BODIPY-based Fluorescent Probe Research Articles

Colorimetric and fluorescent probes for cysteine detection: Applications in food safety and cellular imaging

In this study, three BODIPY-based fluorescent probes were designed and synthesized. The ultraviolet-visible spectra, fluorescence spectra, smartphone color recognition application and bioimaging were utilized to evaluate the capacity of the probes. By comparing key parameters, BDP-SIN had optimal performances including fastest response (10 min), highest signal-to-noise ratio (815 times) and lowest limit of detection (LOD = 49 nM). The recovery rate ranged from 92.04 % to 103.25 %. Meanwhile, BDP-SIN was triumphantly employed for determination of Cys in different daily food samples. Moreover, the test strips and microporous filter membrane loaded with BDP-SIN were developed for the portable real-time visualization and quantitative detection of Cys in food samples, which the contents ranged from 0.27 μM to 0.49 μM. Besides, BDP-SIN could image Cys in the living cells and mice. The novelty of this work was that developed an effective tool for researching the roles of Cys in food industry and living organisms.

Lysosome-Targeted Polarity-Sensitive NIR Fluorescence Probe for Imaging Injured Lung and Liver in Diabetes.

Diabetes is a chronic disease marked by high blood glucose. With the progress of diabetes, complications gradually appear, and various organs may be affected. However, due to the lack of noninvasive in situ detection probes, the diagnosis of organ damage caused by diabetes is significantly delayed, which will cause many complications that cannot be treated in time. Here, we report a BODIPY-based fluorescent probe SNL, which can be used to detect lung and liver damage caused by diabetes. By introducing methylpiperazine and extending the conjugated system, SNL can locate lysosomes and exhibit absorption and emission both in the near-infrared (NIR) region. In addition, SNL is sensitive to polarity and can be used for sensitive detection of lysosomal polarity changes. Unexpectedly, SNL targets and images the lungs and liver of mice. Subsequently, hyperglycemia-stimulated cell models and diabetic mouse models were successfully established, and SNL was utilized to reveal that polarity can be used as a diagnostic signal of diabetic complications. Notably, SNL for the first time confirmed the lung injury and liver injury caused by diabetes using the fluorescent probes method, providing a new approach for the diagnosis of diabetes complications.

A novel lysosome-targeting BODIPY-based fluorescent probe with two near-infrared channel signals for ratiometric detection of HClO and its application in diabetes mice model

Diabetes is an endocrine and metabolic disease characterized by hyperglycemia, which can lead to a variety of serious complications. Hypochlorous acid (HClO), one of the significant members of reactive oxygen species (ROS), is essential for various physiological processes, and the acidic lysosomes of phagocytes are one of its main sources. The related studies suggest that abnormal levels of HClO in the body have a strong association with diabetic process, but the pathological mechanisms of HClO on diabetic diseases are still unclear. Here, a novel ratiometric lysosome-targeted BODIPY-based fluorescence probe Lyso-HClO was designed for specific recognition of HClO. Lyso-HClO could rapidly sense HClO within 20 s, and its limit of detection was calculated as low as 45 nM. Both the emission wavelengths of Lyso-HClO before and after reaction of HClO reached the near-infrared region, which effectively reduced tissue damage and eliminated background interference from organisms to achieve more accurate analysis. Lyso-HClO was successfully employed for fluorescence imaging of both exogenous and endogenous HClO in RAW264.7 cells, HepG2 cells, and zebrafish. Additionally, it was effectively utilized to monitor the HClO fluctuations in diabetic mice, exhibiting a 15-fold increase in the fluorescence signal ratio (I650/I685). Therefore, the probe Lyso-HClO has enormous potential for the early diagnosis and prevention of diabetes and its complications.

Rational Design of a Highly Sensitive Carboxylesterase Probe and Its Application in High-Throughput Screening for Uncovering Carboxylesterase Inhibitors.

