Bakthan Singaram

Monitoring blood glucose levels in diabetics is an important global problem. We introduced a two-component sensing system, consisting of a fluorescent dye and a boronic acid-containing molecule that acts dually as a fluorescence quencher and a saccharide receptor, for continuous glucose monitoring in blood. The object of this chapter is to trace major developments, largely in our own research program, which led from the initial observation of the exceptional sugar signaling capabilities of the ortho-borobenzyl viologen–hydroxypyrene trisulfonic acid (o-BBV–HPTS) system to the present time when it is possible to monitor glucose in the whole blood continuously. Through this research, we have identified several two-component sensing configurations and probes for the detection of saccharides in biological fluids. In this chapter, particular emphasis has been given to the development of the two-component boronic acid-based fluorescent glucose sensors. Mechanistically, in the absence of saccharide, a non-fluorescent ground-state complex forms through ionic association between the dye and the viologen boronic acid, resulting in a greatly reduced fluorescence of the dye. The electrostatic attraction between the dye and the quencher is lost, however, upon the binding of the viologen to the saccharide causing the dye–quencher complex to dissociate. This produces a fluorescence recovery that is directly dependent on the saccharide concentration. These boronic acid appended bipyridinium salts are tunable receptors and can be designed to increase glucose selectivity over that of other boronate-forming analytes. Fluorescent quantum dots can also be used as the reporter component in our system. Powerful solution-phase sensor arrays for neutral and anionic carbohydrates were created with these probes. They can be also used in new label-free fluorescent assays for carbohydrate-modifying enzymes. Immobilizing the two-component chemistry in a hydrogel allowed for continuous monitoring of saccharides. Hydrogels were incorporated in fiber optic devices for continuous monitoring of glucose. Most importantly, our two-component glucose sensing capabilities have contributed significantly to both the basic science of fluorescence-based sensor chemistry and to improving methods for detection of glucose and other carbohydrates in biological fluids.

Asymmetric 1,2-reduction of α,β-unsaturated ketones using TarB-NO₂ and NaBH₄ is reported. Simple cycloalkenones give products in low enantiomeric excess. However, cycloalkenones with α-substituents, such as halides, alkyl, and aryl, have been enantioselectively reduced with this system to yield chiral allylic alcohols in enantiomeric excess up to 99%. The starting materials for TarB-NO₂ are inexpensive, and the boronic acid can be easily recovered in high yield by a simple acid extraction.

The anionic fluorescent dye, aminopyrene trisulfonic acid (APTS), was synthesized and used in a solution-based two-component glucose-sensing system comprising the dye and a boronic acid-appended viologen. The fluorescence of the dye was quenched in the presence of the viologen and the fluorescence restored upon glucose addition. An important feature of this fluorophore is that it can be covalently bonded to a polymer through the amine group without a significant effect on optical properties. Two APTS derivatives, functionalized with polymerizable groups, were synthesized and immobilized in hydroxyethyl methacrylate (HEMA)-based hydrogels. The latter were used to continuously monitor glucose. The fluorescence signal modulation, signal stability, reversibility, reproducibility, and pH sensitivity of the hydrogels were evaluated. The APTS dyes described herein are insensitive to pH changes within the physiological range, both in solution and when immobilized in a hydrogel. When APTS is used in conjunction with boronic acid-appended viologens to sense glucose, the system displays some pH sensitivity because of the presence of the boronic acid.

The simultaneous use of several fluorescent reporter dyes in a multicomponent boronic acid-based glucose sensing system is reported. In one application, two dyes with widely different emission wavelengths are used to report changes in glucose concentration. A third glucose-insensitive dye was then added to act as a reference dye and provide for a ratiometric correction to the two reporter dye signals. The inclusion of such a reference dye reduces errors arising from sources such as fluctuations in lamp intensity and sample dilution. The simultaneous use of multiple fluorescent reporter dyes

High enantioselectivities are obtained for the reduction of a series of ketones using the inexpensive and mild reducing agent NaBH₄ with TarB‐NO₂ (1). This easily prepared tartaric acid‐based reagent combines a Lewis acid with carboxylic acids in a single bifunctional reagent. When combined with NaBH₄, the resulting chiral acyloxyborohydride mediates the reduction of aromatic ketones to provide the product alcohols in enantiomeric excesses of 93–98 %. Several aliphatic ketones were also reduced with moderate to excellent enantioselectivity. A unique mechanism is provided with supporting calculations for the proposed active species and transition state. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2005)