Immobilization of lipases and assay in continuous fixed bed reactor

Lipases are versatile enzymes regarding the range of reactions they catalyse and substrates on which they act. They are as well important as catalyst in organic synthesis. Their immobilization on appropriate supports confer them greater stability besides the possibility of operating in continuous reactors. In order to explore these abilities, the reactions involving hydrolysis of p-nitrophenyl acetate (PNPA) and transesterification of PNPA with n-butanol were chosen. Lipases from two different sources were assayed, namely: microbial (Candida rugosa, CRL, Sigma Type VII) and pancreatic (PPL, Sigma, Type 11). Two immobilization methods were also used, namely: 1) adsorption, using as support the following silica derivatives (150-300μm e 450μ): phenyl, epoxy, amino and without derivation, and 2) covalent binding, using glutaraldehyde as binding agent and silica amino as support. This later method led to better results. Hydrolytic activity was 6.1 U/gsupport for CRL and 0.97U/gsupport for PPL, and of transesterification, 2,8U/gsupport for CRL and 1,9U/gsupport for PPL. Stability of the immobilized enzyme as a function of temperature was evaluated for CRL at 40°C and 50°C and for PPL at 32°C and 40°C. The assays were initially carried out batchwise, both for soluble and immobilized enzymes, aiming to the obtention of parameters for the continues reactor. Lipases immobilized by covalent binding were used in the assays of operacional stability in continuos reactors. For PPL in aqueous medium, at 32°C, and CRL in organic medium at 40°C, both operating continuously, no significant loss of activity was detected along the analysis period of 17 days. In the case of CRL in aqueous medium at 40°C there was a loss of activity around 40% after 18 days. For PPL in organic medium at 40°C the loss was 33% after 20 days. Compairing both sources with each other, very different results were obtained. Higher activitiy was found for CRL, both for hydrolysis and for transesterification reactions, with higher stability in organic medium. PPL showed lower activity as well as higher stability in aqueous medium. The immobilization method by covalent binding showed to be the most appropriate. Immobilized lipases are therefore relatively stable both in aqueous and organic medium.

Determination of cadmium in river water samples by flame AAS after on-line preconcentration in mini-column packed with 2-aminothiazole-modified silica gel

A rapid and sensitive method was developed to determine trace levels of Cd2+ ions in an aqueous medium by flame atomic absorption spectrometry, using on-line preconcentration in a mini-column packed with 100 mg of 2-aminothiazol modified silica gel (SiAT). The Cd2+ ions were sorbed at pH 5.0. The preconcentrated Cd2+ ions were directly eluted from the column to the spectrometer's nebulizer-burner system using 100 μL of 2 mol L-1 hydrochloric acid. A retention efficiency of over 95% was achieved. The enrichment factor (calculated as the ratio of slopes of the calibration graphs) obtained with preconcentrations in a mini-column packed with SiAT (A = -1.3 × 10-3 + 1.8 × 10-3 [Cd2+]) and without preconcentrations (A = 4 × 10-5 + 3.5 × 10-3[Cd2+]), was 51 and the detection limit calculated was 0.38 μg L-1. The preconcentration procedure was applied to determine trace levels of Cd in river water samples. The optimum preconcentration conditions are discussed herein.

Effect of ceramic surface treatment on the microtensile bond strength between a resin cement and an alumina-based ceramic

Purpose: The objective of this study was to test the following hypothesis: the silica coating on ceramic surface increases the bond strength of resin cement to a ceramic. Materials and Methods: In-Ceram Alumina blocks were made and the ceramic surface was treated: G1 - sandblasting with 110-μm aluminum oxide particles; G2 - Rocatec System: tribochemicai silica coating (Rocatec-Pre powder + Rocatec-Plus powder + Rocatec-Sil); G3 - CoJet System: silica coating (CoJet-Sand) + ESPE-Sil. The ceramic blocks were cemented to composite blocks with Panavia F resin cement (under a load of 750 g/1 min). The cemented blocks were stored in distilled water at 37°C for 7 days and sectioned along the x and y axes with a diamond disk. Samples with an adhesive area of ca 0.8 mm 2 (n = 45) were obtained. The samples were attached to an adapted device for the microtensile test, which was performed in a universal testing machine (EMIC) at a crosshead speed of 1 mm/min. Results: The obtained results were submitted to ANOVA and Tukey's test. Mean values of tensile strength (MPa) and standard deviation values were: (G1) 16.8 ± 3.2; (G2) 30.6 ± 4.5; (G3) 33.0 ± 5.0. G2 and 63 presented greater tensile strength than G1. There was no significant difference between G2 and G3. All the failures took place at the ceramic/resin cement interface. Conclusion: The silica coating (Rocatec or CoJet systems) of the ceramic surface increased the bond strength between the Panavia F resin cement and alumina-based ceramic.

