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To develop a new class of neutral fluorescent dyes with mitochondrial staining capacity, a series of functional dyes were obtained from Nile Red (2a-e) and coumarin (3a-e) with different amide compounds via Suzuki coupling reactions. The Nile Red derivatives (2a-e) emitted red fluorescence (590-660 nm) and coumarin derivatives (3a-e) showed blue emission (455-490 nm) in organic solvents. In addition, they exhibited high fluorescence quantum yields (0.27-0.98) in organic solvents and excellent photostability (>92%). Moreover, all of them possessed low cytotoxicity. More importantly, Nile Red borate (2) and coumarin borate (3) only accumulated in lipid droplets, while after being modified by different amide compounds, dyes 2a-e and 3a-e could successfully target mitochondria in HeLa cancer cells via confocal fluorescence experiments. This work provides a new strategy for the design of neutral cellular probes for mitochondrial staining.

Four different neutral fluorescent markers containing nitrogen heterocycles (quinoxaline, 1H-pyrazolo[3,4-b]pyridine, 1H-indazole and 1H-pyrrolo[2,3-b]pyridine) as targeting groups were designed and prepared in order to screen out structural units for targeting mitochondria. Several classical fluorophores (coumarin, 1,8-naphthalimide and Nile Red) were connected with these heterocycles via Suzuki coupling reactions. The derivatives of coumarin (dyes 1a and 2a-c) and 1,8-naphthalimide (dyes 3a-c) fluoresced in the blue-green region, while the Nile Red derivatives (dyes 1b and 4a-c) fluoresced in the red light region. The optical properties of the classical fluorophores, such as emission properties and photostability, were retained in the new dyes. All of them showed low cytotoxicity. Confocal fluorescence experiments in L929 normal cells and HeLa cancer cells indicated that dyes 1a-b targeted dual sites of mitochondria and lipid droplets. Moreover, dyes 2a-c, 3a-c and 4a-c targeted mitochondria; meanwhile, there are only a few mitochondria-targeting markers with neutral skeletons. Furthermore, it was found that nitrogen heterocycles with N-H bonds can improve the mitochondrial targeting ability of partial neutral fluorophores.

Bioactive oxazolopyridine unit was used in the synthesis of fluorescent markers for specific organelles in this paper. The compounds 1a-c are linked with double bond between oxazolopyridine ring and photogenic precursors (3a-c). Compound 1a showed higher fluorescence yield (0.86 in THF), compounds 1b-c showed larger stokes shifts in DMSO. In lipid vesicles environment, they also showed good optical properties. In addition, the three compounds are biomarkers with lower cytotoxicity. Among them, compound 1a based on oxazolopyridine and coumarin unit is a dual targetable fluorescent marker for mitochondria and lipid droplets; while the other two compounds 1b-c are only biomarkers for lipid droplets., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Six mitochondria/lysosomes self-targetable and viscosity-sensitive dyes ( 1a - 1f ) were developed via simple structure modification on cyanine-derived dyes. They all showed remarkable OFF-ON fluorescent response to viscosity in the near-infrared region (652-690 nm) and exhibited good linear relationship with solution viscosity. The transient absorption spectra were used to evaluate the excited-state lifetime of dye 1a in different viscosity environments. Furthermore, cellular imaging assays indicated that different derivatives ( 1a - 1f ) with the same chromophore core exhibited different organelle-targeting abilities. Among them, dyes 1a - 1c could sense lysosomal viscosity fluctuations while dyes 1d - 1f could be applied in mitochondrial viscosity detections.

In this study, a mitochondria-specific fluorescent probe for efficient ratiometric detection of Cys was designed and investigated. Probe 1 is composed of a xanthylene skeleton and a benzyl group containing an acryloyl moiety. The probe showed excellent water solubility, good selectivity and sensitivity toward Cys over other analytes, and afforded an extremely low detection limit of 33.7 nM. The possible detection mechanism was ascertained by HRMS analysis. Moreover, probe 1 had excellent mitochondrial-targeting ability (the Pearson's correlation coefficient was 0.96), and was capable of monitoring endogenous Cys in living HeLa cells by dual channel ratiometric bioimaging, demonstrating its significant potential in biological applications., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Two rosamine-based probes (1a–b) with pyronine-pyridinium skeleton were designed and prepared. Probe 1a bearing boron ester unit was oxidized and eliminated upon addition of hydrogen peroxide (H2O2), and the emission spectrum exhibited OFF–ON response accompanied by 33-fold fluorescent enhancement. In contrast, the fluorescence intensity of probe 1b enhanced 58 times after the dinitrophenyl ether part within the probe was removed by nucleophilic substitution with hydrosulfide (H2S). The design concept was based on the d-PET process in pyronine-pyridinium structures, and free 4-pyridinyl-substituted pyronine dye showing strong fluorescence was released followed by elimination. Furthermore, as biocompatible molecules, probes 1a–b have been successfully applied for imaging in live HeLa and Ges-1 cells, and all of them can serve as mitochondrial targetable probes in red channel for detecting independent species. [ABSTRACT FROM AUTHOR]

