Citation: Li, Y.; Luo, S.; Wang, X.; He,
Y.; Yu, H. CDs-Peroxyfluor
Conjugation for Ratiometric
Fluorescence Detection of Glucose
and Shortening Its Detection Time
from Reaction Dynamic Perspective.
Biosensors 2023, 13, 222. https://
doi.org/10.3390/bios13020222
Received: 12 January 2023
Revised: 24 January 2023
Accepted: 31 January 2023
Published: 3 February 2023
Copyright: © 2023 by the authors.
Licensee MDPI, Basel, Switzerland.
This article is an open access article
distributed under the terms and
conditions of the Creative Commons
Attribution (CC BY) license (https://
creativecommons.org/licenses/by/
4.0/).
Article
CDs-Peroxyfluor Conjugation for Ratiometric Fluorescence
Detection of Glucose and Shortening Its Detection Time from
Reaction Dynamic Perspective
Yangjie Li , Site Luo, Xin Wang, Yang He and Haihu Yu *
National Engineering Laboratory for Fiber Optic Sensing Technology, Wuhan University of Technology,
Wuhan 430070, China
* Correspondence: hhyu@whut.edu.cn
Abstract:
A ratiometric fluorescence probe based on the conjugation of peroxyfluor-NHS (PF) and
carbon dots (CDs) was designed for selective and rapid detection of glucose. When glucose was
catalytically oxidized by glucose oxidase (GOx), the product H
2
O
2
would react with colorless and
non-fluorescent peroxyfluor moiety to give the colored and fluorescent fluorescein moiety which
would absorb the energy of CDs emission at 450 nm due to the Förster Resonance Energy Transfer
(FRET) and generate a new emission peak at 517 nm. The reaction between PF and H
2
O
2
was
slow with a rate constant of about 2.7
×
10
−4
s
−1
under pseudo-first-order conditions (1 uM PF,
1 mM H
2
O
2
), which was unconducive to rapid detection. Given this, a short time detection method
was proposed by studying the kinetics of the reaction between PF and H
2
O
2
. In this method, the
detection time was fixed at three minutes. The linear detection of glucose could be well realized
even if the reaction was partially done. As glucose concentration increased from 0.05 mM to 5 mM,
the fluorescence intensity ratio (I
517
/I
450
) after 3 minutes’ reaction of CDs-PF and glucose oxidation
products changed linearly from 0.269 to 1.127 with the limit of detection (LOD) of 17.19
µ
M. In
addition, the applicability of the probe in blood glucose detection was verified.
Keywords: carbon dots; ratiometric fluorescence; glucose; rapid detection; fluorescein; kinetics
1. Introduction
Diabetes is a serious chronic condition afflicting hundreds of millions of people. It is
among the top 10 causes of death in adults, and 4.2 million people among 20–79-year-old
adults were estimated to die from diabetes and its complications in 2019 [
1
,
2
]. Detecting
and controlling the blood glucose of diabetic patients will limit the long-term damage
of diabetes to the heart, eyes, kidneys, nerves and other organs and reduce risk of pre-
mature death [
3
,
4
]. Therefore, the development of an efficient and accurate method for
blood glucose monitoring is important for the treatment of diabetic patients and the de-
tection of prediabetic individuals. A variety of methods for glucose detection have been
reported in the past few years, such as fiber-optic surface plasmon resonance [
5
–
8
], electro-
chemical [
9
–
13
], surface-enhanced Raman scattering [
14
], electrochemiluminescence [
15
],
fluorescence [
16
–
19
] and so on. Among them, the fluorescence method has received a lot of
attention because of its simplicity, efficiency and sensitivity. Moreover, if two appropriate
fluorophores are combined a ratiometric fluorescence probe possessing self-calibration will
be established to eliminate the instrument errors and external interference to some extent,
thus improving the detection accuracy.
As novel fluorescent materials, carbon dots (CDs) have great application prospects in
biosensing due to their high quantum yield, biocompatibility, low toxicity, low cost and
stable chemical property [
20
–
24
]. CDs have been used for detections of analytes such as
metal ions [
25
] and small organic molecules [
26
–
28
]. However, most CDs-based probes
have single-emission detection, which brings many instrument errors to the detection and
Biosensors 2023, 13, 222. https://doi.org/10.3390/bios13020222 https://www.mdpi.com/journal/biosensors
