Comparisons of immersion and electrochemical properties of highly biocompatible Ti–15Zr–4Nb–4Ta alloy and other implantable metals for orthopedic implants

Metal release from implantable metals and the properties of oxide films formed on alloy surfaces were analyzed, focusing on the highly biocompatible Ti–15Zr–4Nb–4Ta alloy. The thickness and electrical resistance (R_p) of the oxide film on such an alloy were compared with those of other implantable metals. The quantity of metal released during a 1-week immersion test was considerably smaller for the Ti–15Zr–4Nb–4Ta than the Ti–6Al–4V alloy. The potential (E₁₀) indicating a current density of 10 μA cm⁻² estimated from the anodic polarization curve was significantly higher for the Ti–15Zr–4Nb–4Ta than the Ti–6Al–4V alloy and other metals. Moreover, the oxide film (4–7 nm thickness) formed on the Ti–15Zr–4Nb–4Ta surface is electrochemically robust. The oxide film mainly consisted of TiO₂ with small amounts of ZrO₂, Nb₂O₅ and Ta₂O₅ that made the film electrochemically stable. The R_p of Ti–15Zr–4Nb–4Ta was higher than that of Ti–6Al–4V, i.e. 0.9 Ω cm² in 0.9% NaCl and 1.3 Ω cm² in Eagle''s medium. This R_p was approximately five-fold higher than that of stainless steel, which has a history of more than 40 years of clinical use in the human body. Ti–15Zr–4Nb–4Ta is a potential implant material for long-term clinical use. Moreover, E₁₀ and R_p were found to be useful parameters for assessing biological safety.     

An enzyme catalyzing O-prenylation of the glucose moiety of fusicoccin A, a diterpene glucoside produced by the fungus Phomopsis amygdali

Isoprenoids form the largest family of compounds found in nature. Isoprenoids are often attached to other moieties such as aromatic compounds, indoles/tryptophan, and flavonoids. These reactions are catalyzed by three phylogenetically distinct prenyltransferases: soluble aromatic prenyltransferases identified mainly in actinobacteria, soluble indole prenyltransferases mostly in fungi, and membrane‐bound prenyltransferases in various organisms. Fusicoccin A (FC A) is a diterpene glycoside produced by the plant‐pathogenic fungus Phomopsis amygdali and has a unique O‐prenylated glucose moiety. In this study, we identified for the first time, from a genome database of P. amygdali, a gene (papt) encoding a prenyltransferase that reversibly transfers dimethylallyl diphosphate (DMAPP) to the 6′‐hydroxy group of the glucose moiety of FC A to yield an O‐prenylated sugar. An in vitro assay with a recombinant enzyme was also developed. Detailed analyses with recombinant PAPT showed that the enzyme is likely to be a monomer and requires no divalent cations. The optimum pH and temperature were 8.0 and 50 °C, respectively. K_m values were calculated as 0.49±0.037 μM for FC P (a plausible intermediate of FC A biosynthesis) and 8.3±0.63 μM for DMAPP, with a k_cat of 55.3±3.3×10^?3 s. The enzyme did not act on representative substrates of the above‐mentioned three types of prenyltransferase, but showed a weak transfer activity of geranyl diphosphate to FC P.     

Engineering of novel tamavidin 2 muteins with lowered isoelectric points and lowered non-specific binding properties

The avidin-biotin interaction is widely employed as a universal tool in numerous biotechnological applications. In avidin-biotin technology, non-specific binding to biological macromolecules is a hindrance. The major origin of this non-specific binding is the electrical charge of the surface of biotin-binding proteins. Tamavidin 2, a fungal avidin-like protein that binds biotin with an extremely high affinity, can be produced as a soluble recombinant protein in Escherichia coli. The isoelectric point of tamavidin 2 is 7.4-7.5, lower than avidin (10.0), and slightly higher than that of streptavidin (6.0-7.5). Here, we genetically engineered charge mutants of tamavidin 2 to reduce non-specific binding. By substituting an acidic residue (glutamic acid) for basic residues (arginine and lysine), we constructed three mutant proteins (muteins) and confirmed their high-level production in soluble form in E. coli, as well as that of tamavidin 2.?We then tested these proteins for non-specific binding to salmon sperm DNA, glycoproteins (integrin and fibronectin), and IgG from human sera. The muteins showed lower non-specific binding than tamavidin 2 to these macromolecules. In particular, one mutein, tamavidin-R104EK141E, which had the lowest isoelectric point (5.8-6.2) among avidin, streptavidin and tamavidin 2, displayed the lowest non-specific binding. The affinity of this mutein to biotin was high, comparable with that of tamavidin 2. These findings indicate that tamavidin-R104EK141E has the potential to serve as a robust tool in the numerous applications of biotin-binding proteins.     