The neutral fluorescent probes 1a-1c based on D-π-A-π-D structure, with benzopyranoquinoline as the fluorophore and the electron-donating groups at both ends, were developed, which emitted dual-channel fluorescence in different pH environments. Among them, probe 1a with both N,N -diethyl group and morpholine ring as electron-donating groups exhibited the best fluorescence performance, showing a red shift of 85 nm in dual emission at 505 nm and 590 nm, and a Stokes shift of 95 nm in both channels. The neutral 1a displayed green fluorescence specifically targeting mitochondria, while the protonated form 1a+H + resulting in red fluorescence localized in lysosomes. The ability to distinguish healthy and pathological cells such as cancer or inflammation could be achieved by observing the differences in red fluorescence within lysosomes. Probe 1a could migrate and retarget between mitochondria and lysosomes, and it was discovered by the physiological effects of chloral hydrate experiments. In addition, the dynamic changes in morphology and movement of mitochondria and lysosomes during cellular autophagy through the variation of fluorescence intensity in green and red channels induced by rapamycin and starvation conditions had been investigated deeply. [Display omitted] • Novel pH sensitive probes 1a-1c based on D-π-A-π-D structure with ICT mechanism are introduced. • Probe 1a enables distinction between healthy and pathological cells through lysosomal pH monitoring. • The dual-channel strategy of probe 1a allows independent targeting of mitochondria and lysosomes, with organelle migration capability. • Probe 1a enables long-term observation of coordinated interactions between mitochondria and lysosomes during cellular autophagy. [ABSTRACT FROM AUTHOR]

As a special form of autophagy, mitophagy is a pivotal mechanism governing cellular metabolism and physiology. Viscosity monitoring is an important aspect for real-time visualization of the mitophagy process. Hence, the development of high-performance viscosity probes is crucial for investigating dynamic pathological processes related to mitophagy. This study presents the discovery of mitochondria near-infrared and aggregation-induced emission probes with neutral structure. Probes 1a - 1b were connected quinoline/naphthidine malononitrile and coumarin units with a double bond, the absorption maxima were at 433–471 nm; emission maxima were at 605–720 nm with large Stokes shifts of 170–279 nm in solvents of different polarity. As the environmental viscosity increases from 0.89 cP to 856 cP, the fluorescence intensity of probe 1a and probe 1b is enhanced by 418-fold and 769-fold, respectively, which due to the restricted rotation and inhibited twisted intramolecular charge transfer in high viscosity systems. In addition to excellent NIR, hypotoxicity, and mitochondrial targeting capabilities, probes 1a - 1b were successfully applied to detect changes in mitochondrial viscosity during starvation or rapamycin-induced mitophagy. These remarkable features make probes 1a - 1b as promising candidates for investigating dynamic pathological processes related to mitophagy with significant potential for biological applications. [Display omitted] • Two electroneutral, NIR and AIE probes with exhibit mitochondrial targeting abilily. • These probes nearly showed dramatic "OFF–ON" fluorescent response with viscosity increasing. • These probes can monitor the viscosity changes of starvation and drug-induced mitophagy in real time. [ABSTRACT FROM AUTHOR]