Effect of surface treatments on the resin bond to zirconium-based ceramic

Purpose: This study tested the hypothesis that the tribochemical silica coating on ceramic surfaces increases the bond strength of resin cement to a glass-infiltrated zirconium-based ceramic. Materials and Methods: Fifteen blocks of In-Ceram Zirconia from CEREC InLab (5 per group) and 15 composite blocks (Z-250) 5 mm x 5 mm x 4 mm were made. The ceramic surfaces were polished, and the blocks were divided into three groups: (1) airborne abrasion with 110-μm aluminum oxide particles; (2) Rocatec system, tribochemical silica coating; and (3) CoJet system, tribochemical silica coating. The ceramic blocks were cemented to the composite blocks using Panavia F according to the manufacturer's specifications. All samples were stored in 37°C distilled water for 7 days and later sectioned in two axes using a diamond disk under cooling to obtain specimens with a cross-sectional area of approximately 1 mm2 (n = 45). Each specimen was then attached with cyanoacrylate glue to an adapted device for the microtensile test, which was carried out on a universal testing machine. Results: The results were subjected to ANOVA and Tukey's test. Group 2 (23.0 ± 6.7 MPa) and group 3 (26.8 ± 7.4 MPa) showed greater bond strength than group 1 (15.1 ± 5.3 MPa). There was no significant difference between groups 2 and 3. All failures were in the adhesive zone. Conclusion: The hypothesis was confirmed - the tribochemical systems increased the bond strength between Panavia F and In-Ceram Zirconia.

Evaluation of the flexural strength of carbon fiber-, quartz fiber-, and glass fiber-based posts

This study investigated the flexural strength of eight fiber posts (one carbon fiber, one carbon/quartz fiber, one opaque quartz fiber, two translucent quartz fiber, and three glass fiber posts). Eighty fiber posts were used and divided into eight groups (n = 10): G1: C-POST (Bisco); G2: ÆSTHETI-POST (Bisco); G3: ÆSTHETI-PLUS (Bisco); G4: LIGHT-POST (Bisco); G5: D.T. LIGHT-POST (Bisco); G6: PARAPOST WHITE (Coltene); G7: FIBERKOR (Pentron); G8: REFORPOST (Angelus). All of the samples were tested using the three-point bending test. The averages obtained were submitted to the ANOVA and to Tukey's test (p < 0.05). The mean values (MPa) of the groups ÆSTHETI-POST - carbon/ quartz fiber post (Bisco) and ÆSTHETI-PLUS - quartz fiber post (Bisco) were statistically similar and higher than the mean values of the other groups. The mean values of the groups C-POST - carbon fiber post (Bisco), LIGHT-POST - translucent quartz fiber post (Bisco), D.T. LIGHT-POST - double tapered translucent quartz fiber post (Bisco), PARAPOST WHITE - glass fiber post (Coltene) and FIBREKOR - glass fiber post (Pentron) were similar and higher than the group REFORPOST - glass fiber post (Angelus). Copyright © 2005 by the American Association of Endodontists.

On the specific filtration mechanism of a mesoporous silica membrane, prepared with non-connecting parallel pores

We report the singular filtration properties of an ultrafiltration membrane made with mesoporous silica that exhibits cylindrical pores aligned mostly normal to the support. This membrane supported on tubular commercial macroporous alumina supports was prepared by the interfacial growth mechanism between stable silica-surfactant hybrid micelles made of the association of silica oligomers with polyethyleneoxide-based (PEO) surfactants and sodium fluoride, a well-known silica condensation catalyst [Boissière et al., An ultrafiltration membrane made with mesoporous MSU-X silica, Chem. Mater. 15 (2003) 460-463]. It appears that the combined effect of the silica nature of the membrane, whose surface charge can be easily adjusted by changing the pH and the non-connected cylindrical shape of the pores provides a new behavior in the retention properties, as proved by the filtration of polyoxyethylene polymers (PEO) with different molecular weights. Depending on the filtration conditions, a rejection rate of 80% and a steep cut-off at 2000 Da can be obtained or, on the reverse, polymers three times bigger than the pore diameter can diffuse through the membrane. This new filtration mechanism, which opens up new modes of separation modes, is explained in the light of both topology of the porous network and pH-dependent interactions between PEO polymers and silica porous media. © 2004 Elsevier B.V. All rights reserved.