Metal release from stainless steel, Co–Cr–Mo–Ni–Fe and Ni–Ti alloys in vascular implants

A seven-day immersion test using several solutions was conducted on stainless steel, Co-based alloy, and Ni–Ti alloy, which are used for stents and stent grafts. The quantitative data on the release of each metal ion and the correlation between metal ion release rate and pH were obtained. The quantities of Fe and Ni released from stainless steel gradually decreased with increasing solution pH (pH 2–7.5). For Co–Cr–Mo–Ni–Fe alloy, the quantity of Cr released steadily increased as pH decreased (pH ≤ 6) and reached nearly zero at pHs higher than 6 (pH 6–7.5). Co release was slightly affected by a variation in pH. The quantities of Ni and Ti released from Ni–Ti alloy markedly increased with decreasing pH (pH ≤ 4) and they leveled off from pH 4 (pH 4–7.5). Although the rapid increases were observed at approximately pH 2, the quantities were even higher than that of Co released from the Co–Cr–Mo and Co–Cr–Mo–Ni–Fe alloys. For further investigation of the rapid increase in the quantities of metals released at pH 2, an anodic polarization test was employed to study the passive and transpassive behaviours of Ni–Ti alloy. The critical current density for the passivation of Ni–Ti alloy markedly increased as pH decreased (pH ≤ 4) and was low (1.4 μA/cm²) at pH higher than 4 (pH 4–7.5). The potential at a current density of 10 μA/cm², by contrast, markedly rose with decreasing pH (pH ≤ 2), and was 1.2 V from pH 2 (pH 2–7.5).     

Application of β-sheet proteins as a compatibilizer in injection molding of kenaf core/polypropylene composites

ABSTRACT

Injection-molding was performed using ethylene–propylene copolymer as the matrix by varying the physicochemical treatment of the reinforcement, i.e., core portion of kenaf stem that was dry-distilled and coated with or without β-sheet protein derivatives. Although dry distillation reduced the moisture content, the Charpy impact strength of the molded composites was low. When molding was performed using the fillers coated with β-sheet protein derivatives, especially coated with liquid silk (LS), a satisfactory Charpy impact strength of the molded composites was markedly gained. Thus, the β-sheet protein derivatives had potential to work as a compatibilizer. Coating the fillers with β-sheet proteins was also effective in suppressing the generation of silver streaks on the surface of composites and in preventing the occurrence of defective injection molding.     

Reaction products of α-tocopherol with a free radical initiator, 2,2′-azobis(2,4-dimethylvaleronitrile)

α-Tocopherol wss reacted with an alkylperoxyl radical at 37°C in ethanol. 2,2′-Azobis(2,4-dimethylvaleronitrile) was used to generate the alkylperoxyl radicals. The reaction products of α-tocopherol were isolated by reverse-phase and normal-phase high performance liquid chromatography, and their structures were characterized by infrared, ultraviolet,¹H and¹³C nuclear magnetic resonances and mass spectrometry. They were 8aS-hydroperoxy-α-tocopherone, 8aR-hydroperoxy-α-tocopherone, a mixure of 7,8-epoxy-8aS-ethoxy-α-tocopherone, and 7,8-epoxy-8aR-ethoxy-α-tocopherone, 8aS-(1R-cyano-1,3-dimethyl)butylperoxy-α-tocopherone, 8aS-(1S-cyano-1,3-dimethyl)butylperoxy-α-tocopherone, a mixture of 8aR-(1S-cyano-1,3-dimethyl)butylperoxy-α-tocopherone and 8aR-(1S-cyano-1,3-dimethyl)butylperoxy-α-tocopherone, 4aR,5S-epoxy-8aS-ethoxy-α-tocopherone, 4aS,5R-epoxy-8aS-ethoxy-α-tocopherone and 4aR,5S-epoxy-8aR-ethoxy-α-tocopherone.     

Throughput analysis method for hybrid ARQ schemes over burst errorchannels

A definition of a burst error channel using a Markov model was presented by T. Sato et al. in a previous paper (1991). A throughput analysis method of hybrid automatic repeat request (ARQ) under the burst error channel using the three-state Markov model is described. The hybrid ARQ is studied for the random and burst error correction codes as the forward error correction (FEC) code, and multiframe rejection (MREJ) as the ARQ. The throughput efficiency is obtained with both an infinite buffer memory and a finite buffer memory. The applicable range of the burst error channel is clarified for the hybrid ARQ using random and burst error correction codes     

Chinese hamster ovary cells expressing a novel type of acetylated low density lipoprotein receptor. Isolation and characterization

Macrophage scavenger receptors mediate the recognition of a wide range of negatively charged macromolecules including acetylated low density lipoproteins (AcLDL). Chinese hamster ovary (CHO) cells were cultured in the presence of increasing concentrations of simvastatin, a cholesterol biosynthesis inhibitor, and AcLDL as the sole source of exogenous lipoproteins. The cells surviving under these conditions specifically bound 125I-labeled AcLDL with high affinity and degraded them via an endocytic pathway. Unexpectedly, the association and degradation of 125I-labeled AcLDL by these CHO cells were not inhibited by dextran sulfate, fucoidan, and polyinosinic acid, competitors of macrophage scavenger receptors, but were completely inhibited by maleylated bovine serum albumin. Furthermore, these cells effectively took up negatively charged liposomes containing acidic phospholipids such as phosphatidylserine and phosphatidic acid, whereas CHO cells expressing macrophage scavenger receptors did not. AcLDL and negatively charged liposomes were cross-competed with each other. Northern blot analysis using the cDNA for the macrophage scavenger receptor revealed that these CHO cells did not express this receptor. From these observations, we conclude that the isolated CHO cells express a novel type of AcLDL receptor, which is distinct from macrophage scavenger receptors with respect to ligand specificity and competitor sensitivity.