Three hydrogen sulfide (H 2 S) probes based on an azonia–cyanine skeleton were successfully designed and prepared. Probe 1a , containing 4-chloro-7-nitro-1,2,3-benzoxadiazole connected to the cyanine dye, had an emission at 660 nm that was enhanced 4.5-fold by the reduced photoinduced electron transfer process when reacting with H 2 S. Probes 1b and 1c were constructed from cyanine dyes with electron withdrawing 2,4-dinitrophenyl and 7-nitrobenzo[c] [1,2,5]oxadiazol-4-yl groups, respectively. Probes 1b and 1c gave off−on type responses with 169- and 17-fold fluorescent enhancements at 639 nm with H 2 S. Their emission properties were influenced by intramolecular hydrogen bonds and intramolecular charge transfer processes. The detection limits of probes 1a – 1c were calculated at 178, 121, and 9.6 nM, respectively. The intracellular imaging experiments with HeLa cells indicated probe 1a was a mitochondria-targeting H 2 S probe, while probes 1b and 1c were lysosome-targeting H 2 S probes. Image 1 • The cyanine dyes had possess different organelles targetablities with different drivatives. • OFF-ON fluorescent probes were designed based on the elimination and recovery of hydrogen bonds. • Three fluorescent probes all have good selectivity to hydrogen sulfide, and one probe has a lowest detection limit as 9.6 nM. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • Five probes (1a-e) have excellent photo stability. • Four probes (1a-d) exhibit "OFF–ON" fluorescent responses towards nucleic acids. • Probe 1a can image mitochondrial nucleic acid. • Probe 1a was a low-toxic probe for live HeLa cells. Abstract Five cyanine dyes (1a-e) with azonia unit have been prepared by the reaction of pyrido[1,2−a]pyrimidinium ions and suitable aldehydes. Before further research, their photo and thermal stabilities were determined in the first place. At least 88% absorption remained after 5 h irradiating illustrated all of these dyes had excellent photo stability. Meanwhile, the decomposition temperature of dyes was around 200 °C which demonstrated they had good thermal stability. Therefore, the good photo and thermal stability made them possible to be probes. Subsequently, the optical properties of probes toward nucleic acids had been measured. TheirOFF–ON fluorescent responses (1a - d) towards nucleic acids were investigated. In addition, confocal microscopy imaging of probe 1a in live HeLa cells indicated that probe 1a could image mitochondrial nucleic acid. [ABSTRACT FROM AUTHOR]

In this paper, a new compound ( 3 ) with pyronine-pyridine skeleton had a strong fluorescence at pH 5.0−9.0, while the emission turned off at pH = 1.0–2.0. It was a mitochondrial biomarker in cancer cells (KB, 4T1, HeLa) and normal cells (Ges-1, L929, WS1). Based on these above results, probe 1 with pyronine-pyridinium skeleton was further designed and prepared to detect nitroreductase (NTR). The nitrofuran group of probe 1 was reduced and eliminated upon addition of NTR and nicotinamide adenine dinucleotide (NADH) to produce compound 3 , and the d-PET process could be allowed in the pyronine-pyridinium structure of probe 1 but it was prohibited in the pyronine-pyridine skeleton of compound 3 after detection, hence probe 1 showed a nearly turn-on signal in red light region at 565–800 nm with emission maxima at 604 nm. The detection limit was calculated as low as 2.2 ng/mL. Furthermore, fluorescence imaging experiments with live HeLa and Ges-1 cells were performed, and probe 1 could serve as a mitochondrial targetable probe to detect nitroreductase in red channel. [ABSTRACT FROM AUTHOR]

Abnormal mitochondrial viscosity could lead to diabetes and neurodegenerative diseases, therefore three probes 1a-c for monitoring the fluctuation of viscosity were designed and synthesized in this article. Probes 1a-c were composed of 1,8-naphthalimide derivatives and benzoxazole, benzothiophene or oxazole [4,5- b ] pyridine structure through carbon-carbon double bond connection, and their emission wavelengths increased sequentially (1a : 580 nm; 1b : 614 nm; 1c : 630 nm). The single bond between the nitrogen-containing heterocycle and the carbon-carbon double bond served as the viscosity response site. When the viscosity increased from 0.95 cP to 945 cP, the fluorescence intensity of probes 1a-c increased 18.67-fold, 20.34-fold and 18.37-fold, respectively. In 99% glycerol, the fluorescence quantum yields of probes 1a-c were 85.4%, 80.8% and 34.3% respectively. In addition, the excited state lifetime of probe 1c in 80% glycerol (725 ps) was higher than that in 40% glycerol (203 ps). More importantly, the influence of solvent polarity, pH and some potential interfering substances on probes 1a-c were negligible. And cell experiments showed that probes 1a-c could be used for monitoring the fluctuation of the mitochondrial viscosity in HeLa cells. [Display omitted] • Three fluorescent probes containing nitrogen heterocyclic and 1,8-naphthalimide were designed and synthesized. • Probes 1a-c showed specific response to viscosity. They have high fluorescence quantum yield in 99% glycerol (34.3% – 85.4%). • Excited state lifetime of probe 1c in 80% glycerol was higher than that in 40% glycerol due to limited molecular rotation. • Probes 1a-c could be used to detect changes of mitochondrial viscosity in HeLa cells. [ABSTRACT FROM AUTHOR]