Tracking carboxylesterases (CESs) through noninvasive and dynamic imaging is of great significance for diagnosing and treating CES-related metabolic diseases. Herein, three BODIPY-based fluorescent probes with a pyridine unit quaternarized via an acetoxybenzyl group were designed and synthesized to detect CESs based on the photoinduced electron transfer process. Notably, among these probes, BDPN2-CES exhibited a remarkable 182-fold fluorescence enhancement for CESs within 10 min. Moreover, BDPN2-CES successfully enabled real-time imaging of endogenous CES variations in living cells. Using BDPN2-CES, a visual high-throughput screening method for CES inhibitors was established, culminating in the discovery of an efficient inhibitor, WZU-13, sourced from a chemical library. These findings suggest that BDPN2-CES could provide a new avenue for diagnosing CES-related diseases, and WZU-13 emerges as a promising therapeutic candidate for CES-overexpression pathological processes.

A lysosome-targeted fluorescent probe for fluorescence imaging of hypochlorous acid in living cells and in vivo

Lysosomes, as crucial acidic organelles in cells, play a significant role in cellular functions. The levels and distribution of hypochlorous acid (HOCl) within lysosomes can profoundly impact their biological functionality. Hence, real-time monitoring of the concentration of HOCl in lysosomes holds paramount importance for further understanding various physiological and pathological processes associated with lysosomes. In this study, we developed a bodipy-based fluorescent probe derived from pyridine and phenyl selenide for the specific detection of HOCl in aqueous solutions. Leveraging the probe's sensitive photoinduced electron transfer effect from phenyl selenide to the fluorophore, the probe exhibited satisfactory high sensitivity (with a limit of detection of 5.2 nM and a response time of 15 s) to hypochlorous acid. Further biological experiments confirmed that the introduction of the pyridine moiety enabled the probe molecule to selectively target lysosomes. Moreover, the probe successfully facilitated real-time monitoring of HOCl in cell models stimulated by N-acetylcysteine (NAC) and lipopolysaccharide (LPS), as well as in a normal zebrafish model. This provides a universal method for dynamically sensing HOCl in lysosomes.

Dual-Responsive Near-Infrared BODIPY-Based Fluorescent Probe for the Detection of F- and HClO in Organisms.

Fluoride anions (F-) play a crucial role in human physiological processes. However, excessive intake of F- would affect oxygen metabolism and promote the generation of oxygen-free radicals. Hence, it is essential to develop a precise and efficient fluorescent probe for visualizing F--induced oxidative stress. In this work, we developed the first bifunctional BODIPY-based fluorescent probe dfBDP with p-tert-butyldimethylsilanolate benzyl thioether as the sensing site for the detection of F- and HClO via two distinct reactions, the self-immolative removal and the thioether oxidation, which generate the sensing products with two nonoverlap fluorescence bands: 800-1200 and 500-750 nm, respectively. The probe dfBDP displays rapid response, high specificity, and sensitivity for the detection of F- (LOD, 316.2 nM) and HClO (LOD, 33.9 nM) in vitro. Cellular imaging reveals a correlation between F--induced oxidative stress and the upregulation of HClO. Finally, probe dfBDP was employed to detect F- and HClO in mice under the stimulation of F-. The experimental results display that the level of HClO elevates in the liver of mice.

A Benzil- and BODIPY-Based Turn-On Fluorescent Probe for Detection of Hydrogen Peroxide.

Faced with rising threats of terrorism, environmental and health risks, achieving sensitive and selective detection of peroxide-based explosives (PEs) has become a global focus. In this study, a turn-on fluorescent probe (BOD) based on benzil (H2O2-recognition element) and 4,4-difluoro-4-bora-3a,4a-diaza-s-indacene (BODIPY) derivative (fluorophore) was developed to sensitively and specifically detect hydrogen peroxide (H2O2). The synthesized BOD had a very weak fluorescence due to intramolecular donor-excited photo-induced electron transfer (d-PET) effect; however, it could emit a strong fluorescence since H2O2 selectively oxidized the benzil moiety and released free BODIPY fluorophore (BOD-COOH). As a result, the proposed BOD detected H2O2 in linear detection ranged from 25 to 125 µM with a detection limit of 4.41 µM. Meanwhile, the proposed BOD showed good selectivity toward H2O2, which is not affected by other common reactive oxygen species (ROS) and ions from explosive residues. In addition, a blue shift from 508 to 498 nm was observed in the absorption spectra upon addition of H2O2. More importantly, the BOD was successfully applied for rapid detection of H2O2 vapor with good sensitivity (down to 7 ppb), which holds great potential for practical use in public safety, forensic analysis and environmental monitoring.

