteristics. The P2 and P4 showed higher electron mobilities up to 1.5 × 10^?2 cm² V^?1 s^?1 than P1 and P3 , which might be influenced by the orientation of the polymer backbone and the intermolecular orbital overlap. © 2015 Wiley Periodicals, Inc. J. Polym. Sci., Part A: Polym. Chem. 2016 , 54, 359–367     

Synthesis and properties of organosoluble polyimides based on 4,4′‐bis(4‐amino‐2‐trifluoromethylphenoxy)benzophenone

A novel, fluorinated diamine monomer with the ether–ketone group, 4,4′‐bis(4‐amino‐2‐trifluoromethylphenoxy)benzophenone ( 2 ), was prepared through the nucleophilic substitution reaction of 2‐chloro‐5‐nitrobenzotrifluoride and 4,4′‐dihydroxybenzophenone in the presence of potassium carbonate, followed by catalytic reduction with hydrazine and Pd/C. Flourinated polyimides (PIs) 5a – f and copolyimides (co‐PIs) 5c / a – f were synthesized from 2 and various commercial aromatic dianhydrides via thermal or chemical imidization. PIs 5a – f had inherent viscosities ranging from 0.72 to 1.22 dL/g. Besides the chemical imidization of 5c ( C ), the 5 ( C ) series were soluble in amide‐type solvents and even in less polar solvents, but PIs 5a – f prepared via thermal imidization were insoluble. PI films 5a – f exhibited tensile strengths ranging from 92 to 112 MPa, elongations at break from 8 to 15%, and initial moduli from 2.0 to 2.1 GPa. The glass‐transition temperatures of the 5 series were in the range of 232–278 °C, and the 10% weight‐loss temperatures were above 535 °C, with more than a 50% char yield at 800 °C in nitrogen. In comparison of the PI 5 series with the analogous non‐fluorinated PIs 6 series based on 4,4′‐bis(4‐aminophenoxy)benzophenone, the 5 series revealed better solubility, lower color intensity, dielectric constant, and moisture absorption. Their PI films had cutoff wavelengths between 370 and 410 nm, b* values ranging from 9.6 to 58.3, dielectric constants of 3.05–3.64 (1 MHz), with moisture absorption in the range of 0.08–0.38 wt %. © 2003 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 42: 222–236, 2004     

Chitosan‐Glutamate Oxidase Gels: Synthesis,Characterization, and Glutamate Determination

The biopolymer chitosan (CHIT) was chemically modified with glutaric dialdehyde (GDI) and used for the covalent immobilization of enzyme glutamate oxidase (GmOx). The relationships between the loaded, retained, and active units of GmOx in the CHIT‐GDI‐GmOx gels were determined by electrochemical assays. The latter indicated that on average ca. 95% of the GmOx was retained in the CHIT‐GDI matrix that was loaded with 0.10–3.0 units of the enzyme. The maximum activity of the GmOx immobilized in the gels corresponded to ca. 5% of the activity of the free enzyme. Platinum electrodes coated with CHIT‐GDI‐GmOx gels (films) were used as amperometric biosensors for glutamate. Such biosensors displayed good operational and long‐term stability (at least 11 h and 100 days, respectively) in conjunction with low detection limit of 0.10 μM glutamate (S/N=3), linear range up to 0.5 mM (R²=0.991), sensitivity of 100 mA M^?1 cm^?2, and short response time (t_90%=2 s). This demonstrated an efficient signal transduction in the Pt/CHIT‐GDI‐GmOx+glutamate system. The CHIT‐GDI‐GmOx gels constitute a new biosensing element for the development of glutamate biosensors.     

