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Physiologically Stable Fluorophore For Digestion Toxicology Studies

Published Patent Application Numer: 2021/0179639 A1


The diversity of materials used industrially has been increasing, as well as the demand for more renewable feed stocks. As a result, new strategies to assay the impact on the environment and human physiology is needed. Typical methods involving radioactive isotopic labeling and fluorescent labeling to track uptake and analyze compounds after excretion. Isotopic labeling can be very expensive, and the common fluorescent dyes and attachment chemistries are sensitive to pH fluctuations and enzymatic attack. This fluorophore is highly resistant to pH fluctuations found in the digestive track and is resistant to enzymatic attack.  The dye will retain its fluorescent properties at every stage of digestion with enables the pathway and absorbance rate of materials to readily be tracked.

patent description

The invention is a fluorescent molecule with an inert fluorescent center and a chemical handle that can react readily with alcohols or amines forming an either type bond. Towards the molecular design, the fluorescent center of choice is 2,8-diethyl-5,5-difluo ro-1,3,7,9,1O­ pentamethy l-5H-4A4,SA4. - dipyrrolo[1,2 -c:2 ', l '1][ l ,3, 2]diazaborinine  (Bodipy). The Bodipy family of dyes is known for their photo stability and lack of toxicity. The moiety in the meso position of the fluorescent center is functionalized with a chlorotriazine. This functional group can react easily with nucleophiles such as alcohols or amines under mildly basic conditions to yield a stable aromatic ether linkage. Although not as stable as aliphatic eithers, this bond is highly resistant to pH and enzymatic attack, which sets it apart from the usual amide or thiourea bonds commonly formed with commercial fluorescent dyes. In addition, the 2 and 8 positions are protected with hydrophobic groups and renders these electrophilic centers inert. The photophysical characteristics of the dye 10-(4-((4,6-dichloro- l ,3,5-triazin-2-yl)oxy)phenyl)-2,8-diethyl-5,5- difluoro-l ,3,7,9-tetramethyl-SH-414,514- dipyrrolo[l ,2-c:2', l'-f][l ,3,2]diazaborinine (mDTEB) is such that commonly available instrumentation can be utilized.


The current methods for testing or tracking nano or micron sized particles for these studies either uses radioactive isotopes like l4C or fluorescent dyes with pH sensitivities or labile attachment chemistries. The isotopic method is quite expensive and requires specialized facilities. The common fluorophores used typically are quenched either in acidic conditions pH < 5, or basic conditions pH > 8. Both scenarios are found in the digestive tract. In addition, amides and thioureas can be attacked by digestive enzymes in addition to pH, producing free dye molecules in the study. mDTEB offers a chemical label that is both nontoxic and resistant to known digestive process and conditions. Synthesis of this compound is only two steps which puts in the realm of commercial viability, and less complex than several other fluorescent dyes.

Created September 7, 2022, Updated December 14, 2023