A highly selective and sensitive fluorescence probe based on BODIPY-cyclen for hydrogen sulfide detection in living cells and serum

Hydrogen sulfide (H2S) is a multifunctional gaseous signaling molecule that plays a vital role in several biological processes. In the present study, a BODIPY-based fluorescent probe called 8-[4-((1,4,7,10-tetraazacyclododecane)methyl)phenyl]-4,4-difluoro-1,3,5,7-tetramethyl-4-bora-3a, 4a-diaza-s-indacene (BA–Cyclen)–Cu was designed and synthesized; this probe is a Cu(Ⅱ) complex that uses Cu(Ⅱ) decomplexation to achieve the sensitive and rapid detection of aqueous H2S via the “turn-on” mode. We observed that BA–Cyclen–Cu exhibited good membrane permeability, low toxicity, and lysosome-targeting ability, facilitating H2S detection in living cells. Furthermore, we demonstrated the potential biological applications of the probe by measuring exogenous H2S originating from Na2S and GYY4137, a slow-release donor, and endogenous H2S generated via the catalysis of cystathionine-β-synthase in both normal (H9c2) and cancerous (U87) cells. Moreover, BA–Cyclen–Cu was successfully used to detect exogenous H2S by the external standard method in fetal bovine serum, the serum of a healthy person, and the serum of a patient with liver cancer.

A simple BODIPY-based fluorescent probe for sequential recognition of Cu2+ and GSH and its application on test strips and bioimaging in living cells

A novel, highly sensitive “on-off-on” fluorescent probe (E)-N’-((1H-imidazol-2-yl) methylene) -4-(5, 5-difluoro-1, 3, 7, 9-tetramethyl-5H-4λ4,5λ4-dipyrrolo [1, 2-c:2′, 1′-f] [1,3,2] diazaborinin-10-yl) benzohydrazide (BDB) was developed in this study, and its structural and spectral properties were characterized by 13C NMR, 1H NMR, X-ray, ESI-MS, ultraviolet and fluorescence photometry. BDB could specifically and efficiently identify Cu2+ among many other metal ions with a fluorescence quenching efficiency of 92.7% and a recognition limit of 0.65 μM in CH3CN/HEPES (7:3, v/v, pH = 7.23) buffer. The energy difference between BDB and BDB-Cu2+ complex was calculated by DFT. The BDB-Cu2+ complex could continuously identify GSH in the same system and show an obvious fluorescence enhancement response with a recovery efficiency of approximately 91% and a low recognition limit of 0.02 μM, which is significantly lower than the physiological concentration of GSH in living cells. The applicability of BDB is verified by the strip test. BDB can also be used to detect GSH in HeLa cells, which expand the application of fluorescent chemical sensors in biology.

BODIPY-based fluorescent probe for cysteine detection and its applications in food analysis, test strips and biological imaging.

Cysteine, as one of semi-essential amino acids, which is absorbed from protein-rich foods and acts considerable role in various physiological processes. Here, we designed and synthesized a BODIPY-based turn-on fluorescent probe BDP-S for detecting Cys. The probe displayed short reaction time (10min), distinct color response (from blue to pink), large signal noise ratio (3150-fold), high selectivity and sensitivity (LOD=11.2nM) toward Cys. Moreover, BDP-S could not only be used for quantitative determination of Cys in food samples, but also be conveniently deposited on the test strips for qualitative detection of Cys. Notably, BDP-S was successfully used for imaging Cys in living cells and in vivo. Consequently, this work provided a hopefully powerful tool for detecting Cys in food samples and complex biological systems.

Development of meso-Five-Membered Heterocycle BODIPY-Based AIE Fluorescent Probes for Dual-Organelle Viscosity Imaging.

