Chemistry: analytical and immunological testing – Heterocyclic carbon compound – Hetero-o
Reexamination Certificate
2000-12-05
2003-11-25
Snay, Jeffery (Department: 1743)
Chemistry: analytical and immunological testing
Heterocyclic carbon compound
Hetero-o
C436S172000, C422S082080
Reexamination Certificate
active
06653141
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
This invention relates to an improved optical method and sensor for polyhydroxy-substituted organic molecules that measure the concentration of these molecules in aqueous or organic media. In particular, the method and sensor monitor the concentration of sugars, i.e. glucose or fructose, in aqueous solution in vitro. The determination of glucose in samples of body fluid in vitro is of particular importance. Some of the novel components of the optical method and device are also considered to be inventions in their own right.
2. Description of Related Art
There has been an ongoing effort over many years to use fluorescence techniques to measure polyhydroxyl compound (e.g. glucose) concentrations in body fluids. Although the term “glucose” is used herein below, it is to be understood that the concentration of most polyhydroxyl-containing organic compounds (carbohydrates, 1,2-diols, 1,3-diols and the like) in a solution are capable of being determined. But in spite of the intense effort, no practical system has been developed and commercialized. Several attempts have been made to detect glucose by fluorescence using dyes to which a boronic acid group has been attached. Boronic acids are known to bind sugars reversibly. When the boronic acid functional dye binds to a sugar, the properties of the dye are affected. These changes have been used in the past to measure sugar concentration.
One use of this approach to a glucose sensor was reported by Russell, U.S. Pat. No. 5,137,833 (See also Russell & Zepp, U.S. Pat. No. 5,512,246) which disclosed the use of a boronic acid functionalized dye that binds to glucose and generates a signal dependent on glucose concentration. James et al, U.S. Pat. No. 5,503,770,used the same principle but combined a fluorescent dye, an amine quenching functionality, and a boronic acid in a single complex moiety, the fluorescence emission from which varies with extent of glucose binding. Van Antwerp et al, U.S. Pat. Nos. 6,002,954 and 6,011,984 combined features of the previously cited references and also taught fabrication of a device that is purported to be implantable.
Patents of interest include but are not limited to:
Russell, U.S. Pat. No. 5,137,833 (1992)
James et al, U.S. Pat. No. 5,503,770 (1996)
Russell & Zepp, U.S. Pat. No. 5,512,246 (1996)
Van Antwerp et al, U.S. Pat. No. 6,002,954 (1999)
Van Antwerp and Mastrototaro, U.S. Pat. No. 6,011,984 (2000)
Related U.S. patents of interest include:
Wolfbeis et al, U.S. Pat. No. 4,586,518 (1986)
Gallop & Paz, U.S. Pat. No. 4,659,817 (1989)
Yafuso & Hui, U.S. Pat. No. 4,798,738 (1989)
Yafaso & Hui, U.S. Pat. No. 4,886,338 (1989)
Saaski et al, U.S. Pat. No. 5,039,491 (1991)
Lanier et al, U.S. Pat. No. 5,114,676 (1992)
Wolfbeis et al, U.S. Pat. No. 5,232,858 (1993)
Colvin, U.S. Pat. No. 5,517,313 (1996)
Sundrehagen et al, U.S. Pat. No. 5,631,364 (1997)
James et al, U.S. Pat. No. 5,763,238 (1998)
Siegmund et al, U.S. Pat. No. 5,711,915 (1998)
Barnard & Rouilly, U.S. Pat. No. 5,852,126 (1998)
Colvin, U.S. Pat. No. 5,894,351 (1999)
Alder et al, U.S. Pat. No. 5,922,612 (1999)
Arnold et al, U.S. Pat. No. 6,063,637 (2000)
Song et al, U.S. Pat. No. 6,046,312 (2000)
Kimball et al, U.S. Pat. No. 6,139,799 (2000)
Chick et al, U.S. Pat. No. 6,040,194 (2000)
Related articles and publications of interest include:
Yoon & Czarnik,
J. Amer. Chem. Soc
. (1992) 114, 5874-5875
James, Linnane, & Shinkai,
Chem. Commun
. (1996), 281-288
Suenaga et al,
Tetrahedron Letters
(1995), 36, 4825-4828
Eggert et al,
J.Org.Chem
. (1999), 64, 3846-3852
Wolfbeis et al,
Analytica Chimica Acta
(1995), 304, 165-170
Wang et al,
Organic Letters
(1999), 1, 1209-1212
Chen et al,
Proc. Nat. Acad. Sci
. (1999), 96, 12287-12292
P. D. Hale et al,
Analytica Chimica Acta
(1999), 248, 155-161
A. E. Colvin, Jr. et al,
Johns Hopkins Technical Digest
, Vol. 12, #17, p.378 (1996)
Murakami et al,
Chem. Letters
(
Japan
) (2000), (8), p. 940-941.