Polyacrylonitrile enzyme ultrafiltration and polyamide enzyme microfiltration membranes prepared by diffusion and convection

Glucose oxidase (GOD) and catalase (CAT) were covalently immobilized onto three types of polyacrylonitrile (PAN 1, PAN 2, and PAN 3) ultrafiltration (UF) membranes with different pore sizes and one type of polyamide (PA) microfiltration (MF) membrane by the bifunctional reagent, glutaraldehyde. The initial membranes were pre-modified to generate active amide groups in the PAN membranes and active amino groups in the PA membranes. The PAN 3 membrane contained the highest amount of active groups, and the membrane PA the lowest. The modified membranes were enzyme-loaded by diffusion and convection (UF). The effect of membrane pore size and immobilization methods on enzymatic activity and bound protein were studied. The most effective immobilized system was prepared by diffusion using a PAN 3 membrane as a carrier (bound protein: 0.055 mg/cm(2), relative activity: 87.6%). This membrane had the highest pore size of all the PAN membranes. Despite the highest pore size of PA membrane, the enzyme PA membranes prepared by diffusion showed the lowest amount of bound protein (0.03 mg/cm(2)) and the lowest relative activity (35.38%). This correlates with the lowest amount of active groups found in these membranes. The relative activity was higher for all the enzyme systems loaded by diffusion. The systems prepared by convection of the enzyme solution contained higher amounts of enzymes (0.035-0.13 mg/cm(2) protein), which led to internal substrate diffusion resistance and a decrease in the GOD relative activity (21.55-68.5%) in these systems. The kinetic parameters (V(max) and K(m)) and the glucose conversion of the immobilized systems prepared by diffusion were also studied. [diagram in text].     

Synthesis and characterization of high refractive index polyimides derived from 2,5‐Bis(4‐Aminophenylenesulfanyl)‐3,4‐Ethylenedithiothiophene and aromatic dianhydrides

The authors describe the synthesis and characterization of the polyimide (PI) series containing a 2,5‐bis(4‐aminophenylenesulfanyl)‐3,4‐ethylenedithiothiophene (APSEDTT) moiety in their main chain. The APSEDTT monomer with high sulfur content was prepared and polymerized with several aromatic dianhydrides such as 4,4′‐[p‐thio bis(phenylenesulfanyl)]diphthalic anhydride (3SDEA), 4,4′‐biphthalic anhydride (BPDA), and 4,4′‐oxydiphthalic anhydride (ODPA) by the traditional two‐step polycondensation procedure. All PIs exhibited high transparency, higher than 75% at 550 nm for a thickness of about 20 μm and good thermal properties such as thermal decomposition temperatures (T_10%) in the range of 409–521 °C. In addition, the PIs have extraordinarily excellent optical properties in refractive index and birefringence as originally designed. In particular, the PI derived from APSEDTT and 3SDEA showed a high refractive index (1.7586), and a low birefringence (0.0087) because of their very high sulfur content (27.7%). © 2015 Wiley Periodicals, Inc. J. Polym. Sci., Part A: Polym. Chem. 2015 , 53, 944–950     

Dual‐enzyme,co‐immobilized capillary microreactor combined with substrate recycling for high‐sensitive glutamate determination based on CE

A new method for high‐sensitive determination of glutamate was developed and evaluated based on CE by using dual‐enzyme co‐immobilized capillary microreactor combined with substrate recycling. The capillary microreactor was prepared by covalently co‐immobilizing glutamate dehydrogenase (GDH) and glutamic pyruvic transaminase (GPT) on the inner surface of a capillary and was characterized by SEM, ultraviolet‐visible spectroscopy, and fluorescence spectroscopy. The GDH‐GPT co‐immobilized capillary microreactor showed great stability and reproducibility. The apparent K_m for glutamate with GDH‐GPT coupled reaction was determined to be 0.61±0.06 mM but 2.56±0.24 mM when only GDH was immobilized. Glutamate determination was based on on‐column monitoring UV absorption at 340 nm of the reaction product reduced nicotinamide adenine dinucleotide, of which peak area was directly related to the glutamate concentration. The response of the present co‐immobilized GDH‐GPT assay for glutamate is greatly enhanced over single enzyme system, and a 15.7‐fold improvement in sensitivity was obtained. The detection limit of the proposed method is 0.15 μM glutamate (S/N=3). Selectivity for glutamate is good over most of the 20 amino acids. Finally, this method was successfully applied to determine the glutamate content in rat plasma and serum samples.