Adsorption of silanes bearing nitrogenated Lewis bases on SiO 2/Si (100) model surfaces

The present paper describes the one-pot procedure for the formation of self-assembled thin films of two silanes on the model oxidized silicon wafer, SiO2/Si. SiO2/Si is a model system for other surfaces, such as glass, quartz, aerosol, and silica gel. MALDI-TOF MS with and without a matrix, XPS, and AFM have confirmed the formation of self-assembled thin films of both 3-imidazolylpropyltrimethoxysilane (3-IPTS) and 4-(N- propyltriethoxysilane-imino)pyridine (4-PTSIP) on the SiO2/Si surface after 30 min. Longer adsorption times lead to the deposition of nonreacted 3-IPTS precursors and the formation of agglomerates on the 3-IPTS monolayer. The formation of 4-PTSIP self-assembled layers on SiO2/Si is also demonstrated. The present results for the flat SiO2/Si surface can lead to a better understanding of the formation of a stationary phase for affinity chromatography as well as transition-metal-supported catalysts on silica and their relationship with surface roughness and ordering. The 3-IPTS and 4-PTSIP modified SiO2/Si wafers can also be envisaged as possible built-on-silicon thin-layer chromatography (TLC) extraction devices for metal determination or N-heterocycle analytes, such as histidine and histamine, with on-spot MALDI-TOF MS detection. © 2005 Elsevier Inc. All rights reserved.

Effect of cyclic compressive loading on the bond strength of an adhesive system to dentin after collagen removal

Purpose: The objective of this study was to verify the effect of cyclic compressive loading on the shear bond strength of an adhesive system following collagen removal. Materials and Methods: Sixty bovine teeth were divided into 4 groups based on the adhesive procedure used: groups 1 and 2 - etching with 35% phosphoric acid and application of the Single Bond adhesive system; groups 3 and 4 - after etching, a 10% sodium hypochlorite solution was applied for 1 min before the application of the adhesive. In all the specimens, a Z100 resin cylinder was built up over the bond area. Groups 2 and 4 were submitted to 500,000 cycles with a load of 100 N. Results: The mean values for the shear bond test (MPa) were: group 1: 7.37 ± 1.15; group 2: 5.72 ± 1.66; group 3: 5.95 ± 1.21; group 4: 3.66 ± 1.12. There was no difference between groups 1 and 2 (p > 0.01). Between groups 1 and 3, 2 and 4, and 3 and 4 there was a significant difference (p < 0.01). The majority of the specimens demonstrated an adhesive failure. Conclusion: The application of sodium hypochlorite on dentin decreased the values of shear bond strength, as did the load cycling in the group treated with sodium hypochlorite.

Preconcentration and determination of mercury(II) at a chemically modified electrode containing 3-(2-thioimidazolyl)propyl silica gel

A mercury-sensitive chemically modified graphite paste electrode was constructed by incorporating modified silica gel into a conventional graphite paste electrode. The functional group attached to the (3-chloropropyl) silica gel surface was 2-mercaptoimidazole, giving a new product denoted by 3-(2-thioimidazolyl)propyl silica gel, which is able to complex mercury ions. Mercury was chemically adsorbed on the modified graphite paste electrode containing 3-(2-thioimidazolyl)propyl silica (TIPSG GPE) by immersion in a Hg(II) solution, and the resultant surface was characterized by cyclic and differential pulse anodic stripping voltammetry. One cathodic peak at 0.1 V and other anodic peak at 0.34 V were observed on scanning the potential from -0.1 to 0.8 V (0.01 M KNO3; ν = 2.0 mV s-1 νs. Ag/AgCl). The anodic peak at 0.34 V show an excellent sensitivity for Hg(II) ions in the presence of several foreign ions. A calibration graph covering the concentration range from 0.02 to 2 mg L-1 was obtained. The detection limit was estimated to be 5 μg L-1. The precision for six determinations of 0.05 and 0.26 mg L-1 Hg(II) was 3.0 and 2.5% (relative standard deviation), respectively. The method can be used to determine the concentration of mercury(II) in natural waters contaminated by this metal. 2005 © The Japan Society for Analytical Chemistry.

Effect of silica coating on flexural strength of fiber posts

Purpose: Fiber-reinforced composite (FRC) posts can be air-abraded to obtain good attachment to the resin cement. This study tested the effect of silica coating on the flexural strength of carbon, opaque, and translucent quartz FRC posts. Materials and Methods: Six experimental groups of FRC posts (n = 10 per group) were tested, either as received from the manufacturer or after chairside silica coating (30-μm CoJet-Sand). Results: There was no significant difference in the flexural strength of nonconditioned (504 to 525 MPa) and silica-coated (514 to 565 MPa) specimens (P > .05) (analysis of variance). The type of post did have a significant effect on flexural strength (P < .05). Conclusion: Chairside silica coating did not affect the flexural strength of both carbon and quartz FRC posts.

