Screen-printed transcutaneous oxygen sensor employing polymer electrolytes.

A disposable, transcutaneous oxygen sensor has been designed and implemented using screen-printing technology for all fabrication stages. The sensor incorporates an integral heating element to promote transcutaneous diffusion of blood gases so that a reliable estimation of arterial blood gas concentration can be obtained. The oxygen sensing part of the device consists of a screen-printed Clark cell implemented as electrodes, electrolyte and membrane. A three-electrode configuration is employed with gold working and counter electrodes and a silver/silver chloride reference electrode. Several different polymer electrolyte and membrane materials were evaluated in the construction of the device, and their performances were compared. A fully automated gas testing rig was constructed to enable oxygen levels to be varied under computer control. Cyclic voltammetry and static analysis of the sensors were carried out at different oxygen concentration levels and in various test environments. Linear relationships were established with an averaged sensitivity level of 0.02 microA(mmHg)(-1) and high regression coefficients of 0.99. The prototype covered with a polytetrafluoroethylene membrane gave the experimental result of I (microA) = -0.025PO2 (mmHg) - 0.085. Several factors influenced the performance of the sensors. The investigations have greatly contributed towards an understanding of the suitability of the materials in achieving a viable, low-cost sensor.

Effects of a hydroxy-cinnamoyl conjugate of spermidine on arthropod neuromuscular junctions.

N1-coumaroyl spermidine is structurally similar to acylpolyamines found in spider and wasp venoms, which are known to block arthropod glutamate receptors. N1-coumaroyl spermidine reduced the amplitude of excitatory postsynaptic potentials recorded in crayfish muscle. This effect was dose dependent, with an IC50 value of 70 micromol l(-1). N1-coumaroyl spermidine reversibly reduced the amplitude of potentials elicited by iontophoretic application of L-glutamate, indicating a direct effect on postsynaptic glutamate receptors. Neither 1 mmol l(-1) spermidine nor 1 mmol l(-1) coumaric acid altered excitatory postsynaptic potential amplitude, indicating that blockage requires the conjugated phenolic polyamine. N1-coumaroyl spermine, a slightly longer phenolic polyamine, reduced excitatory postsynaptic potential amplitude with approximately the same potency as N1-coumaroyl spermidine. Thus, potency of blockage does not appear to be affected in this experimental preparation by small changes in length of the polyamine. N1-coumaroyl spermidine also reduced excitatory postsynaptic potentials in muscles of the insect Drosophila. The ability of N1-coumaroyl spermidine to attenuate synaptic transmission at insect neuromuscular synapses lends support to the notion that plant-derived phenolic polyamines might serve as natural insecticides.

Synthesis of photoaffinity label analogues of alpha-tocopherol.

Photoaffinity analogues of alpha-tocopherol have been synthesized that incorporate the photosensitive 4-azido-2,3,5,6-tetrafluorobenzyloxy group at the terminus of unbranched analogues of the naturally occurring phytyl side chain. An intermediate from these syntheses has also been used to generate a supported ligand for bioaffinity chromatography of alpha-tocopherol binding proteins.

Cytochrome P450-catalyzed hydroxylation of hydrocarbons: kinetic deuterium isotope effects for the hydroxylation of an ultrafast radical clock.