Fluorescent rotors with aggregation-induced emission (AIE) and organelle-targeting properties have attracted great attention for sensing subcellular viscosity changes, which could help understand the relationships of abnormal fluctuations with many associated diseases. Despite the numerous efforts spent, it remains rare and urgent to explore the dual-organelle targeting probes and their structural relationships with viscosity-responsive and AIE properties. Therefore, in this work, we reported four meso-five-membered heterocycle-substituted BODIPY-based fluorescent probes, explored their viscosity-responsive and AIE properties, and further investigated their subcellular localization and viscosity-sensing applications in living cells. Interestingly, the meso-thiazole probe 1 showed both good viscosity-responsive and AIE (in pure water) properties and could successfully target both mitochondria and lysosomes, further imaging cellular viscosity changes by treating lipopolysaccharide and nystatin, attributing to the free rotation and potential dual-organelle targeting ability of the meso-thiazole group. The meso-benzothiophene probe 3 with a saturated sulfur only showed good viscosity-responsive properties in living cells with the aggregation-caused quenching effect and no subcellular localization. The meso-imidazole probe 2 showed the AIE phenomenon without an obvious viscosity-responsive property with a C═N bond, while the meso-benzopyrrole probe 4 displayed fluorescence quenching in polar solvents. Therefore, for the first time, we investigated the structure-property relationships of four meso-five-membered heterocycle-substituted BODIPY-based fluorescent rotors with viscosity-responsive and AIE properties, and among these, 1 with a C═N bond and a saturated sulfur on the meso-thiazole, potentially contributing to their corresponding AIE and viscosity-responsive properties, served as a sensitive AIE fluorescent rotor for imaging dual-organelle viscosity in both mitochondria and lysosomes.

Monitoring of Hypochlorite Level in Fruits, Vegetables, and Dairy Products: A BODIPY-Based Fluorescent Probe for the Rapid and Highly Selective Detection of Hypochlorite.

Hypochlorite/hypochlorous acid (ClO-/HOCl), among the diverse reactive oxygen species, plays a vital role in various biological processes. Besides, ClO- is widely known as a sanitizer for fruits, vegetables, and fresh-cut produce, killing bacteria and pathogens. However, excessive level of ClO- can lead to the oxidation of biomolecules such as DNA, RNA, and proteins, threatening vital organs. Therefore, reliable and effective methods are of utmost importance to monitor trace amounts of ClO-. In this work, a novel BODIPY-based fluorescent probe bearing thiophene and a malononitrile moiety (BOD-CN) was designed and constructed to efficiently detect ClO-, which exhibited distinct features such as excellent selectivity, sensitivity (LOD = 83.3 nM), and rapid response (<30 s). Importantly, the probe successfully detected ClO- in various spiked water, milk, vegetable, and fruit samples. In all, BOD-CN offers a clearly promising approach to describe the quality of ClO--added dairy products, water, fresh vegetables, and fruits.

A lysosomal targeted fluorescent probe based on BODIPY for monitoring NO in living cells and zebrafish imaging

Nitric oxide (NO), as a special biological small molecule, plays an important role in physiology and pathology. Studies have revealed a strong correlation between human illness and the amount of NO in lysosomes. In this research, we synthesized a BODIPY-based fluorescent probe BML that can be localized in the lysosome to detect NO. The secondary amine in the BML structure can be N-nitrosation reaction with NO in the lysosome, preventing the initial photo-induced electron transfer (PET) process and leading to the creation of the highly luminescent BML-NO. The reaction mechanism was proved in the high-resolution mass spectrometry and DFT simulation calculations. Additionally, BML was discovered to have some evident benefits, such as fast detection time (3.5 min), low detection limit (LOD = 10.3 nM), and the resistance to interference and acidity, which are essential for the detection of NO in lysosomes. Furthermore, the BML was effectively employed to detect endogenous and exogenous nitric oxide in living cells and zebrafish.

A BODIPY-based fluorescent probe for simultaneous detection of H2O2 and viscosity during the pyroptosis process.

