Reaction-based small-molecule fluorescent probes for the detection of biologically important analytes.

Abstract

BODIPY-based dyes for the study of Fenton chemistry: Three novel BODIPY dyes are synthesized where [$O_{phenol}, N_{tert-amine}, N_{py}, X$] (X = S (thienyl), O (furyl), N (pyridyl) one pendent arm of the chelation pocket is hemi-labile and which can be used as a probe for the selective detection of $Fe^{3+}$. Also, $H_2O_2$ can be detected in Fen-ton-like conversions by $Fe^{2+}\cdot$ probe ensemble fluorescence “turn-on” responses that operate in neuronal cells with multiple inputs. A new BODIPY-based probe was synthesised with one pendent arm of the [$O_{phenol}, N_{tert-amine}, N_{py}, Se_2$], which can be used to detect $Fe^{3+}$ via enhancement in fluorescence intensity which can be used as simplistic po-dium to discriminate $Fe^{2+}$ and $Fe^{3+}$ via Fenton Chemistry utilizing hydrogen peroxide. This is the first BODIPY system which works as a 2:1 multiplexer for $Fe^{3+}$, $Fe^{2+}$ and $H_2O_2$. A novel fluorescent difluoroboroazaoxobenzazulene [8a-aza-8-boro-8,8-difluoro-7-oxo-7,8-dihydro(diethylamino)benz(f)azulene] dye has been synthesized from commercially available substrates in “one pot,” and was characterized structurally and spectroscopically, was derivatized and finally compared to a related BODIPY system that demonstrates “turn-on” $H^+$ (42-fold) and $F^-$ (~5-fold) responses. It is the first example of its type. Its 5-7-6 fused structure is unique and has some interesting structural properties that can be compared to few other related dye molecules / organic fragments. Fluorescein-based dyes for detection of biologically important analytes: Here we synthesized four different fluorescein based molecules which can be used as reaction based fluorescent probes for detection reactive oxy-gen species and biothiols. Firstly 3,5-dinitrobenzoyl ester of fluorescein was synthesised which can be used as a chemo-dosimeter for the detection of cysteine and homocysteine (310- and 290-fold) and superoxide inputs (336-fold increase) gives significant fluorescence intensity increases with detection limits of $2.13 \times 10^{-5} M$, $1.37 \times 10^{-5} M$ and $2.71 \times 10^{-5} M$ respectively. “OR” logic gating was constructed where biothiol and superoxide are used as two separate types of inputs. This dye is biorelevant and used to detect intracellular biothiol detection in neu-ronal cells. As per our knowledge, this is the first example of a novel fluorescent probe based on the nucleo-philic substitution reaction of biothiols and superoxide through ester cleavage. Dimethyl-thiocarbamoyl (DMTC) derivative was synthesized by simple DMTC protection of fluroescein. This can be used as new aqueous fluorescein-based chemodosimeter for the detection of $H_2O_2$ in living neuronal cells in the presence of $Hg^{2+}$ concentration. This probe gives “AND” logic gate responses with $Hg^{2+}$ and $H_2O_2$ with 55- and 100-fold, respectively increase in fluorescence intensity. Simple esterification of Fluorescein with vinylacetic acid affords a novel ratiometric, fluores-cent and selective probe for detection of Cys over other thiol-containing amino acids with a detection limit of $11.3 \mu M$ and 1390-fold increase in fluorescence intensity within 20 minutes. This sensing was also confirmed in living neuronal cells and in human blood plasma. Finally a fluorescein chloropropionate ester was synthesized by a simple one-step reaction. The chloropropionate caged fluorescein probe allows for prompt detection of cysteine over other biothiols e.g., ho-mocysteine with a time and limit of detection of $12.8 \mu M$. The detection was confirmed in living neuronal cells and human blood plasma. 2-(benzothiazol-2-yl)-phenol (HBT)-based probes for detection of superoxide: Two novel fluorescent probe conjugates of 2-(benzothiazol-2-yl)-phenol (HBT) (1) phosphinate ester and (2) 4-nitro ether enable ratiometric and selective superoxide detection by an established excited state intramolecular proton transfer (ESIPT) mechanism giving a 60-fold increase in fluorescence intensity and a detection limit of $3.0 × 10^{-3} M$ and $6.38 × 10^{-5} M$, respectively.