• Probe 1 based on benzo[ e ]indole and aromatic azonia exhibited near infrared emission and high sensitivity for viscosity. • A good linear relationship between lg (I 650nm) and lg η (viscosity values from 0.89 cP to 8.6 cP). • Probe 1 with the low concentration (500 nM) could simultaneously detect changes of viscosity in mitochondria and lysosomes. • Probe 1 could be used to distinguish between cancer cells and normal cells. • Probe 1 could detect the mitochondrial and lysosomal viscosity fluctuation during autophagy. This article reported a hemicyanine dye fluorescent probe (1), based on benzo[ e ]indole and aromatic azonia skeleton, for simultaneous detection of viscosity fluctuation in mitochondria and lysosomes. Probe 1 exhibited near infrared emission and high sensitivity towards viscosity of the medium. As the viscosity increases, the rotation of C C single bond between the benzo[ e ]indole and the nitrogen heterocycle could be suppressed, and the entire molecular scaffold was coplanar and emitted strong fluorescence due to more delocalized electron distribution. The fluorescence intensity of probe 1 was enhanced 16.2 times at 650 nm as the viscosity increased from 0.89 cP to 865 cP. Correspondingly, the fluorescence quantum yield of probe 1 increased from 4.2% to 49.6%. The change of lg (I 650nm) had a good linear relationship with lg η (viscosity values: 0.89 cP to 8.6 cP), which meant the viscosity of dilute solution could be detected quantitatively. In cell experiments, probe 1 with low concentration (500 nM) had stronger fluorescence intensity in cancer cells than in normal cells. Besides, probe 1 could monitor the viscosity changes of mitochondria and lysosomes caused by the addition of ionophores (monensin, nystatin and dexamethasone) or in the process of autophagy. The results provide a simple and effective method for the early diagnosis of related diseases. [ABSTRACT FROM AUTHOR]

• Cyanine dye with benzo[e]indol and aromatic azonia unites have dual organelles target abilities. • A probe was designed for the detection of NTR in mitochondria and lysosome simultaneously. • NIR emission, fast OFF-ON response (180 s) and low detection limit (3.2 ng/mL). In this article, a benzo[ e ]indol aromatic azonia skeleton cyanine dye (3) with excellent performance was developed successfully through simple modification of an original aromatic azonia skeleton cyanine dye. Dye 3 had mitochondria/lysosome-targeting ability and its maximum emission wavelength extended to the near infrared region (655 nm). Inspired by this, probe 1a was designed, which connecting 2-bromomethyl-5-nitrofuran to the cyanine dye 3 , and was the first mitochondria and lysosome dual-targeting probe for nitroreductase (NTR) detection. In addition to exhibiting good selectivity and fast response (180 s), probe 1a also exhibited high sensitivity to NTR (the detection limit was 3.2 ng/mL). The cell imaging experiments showed probe 1a could work as a dual mitochondria/lysosome-targeting probe to detect NTR in HeLa cells. [ABSTRACT FROM AUTHOR]

A small molecule fluorescent probe (probe 1) based on adenine-coumarin derivative was designed and synthesized in this paper. Probe 1 exhibited a significant fluorescence-enhancing response to nucleic acids at 495 nm (for DNA) and 487 nm (for RNA). The fluorescence enhancement of probe 1 for DNA and RNA was 5.68 and 9.73 times respectively, the fluorescence quantum yield was changed from 2.5% to 11.7% and 22.5% accordingly. Meanwhile, an excellent linear relationship of fluorescence intensity at 495 nm or 487 nm versus the nucleic acid concentration (1 μM for probe 1 , 0–350 μg/mL for DNA and 0–300 μg/mL for RNA) was obtained. Co-staining and nucleic acid digestion experiments showed that probe 1 could selectively image nucleic acids in mitochondria and nucleoli in HeLa cells. Unlabelled Image • An adenine-coumarin based probe was designed and synthesized. • Excellent light stability and high selectivity for DNA and RNA • Low cell cytotoxicity in HeLa cells • Image DNA in mitochondria of HeLa cells [ABSTRACT FROM AUTHOR]