A dual functional BODIPY fluorescent probe was developed for simultaneous detection of H2O2 and viscosity, by collecting fluorescence from 800-1100 nm and 550-750 nm, respectively. Bioimaging based on the probe shows that H2O2 accumulates and cytoplasmic viscosity increases during the palmitic acid (PA)-induced pyroptosis process.

Bioimaging of Lysosomes with a BODIPY pH-Dependent Fluorescent Probe.

Fluorescence-based probes represent a powerful tool for noninvasive imaging of living systems in real time and with a high temporal and spatial resolution. Amongst several known fluorophores, 3-difluoroborodipyrromethene (BODIPY) derivatives have become a cornerstone for innovative fluorescent labelling applications, mainly due to their advantageous features including their facile synthesis, structural versatility and exceptional photophysical properties. In this context, we report a BODIPY-based fluorescent probe for imaging of lysosomes in living cells. The BODIPY derivative displayed a remarkable fluorescence enhancement at low pH values with a pKa* of 3.1. In vitro studies by confocal microscopy in HeLa cells demonstrated that the compound was able to permeate cell membrane and selectively label lysosome whilst remaining innocuous to the cell culture at the maximum concentration tested. Herein, the BODIPY derivative holds the promise of investigating lysosomal dynamics and function in living cells through fluorescence imaging.

BODIPY-Based Fluorescent Probes for Selective Visualization of Endogenous Hypochlorous Acid in Living Cells via Triazolopyridine Formation.

In this work, the two pyridylhydrazone-tethered BODIPY compounds (2 and 3) were synthesized. These compounds aimed to detect hypochlorous acid (HOCl) species via cyclic triazolopyridine formation. The open forms and the resulting cyclic forms of BODIPYs (2, 3, 4, and 5) were fully characterized by nuclear magnetic resonance, mass spectrometry, infrared spectroscopy, and single-crystal X-ray diffraction. These two probes can selectively detect HOCl through a fluorescence turn-on mechanism with the limit of detections of 0.21 µM and 0.77 µM for compounds 2 and 3, respectively. This fluorescence enhancement phenomenon could be the effect from C = N isomerization inhibition due to HOCl-triggered triazolopyridine formation. In cell imaging experiments, these compounds showed excellent biocompatibility toward RAW 264.7 murine live macrophage cells and greatly visualized endogenous HOCl in living cells stimulated with lipopolysaccharide.

Novel Meso-Benzothiazole-Substituted BODIPY-Based AIE Fluorescent Rotor for Imaging Lysosomal Viscosity and Monitoring Autophagy.

Meso-substituted boron dipyrromethenes (BODIPYs) provide a potential and innovative strategy for the synergistic construction of aggregation-induced emission (AIE) probes and fluorescent rotors for monitoring cellular viscosity changes, which play critical roles in understanding the function of viscosity in its closely associated diseases. Therefore, for the first time, a BODIPY-based fluorescent probe (1) with a rotatable meso-benzothiazole group was rationally designed and synthesized, showing both good viscosity-responsive and AIE properties. Probe 1 through direct linkage with the thiazole group, showed nearly no emission in low viscous solvents; however, a strong emission at 534 nm appeared and increased gradually with the increase in viscosity, attributing to the efficient restriction of the rotatable meso-benzothiazole group. The intensity (log I534) displayed a good linear relationship with viscosity (log η) in the viscous range of 0.59-945 cP in methanol/glycerol mixtures. Interestingly, 1 showed enhanced emission at 534 nm in 70% water compared to pure acetonitrile due to the aggregation-induced inhibited rotations. Cellular imaging suggested that 1 could successfully sense lysosomal viscosity changes induced by lipopolysaccharide, nystatin, low temperature, and dexamethasone in living cells, which could be further applied in autophagy monitoring by tracing viscosity changes. As a comparison, its analogue 2 directly linking with the phenyl group showed no viscosity-responsive or AIE properties. Therefore, for the first time, we reported a meso-benzothiazole-BODIPY-based fluorescent rotor with AIE and lysosomal viscosity-responsive properties in nervous cells, which was further applied in monitoring autophagy, and this work thus could provide an innovative strategy for the design of potential AIE and viscosity-responsive probes.

