title: Selective optical sensing of biothiols with Ellman's reagent: 5,5'-Dithio-bis(2-nitrobenzoic acid)-modified gold nanoparticles.
abstract: Development of sensitive and selective methods of determination for biothiols is important because of their significant roles in biological systems. We present a new optical sensor using Ellman's reagent (DTNB)-adsorbed gold nanoparticles (Au-NPs) (DTNB-Au-NP) in a colloidal solution devised to selectively determine biologically important thiols (biothiols) from biological samples and pharmaceuticals. 5,5'-Dithio-bis(2-nitrobenzoic acid) (DTNB), a versatile water-soluble compound for quantitating free sulfhydryl groups in solution, was adsorbed through non-covalent interaction onto Au-NPs, and the absorbance changes associated with the formation of the yellow-colored 5-thio-2-nitrobenzoate (TNB(2-)) anion as a result of reaction with biothiols was measured at 410nm. The sensor gave a linear response over a wide concentration range of standard biothiols comprising cysteine, glutathione, homocysteine, cysteamine, dihydrolipoic acid and 1,4-dithioerythritol. The calibration curves of individual biothiols were constructed, and their molar absorptivities and linear concentration ranges determined. The cysteine equivalent thiol content (CETC) values of various biothiols using the DTNB-Au-NP assay were comparable to those of the conventional DTNB assay, showing that the immobilized DTNB reagent retained its reactivity toward thiols. Common biological sample ingredients like amino acids, flavonoids, vitamins, and plasma antioxidants did not interfere with the proposed sensing method. This assay was validated through linearity, additivity, precision and recovery, demonstrating that the assay is reliable and robust. DTNB-adsorbed Au-NPs probes provided higher sensitivity (i.e., lower detection limits) in biothiol determination than conventional DTNB reagent. Under optimized conditions, cysteine (Cys) was quantified by the proposed assay, with a detection limit (LOD) of 0.57μM and acceptable linearity ranging from 0.4 to 29.0μM (r=0.998).
authors: Department of Chemistry, Faculty of Engineering, Istanbul University, Avcilar, 34320 Istanbul, Turkey.
authors: Kubilay
authors: K
authors: Güçlü
authors: Mustafa
authors: M
authors: Ozyürek
authors: Nilay
authors: N
authors: Güngör
authors: Sefa
authors: S
authors: Baki
authors: Reşat
authors: R
authors: Apak
mesh_terms: D000818:Animals; D000975:Antioxidants / Q000032:analysis / Q000737:chemistry; D002623:Chemistry Techniques, Analytical / Q000379:methods*; D004228:Dithionitrobenzoic Acid / Q000737:chemistry*; D006046:Gold / Q000737:chemistry*; D057230:Limit of Detection; D053758:Nanoparticles / Q000737:chemistry*; D055095:Optics and Photonics; D051381:Rats; D017208:Rats, Wistar; D013438:Sulfhydryl Compounds / Q000032:analysis* / Q000737:chemistry
chemical_list: D000975:Antioxidants; D013438:Sulfhydryl Compounds; D006046:Gold; D004228:Dithionitrobenzoic Acid
keywords: Biothiols; Conventional 5,5′-dithio-bis(2-nitrobenzoic acid) (DTNB) assay; Gold nanoparticles; HPLC; Optical sensor