Microtensile bond strength of a resin cement to glass infiltrated zirconia-reinforced ceramic: The effect of surface conditioning

This study evaluated the effect of three surface conditioning methods on the microtensile bond strength of resin cement to a glass-infiltrated zirconia-reinforced alumina-based core ceramic. Thirty blocks (5×5×4 mm) of In-Ceram Zirconia ceramics (In-Ceram Zirconia-INC-ZR, VITA) were fabricated according to the manufacturer's instructions and duplicated in resin composite. The specimens were polished and assigned to one of the following three treatment conditions (n=10): (1) Airborne particle abrasion with 110 μm Al2O3 particles + silanization, (2) Silica coating with 110 μm SiOx particles (Rocatec Pre and Plus, 3M ESPE) + silanization, (3) Silica coating with 30 μm SiOx particles (CoJet, 3M ESPE) + silanization. The ceramic-composite blocks were cemented with the resin cement (Panavia F) and stored at 37 °C in distilled water for 7 days prior to bond tests. The blocks were cut under coolant water to produce bar specimens with a bonding area of approximately 0.6 mm2. The bond strength tests were performed in a universal testing machine (cross-head speed: 1 mm/min). The mean bond strengths of the specimens of each block were statistically analyzed using ANOVA and Tukey's test (α≤0.05). Silica coating with silanization either using 110 μm SiOx or 30 μm SiOx particles increased the bond strength of the resin cement (24.6±2.7 MPa and 26.7±2.4 MPa, respectively) to the zirconia-based ceramic significantly compared to that of airborne particle abrasion with 110-μm Al2O3 (20.5±3.8 MPa) (ANOVA, P<0.05). Conditioning the INC-ZR ceramic surfaces with silica coating and silanization using either chairside or laboratory devices provided higher bond strengths of the resin cement than with airborne particle abrasion using 110 μm Al2O3. © 2005 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.

Microtensile bond strength between a quartz fiber post and a resin cement: Effect of post surface conditioning

Purpose: To test the bond strength between a quartz-fiber-reinforced composite post (FRC) and a resin cement. The null hypothesis was that the bond strength can be increased by using a chairside tribochemical silica-coating system. Materials and Methods: Thirty quartz-FRCs (Light-Post) were divided into 3 groups according to the post surface treatment: G1) Conditioning with 32% phosphoric acid (1 min), applying a silane coupling agent; G2) etching with 10% hydrofluoric acid (1 min), silane application; G3) chairside tribochemical silica coating method (CoJet System): air abrasion with 30-μ SiO x-modified Al2O3 particles, silane application. Thereafter, the posts were cemented into a cylinder (5 mm diameter, 15 mm height) with a resin cement (Duo-Link). After cementation, the specimens were stored in distilled water (37°C/24 h) and sectioned along the x and y axes with a diamond wheel under cooling (Lab-cut 1010) to create nontrimmed bar specimens. Each specimen was attached with cyanoacrylate to an apparatus adapted for the microtensile test. Microtensile testing was conducted on a universal testing machine (1 mm/min). The data obtained were submitted to the one-way ANOVA and Tukey test (α = 0.05). Results: A significant influence of the conditioning methods was observed (p < 0.0001). The bond strength of G3 (15.14 ± 3.3) was significantly higher than the bond strengths of G1 (6.9 ± 2.3) and G2 (12.60 ± 2.8) (p = 0.000106 and p = 0.002631, respectively). Notwithstanding the groups, all the tested specimens showed adhesive failure between the resin cement and FRC. Conclusion: The chairside tribochemical system yielded the highest bond strength between resin cement and quartz-fiber post. The null hypothesis was accepted (p < 0.0001).