Near-Infrared Turn-On Fluorescent Probe for Aqueous Fluoride Ion Detection and Cell Imaging.

Fluoride ions are one of the most essential anions in the human body and have been implicated in various pathological and physiological processes. The detection of fluoride ions in aqueous solution, as well as the imaging of fluoride ions in living cells, remains a challenge. We herein report a BODIPY-based fluorescent probe employing a pinacol borate group as the recognition moiety for the detection of fluoride ions in aqueous solutions. This probe shows high selectivity and sensitivity to fluoride ions with a significant near-infrared fluorescence turn-on response. In addition, this probe was successfully employed in fluorescence bioimaging of fluoride ions in the human cervical cancer cell and mouse mammary cancer cell, demonstrating its good cell permeability and stability under physiological conditions.

An ultrasensitive fluorescent platform for monitoring GSH variation during ischemic stroke

The growing evidence suggests that the disturbance of redox homeostasis in the brain tissue contributes to the process of tissue damage following ischemic stroke (IS). Tracking redox process in situ is critical for diagnosis, treatment and drug design of IS. Monitoring of substances in the reduction system is more suitable than detection of molecules of oxidation system, such as reactive oxygen species (ROS), due to their difficult capture of fleeting existence. Glutathione (GSH), a dominated component of reduction system, has higher concentration than other reduced substances and is the major cellular antioxidant for maintaining the redox balance but dramatically decreases during cerebral ischemia. Efficient tools can realize the real-time tracing of endogenous GSH at ultra low concentration during ischemic damage are significant and indispensable for dynamic monitoring the process of IS. Till now, only a few fluorescent probes have been reported for imaging GSH. The direct visualization of reductive substances in the cerebral vasculature of live mice is still a significant challenge. More importantly, due to the presence of the blood–brain barrier (BBB), monitoring GSH at ultralow concentration in brain tissue seems more difficult, which tremendously restricts the development of new strategies for targeting, detecting, and imaging GSH in vivo. In present study, we successfully synthesized a series of BODIPY-based fluorescent probes, WD-1-SH, WD-2-SH, and WD-3-SH, and detected many properties, including the capacity to penetrate the BBB, sensitivity to GSH and tissue penetration depth of excitation, and emission wavelengths. In vitro experiments revealed that among the probes, WD-1-SH exhibited the highest sensitivity to reducing substances; however, WD-2-SH exhibited higher sensitivity in vivo. These desired photophysical and pharmacokinetic properties endow our fluorescent probes with the capability to monitor brain tissue low concentration GSH after injury with excellent temporal and spatial resolution. Due to these favourable properties, our fluorescent probes hold great promise for visualizing endogenous reducing substances in a variety of pathophysiological processs in vitro and in vivo, such as during stroke induced ferroptosis.

Red fluorescent BODIPY molecular rotor for high microviscosity environments

Microviscosity has a strong impact for diffusion-controlled processes in biological environments. BODIPY molecular rotors are viscosity-sensitive fluorophores that provide a simple and non-invasive way to visualise microviscosity. Although green fluorescent probes are already well developed for imaging, thick biological samples require longer wavelengths for investigation. This work focuses on the examination of novel β-substituted meso-phenyl-BODIPYs possessing a red emission. We report a new red fluorescent BODIPY-based probe BP-Vinyl-NO2 suitable for sensing microviscosity in rigid environments of over 100 000 cP viscosities. Furthermore, we demonstrate that changing the methyl position from ortho to meta on the β-phenyl-substituted conjugate BP-PH-m2M-NO2 redshifts absorbance and fluorescence spectra while maintaining viscosity sensitivity. Finally, we show that nitro-substitution of meso-phenyl is a versatile approach to improve the sensitivity to viscosity while suppressing sensitivity to polarity and temperature of such derivatives. In summary, we present two nitro-substituted red fluorescent probes that could be used as lifetime-based microviscosity sensors.