References of a general nature include:
A. W. Czarnik (ed),
Fluorescent Chemosensors for Ion and Molecule Recognition
ACS Washington, D.C. 1992.
F. W. Scheller et al (eds),
Frontiers in Biosensorics I Fundamental Aspects,
Birkhaüser Vertag, Basel 1997.
J. R. Lakowicz,
Principles of Fluorescence Spectroscopy,
2
nd
ed. Kluwer Academics/Plenum Publishers, New York, N.Y. (1999).
Haugland, R. P.
Handbook of Fluorescent Probes and Research Chemicals
6
th
ed. Molecular Probes Inc., Eugene, Oreg. (1996).
All patents, articles, references, standards and the like cited in this application are incorporated herein by reference in their entirety.
All of these prior art sensors are deficient in one or more aspects, such as operability under physiological conditions, stability of operation, simplicity of design, reliability, and sensitivity. The present invention overcomes these deficiencies.
SUMMARY OF THE INVENTION
This present invention concerns an optical method and an optical device for determining in vitro the concentrations of polyhydroxyl compounds, especially sugars such as glucose or fructose, in aqueous and/or organic media. These compounds, the analytes, are in a system with a fluorescence sensing device comprised of a light source, a detector, a fluorophore (fluorescent dye), a quencher and an optional polymer matrix. Some components may be inventions in their own right. When excited by light of appropriate wavelength, the fluorophore emits light (fluoresces). The intensity of the light is dependent or, the extent of quenching. The fluorophore and quencher are preferably independent entities, optionally they are immobilized in or covalently attached to a polymeric matrix which is permeable to or in contact with the compounds of interest to be detected and quantified.
In one aspect, the present invention comprises a class of fluorescence quenching compounds that are responsive to the presence of polyhydroxyl compounds such as glucose in aqueous or organic media optionally at or near physiological pH. In other words, the quenching efficiency is controlled by the concentration of these compounds in the medium. The quencher is comprised of a viologen substituted with at least one boronic acid group as a discrete molecule, or wherein the adduct is optionally immobilized or covalently bonded to a polymer. The quencher, dye and an optional polymer may also be covalently bonded to each other.
The combination of boronic acid and viologen, and the resultant effect on viologen properties are important embodiments of the present invention.
In another aspect, the present invention is a class of polymeric fluorescent dyes which are susceptible to quenching by the viologen/boronic acid adduct. Useful dyes include pyranine derivatives (e.g. hydroxypyrene trisulfonamide derivatives and the like (See FIGS.
1
A and
1
B).
In one embodiment, the dye is comprised of a hydroxypyrene trisulfonamide moiety bonded to a polymer. Converting sulfonic acid groups to sulfonamide groups shifts the pKa of pyranine into a range more suitable for measurement at physiological pH. These derivatives are typically prepared by reacting a trisulfonyl chloride intermediate with 1) a polyamine, 2) an amine functional ethylenically unsaturated monomer which adduct is subsequently polymerized, 3) or an amine functional polymer. Preferably, the dye is a fully substituted trisulfonamide containing no residual sulfonic acid groups.
In another aspect, the present invention is a composite water or organic solvent-compatible polymer matrix, preferably a hydrogel, which comprises the dye and quencher moieties. The matrix is a water- or organic liquid-swellable copolymer, preferably crosslinked, to which the dye and quencher moieties are covalently bonded. More preferably, the matrix is an interpenetrating polymer network (IPN) with the dye incorporated in one polymer network and the quencher in the other polymer. Most preferably, the matrix is a semi-IPN wherein the dye component is a high molecular weight water- or organic-soluble or dispersible polymer trapped in a crosslinked network comp
Singaram Bakthan
Wessling Ritchie A.
Myers Howard M.
Peters, Verny, Jones & Schmitt, LLP
Snay Jeffery
The Regents of the University of California
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