The ultrafast radical clock probe trans-1-methyl-2-phenylcyclopropane (1CH3) and its mono-, di-, and trideuteriomethyl analogues were oxidized by phenobarbital-induced rat liver microsomal enzymes. This cytochrome P450-catalyzed hydroxylation of 1CH3 gave three products: the alcohol trans-(2-phenylcyclopropyl)methanol (2), the rearranged alcohol 1-phenylbut-3-en-1-ol (3), and the phenol trans-2-(p-hydroxyphenyl)-1-methylcyclopropane (4). The identification of both the unrearranged and rearranged products of oxidation, 2 and 3, is consistent with the formation of a radical intermediate via a hydrogen atom abstraction from the methyl group by the catalytically active iron-oxo center. Hydroxylation of three deuteriomethyl forms of 1CH3 produced the analogous deuterated products, although in different amounts of each. Perdeuteration of the methyl group (1CD3) disfavored oxidation at the methyl group and caused an increase in the oxidation of the phenyl ring (metabolic switching). By comparing the amounts of alcohols and phenol formed from the individual, noncompetitive oxidation of 1CH3 and 1CD3 the overall (i.e., combined primary and secondary) deuterium kinetic isotope effect (DKIE) was found to be 12.5. Intramolecular DKIEs for 1CHD2 and 1CH2D were 2.9 and 13.2, respectively. From these results, the primary and secondary DKIEs were calculated to be 7.87 and 1.26, respectively, values that indicate that there is extensive C--H bond stretching in the transition state for the rate-controlling step in P450-catalyzed hydroxylation of 1CH3.(ABSTRACT TRUNCATED AT 250 WORDS)

A nuclear juvenile hormone-binding protein from larvae of Manduca sexta: a putative receptor for the metamorphic action of juvenile hormone.

A 29-kDa nuclear juvenile hormone (JH)-binding protein from the epidermis of Manduca sexta larvae was purified by using the photoaffinity analog for JH II ([3H]epoxyhomofarnesyldiazoacetate) and partially sequenced. A 1.1-kb cDNA was isolated by using degenerate oligonucleotide primers for PCR based on these sequences. The cDNA encoded a 262-amino acid protein that showed no similarity with other known proteins, except for short stretches of the interphotoreceptor retinoid-binding protein, rhodopsin, and human nuclear protein p68. Recombinant baculovirus containing this cDNA made a 29-kDa protein that was covalently modified by [3H]epoxyhomofarnesyldiazoacetate and specifically bound the natural enantiomer of JH I (Kd = 10.7 nM). This binding was inhibited by the natural JHs but not by methoprene. Immunocytochemical analysis showed localization of this 29-kDa protein to epidermal nuclei. Both mRNA and protein are present during the intermolt periods; during the larval molt, the mRNA disappears but the protein persists. Later when cells become pupally committed, both the mRNA and protein disappear with a transient reappearance near pupal ecdysis. The properties of this protein are consistent with its being the receptor necessary for the antimetamorphic effects of JH.

Cytochrome P450 hydroxylation of hydrocarbons: variation in the rate of oxygen rebound using cyclopropyl radical clocks including two new ultrafast probes.

The oxidation of eight methyl-substituted and three alkyl-substituted cyclopropanes by rat liver microsomal cytochrome P450 and pure reconstituted rabbit P450 2B4 was studied. Alkane hydroxylation catalyzed by P450 is generally believed to proceed by hydrogen abstraction followed by reaction of the carbon-centered radical with an iron-bound hydroxyl radical, a process called oxygen rebound. Hydrogen abstraction from methylcyclopropanes generates cyclopropylcarbinyl radicals whose solution rate constants for ring opening are known [Bowry, V.W., et al. (1991) J. Am. Chem. Soc. 113, 5687-5698]. Rearranged products were only observed with the five substrates which, upon hydrogen abstraction, would generate a cyclopropylcarbinyl radical that undergoes ring opening with a rate constant > or = 2.0 x 10(9) s-1 in solution. Values of the rate constants for oxygen rebound (kOH) were calculated by determining the ratio of unrearranged products (cyclopropylmethanols) to rearranged products (alkenols). For each substrate this ratio was generally about the same for the oxidations catalyzed by microsomal P450 and by P450 2B4. It is concluded that all of the substrates are oxidized via an intermediate cyclopropylcarbinyl radical. Two ultrafast probes, trans-1-methyl-2-phenylcyclopropane and 1,1-diphenyl-2-methylcyclopropane, gave alcohol product ratios which yielded unreasonably high values for kOH, viz., ca. 1.5 x 10(12) and ca. 7 x 10(12) s-1, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)