Self-assembled monolayers with biospecific affinity for lactate dehydrogenase for the electroenzymatic oxidation of lactate

Surface modified gold electrodes with high biospecific affinity for NAD(H)-dependent lactate dehydrogenase have been prepared by covalent attachment of several traizine dyes to stepwise functionalized mixed alkanethiol self-assembled monolayers. The biospecific affinity of such ligand-anchored monolayers to bind submonolayer amounts of enzyme was demonstrated from the course of the protein adsorption events monitored by surface plasmon resonance. Electroenzymatic activity measurements of lactate dehydrogenase modified surfaces for the reaction of lactate oxidation, carried out `ex situ¿ at different stages of protein layer growth, allowed the optimization of the preparative procedure to yield reproducible enzymatic electrodes with a low amount of unspecifically bound protein. A short adsorption time, as well as a high concentration of enzyme in the solution used for protein layer growth, led to lactate dehydrogenase-modified gold electrode surfaces with a high electroenzymatic activity arising mainly from biospecifically bound species. The lowest amount of unspecifically adsorbed protein was found for ligand-anchored monolayers prepared from mixed alkanethiol underlayers with an excess of positively charged groups. The lack of electroenzymatic activity shown by lactate dehydrogenase modified electrodes in the absence of soluble coenzyme (NAD+) indicates that none of the investigated ligand-anchored monolayers could provide an efficient electronic pathway from the metal to the active site of the enzyme. Therefore, the monolayers acted just as an anchoring system for lactate dehydrogenase

Polarimetric optical-fibre sensor for biochemical measurements

The use of an optical-fibre polarimeter as a chemical sensor is demonstrated. The compound to be detected is allowed to adsorb onto a decladded 5 cm length of the fibre. The fibre is polarization maintaining with an elliptical fibre core and a D-shaped geometry. The overall retardation stability of this fibre polarimeter is ≈ 0.5 × 2π rad m−1 K−1. With this sensor adsorption processes of proteins can be followed on-line. The resulting relative phase retardations caused by the growth of a monolayer of antibodies (αhCG, αhSA) are 0.25 × 2π. For the much smaller protein hSA, this value is 0.1 × 2

Self-assembled monolayers of cavitand receptors for the binding of neutral molecules in water

Resorcin[4]arene cavitands 2-4 were synthesized starting from tetrol 5. With these cavitand tetrasulfides, self-assembled monolayers (SAMs) on gold were prepared and characterized by electrochemical capacitance and resistance measurements, contact angle (CA) measurements, grazing angle FT-IR spectroscopy, and X-ray photoelectron spectroscopy (XPS). In situ surface plasmon resonance (SPR) spectroscopy was employed to study the host-guest interactions at the monolayer-water interface, between receptor adsorbates 1-4 and p-toluic acid, benzoic acid, p-nitrobenzoic acid, p-hydroxybenzoic acid, p-cresol, p-nitrophenol, and p-methoxyphenol. SPR results show that the presence of functional groups on the upper rim of the adsorbate molecules induces selectivity in the guest recognition process. Furthermore, it is interesting to note that such interactions are not directly related to the degree of hydrophobicity of the receptor adsorbate or of the guest. Concentration-dependent experiments were performed with p-nitrophenol as guest for a monolayer of adsorbate 1. At a 3 mM concentration of p-nitrophenol, approximately five to eight guest molecules associate with each adsorbate molecule, implying the formation of 1-4 layers of guests on the receptor surface