Copper (Cu) is one of the vital heavy metal elements that plays a major role in many physiological aspects of biological organisms. It is known to serve as an important factor in several enzymes in the human body such as superoxide dismutase, cytochrome c oxidase and tyrosinase (Tyr). Inside the cells Cu is regulated by the CuATPases, either for Cu transportation to the Golgi lumen or for excess Cu extracellular discharge. Genetic disorders in these ATPases can cause defects in the system, specifically in brain and liver, where they have been found to be abundant. Specifically, the Cu dyshomeostasis in the brain is related to several major neurodegenerative diseases which have become major concerns of the society such as Alzheimer’s disease and Parkinson’s disease. Thus, Cu-containing enzyme detection is important and worth considerable effort. In this regard, fluorescent detection methods based on various nanoparticles (NPs) are becoming popular. It is of utmost importance that these NPs are capable of specifically binding to cuproenzymes, without binding to other enzymes. Herein, a fluorescent detection technique of Cu-containing Tyr was developed using carbon nitride dots (CNDs). CNDs were synthesized using abundant materials, citric acid and urea and was found to have excellent photoluminescence (PL) properties with high quantum yields and biocompatibility. Tyr fluorescence inhibitory studies were conducted in Tris-HCl (pH=7.4) buffer by mixing with CNDs. A clear fluorescence inhibition of CNDs at its maximum PL emission, in the presence of Tyr was observed which follows a linear relation in relevance with the Tyr concentration. Further, the PL inhibition was not observed when non-metal enzymes such as bovine serum albumin (BSA), Glyceraldehyde 3-phosphate dehydrogenase (GADPH), Apo-transferrin (Apo-Tf) were used confirming the specific binding, thus resulting in better detection. Through circular dichroism (CD) spectrometry, it was confirmed that the CNDs does not change the tertiary structure of the enzymes. The proposed mechanism of this inhibition is that by CNDs interaction of the Cu ions through the nitrogen available on the surface of the CNDs, which is abundant with -NH<sub>2</sub> and -CONH<sub>2</sub> functionalities, thus resulting in a decrease of PL emission of CNDs. Therefore, CNDs can be a promising tool for Cu-enzyme detection in biological bodies.

Carbon nitride dots based biosensor for Cu-containing enzyme detection

Macromolecular Chemistry: The Second Century

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ACS Spring 2020 National Meeting & Expo

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