Fatigue resistance of the bond of a glass-infiltrated alumina ceramic to human dentin

Purpose: To evaluate the fatigue resistance of the bond between dentin and glass-infiltrated alumina ceramic, using different luting protocols. Materials and Methods: The null hypothesis is that the fatigue resistance varies with the luting strategy. Forty blocks of In-Ceram Alumina were prepared, and one surface of each block was abraded with 110-μm aluminum oxide particles. Then, the blocks were luted to flat dentin surfaces of 40 human third molars, using 4 different luting strategies (luting system [LS]/ceramic surface conditioning [CSC]) (n=10): (G1) [LS] RelyX-Unicem/[CSC] airborne abrasion with 110-μm Al2O3 particles; (G2) [LS] One-Step + Duo-Link (bis-GMA-based resin)/[CSC] etching with 4% hydrofluoric acid + silane agent; (G3) [LS] ED-Primer + Panavia F (MDP-based resin)/[CSC] Al2O 3; (G4) [LS] Scotchbond1+RelyX-ARC (bis-GMA-based resin)/[CSC] chairside tribochemical silica coating (air abrasion with 30-μm SiO x particles + silane). After 24 h of water storage at 37°C, the specimens were subjected to 106 fatigue cycles in shear with a sinusoidal load (0 to 21 N, 8 Hz frequency, 37°C water). A fatigue survivor score was given, considering the number of the fatigue cycles until fracture. The failure modes of failed specimens were observed in a SEM. Results: G3 (score = 5.9, 1 failure) and G4 (score = 6, no failures) were statistically similar (p = 0.33) and had significantly higher fatigue resistance than G1 (score = 3.9, 5 failures) and G2 (score = 3.7, 6 failures) (p < 0.03). SEM analysis of fractured specimens of G1 and G2 showed that almost all the failures were between ceramic and cement. Conclusion: The MDP-based resin cement + sandblasting with Al2O3 particles (G3) and bis-GMA-based resin cement + tribochemical silica coating (G4), both using the respective dentin bonding systems, were the best luting protocols for the alumina ceramic. The null hypothesis was confirmed.

Bond strength of a resin cement to high-alumina and zirconia-reinforced ceramics: The effect of surface conditioning

Purpose: The aim of this study was to evaluate the effect of two surface conditioning methods on the microtensile bond strength of a resin cement to three high-strength core ceramics: high alumina-based (In-Ceram Alumina, Procera AllCeram) and zirconia-reinforced alumina-based (In-Ceram Zirconia) ceramics. Materials and Methods: Ten blocks (5 ×6 × 8 mm) of In-Ceram Alumina (AL), In-Ceram Zirconia (ZR), and Procera (PR) ceramics were fabricated according to each manufacturer's instructions and duplicated in composite. The specimens were assigned to one of the two following treatment conditions: (1) airborne particle abrasion with 110-μm Al2O3 particles + silanization, (2) silica coating with 30 μm SiOx particles (CoJet, 3M ESPE) + silanization. Each ceramic block was duplicated in composite resin (W3D-Master, Wilcos, Petrópolis, RJ, Brazil) using a mold made out of silicon impression material. Composite resin layers were incrementally condensed into the mold to fill up the mold and each layer was light polymerized for 40 s. The composite blocks were bonded to the surface-conditioned ceramic blocks using a resin cement system (Panavia F, Kuraray, Okayama, Japan). One composite resin block was fabricated for each ceramic block. The ceramic-composite was stored at 37°C in distilled water for 7 days prior to bond tests. The blocks were cut under water cooling to produce bar specimens (n = 30) with a bonding area of approximately 0.6 mm2. The bond strength tests were performed in a universal testing machine (crosshead speed: 1 mm/min). Bond strength values were statistically analyzed using two-way ANOVA and Tukey's test (≤ 0.05). Results: Silica coating with silanization increased the bond strength significantly for all three high-strength ceramics (18.5 to 31.2 MPa) compared to that of airborne particle abrasion with 110-μm Al2O3 (12.7-17.3 MPa) (ANOVA, p < 0.05). PR exhibited the lowest bond strengths after both Al2O3 and silica coating (12.7 and 18.5 MPa, respectively). Conclusion: Conditioning the high-strength ceramic surfaces with silica coating and silanization provided higher bond strengths of the resin cement than with airborne particle abrasion with 110-μm Al2O3 and silanization.

Bond strength of acrylic teeth to denture base resin after various surface conditioning methods before and after thermocycling

This study aimed to evaluate the durability of adhesion between acrylic teeth and denture base acrylic resin. The base surfaces of 24 acrylic teeth were flatted and submitted to 4 surface treatment methods: SM1 (control): No SM; SM2: application of a methyl methacrylate-based bonding agent (Vitacol); SM3: air abrasion with 30-μm silicone oxide plus silane; SM4: SM3 plus SM2. A heat-polymerized acrylic resin was applied to the teeth. Thereafter, bar specimens were produced for the microtensile test at dry and thermocyled conditions (60 days water storage followed by 12,000 cycles). The results showed that bond strength was significantly affected by the SM (P < .0001) (SM4 = SM2 > SM3 > SM1) and storage regimens (P < .0001) (dry > thermocycled). The methyl methacrylate-based adhesive showed the highest bond strength.