# Carbon Nanodots as Peroxidase Nanozymes for Biosensing
## 1 Introduction
Typically, the catalytic reactions of PODs involve the oxidation of an electron-donor substrate with the simultaneous reduction of H\({}_{2}\)O\({}_{2}\) or occasionally alkyl hydroperoxide (Scheme 1a). Horseradish peroxidase (HRP) containing the heme-iron cofactor is the most abundant POD isoenzyme, which functions like a ping-pong ball, bouncing back and forth from an intermediate state (E*) to its standard state (E). Consequently, one substrate (A) converts to the product (C) and releases before the second substrate (B) binds to the enzyme. The overall process of this double displacement reaction is highlighted in Scheme 1b. The POD-like activity of a desired material, for example, C-dots, can be evaluated by measuring the concentration of H\({}_{2}\)O\({}_{2}\) using a variety of chromogenic substrates such as hydroquinone, 1,2,3-trihydroxybenzene (THB) or pyrrogallol, \(o\)-phenylenediamine (OPD), 2,2'-azino-bis-(3-ethylbenzothiozoline-6-sulfonic acid (ABTS), and 3,3',5,5'-tetramethylbenzidine (TMB). However, owing to the low carcinogenicity and high absorption coefficient of its reaction products, TMB serves as the most-studied chromogen for HRP mimics, in acidic conditions. In general, the POD-catalyzed oxidation of TMB + H\({}_{2}\)O\({}_{2}\) system involves two electron transfer steps. In the first step, a one electron oxidation generates a TMB radical cation. Two of these intermediate radical cations then form a blue-colored charge-transfer  
Figure 1: Schematic illustration of the structure of C-dots. Reproduced with permission from [19]. Copyright (2010) American Chemical Society. complex (\(\lambda\) = 370 & 652 nm), a diagnostic for HRP mimic, as shown in Scheme 1c. The cation radical is further oxidized in the second electron transfer step, producing diimine derivative of TMB (TMBDI; \(\lambda\) = 450 nm). That is why; the absorption spectrum of the oxidation product of TMB usually exhibits three absorption bands. ABTS is yet another important chromogen in POD reactions, which typically produces green color due to the formation of ABTS radical cation in the presence of H\({}_{2}\)O\({}_{2}\). The one electron transfer step in the oxidation of ABTS is shown in Scheme 1d. Very recently we have provided an authoritative detail and comprehensive account on the applications of graphene-based nanomaterials as versatile catalysts in POD-mimicking reactions [16]. In this mini review, we will provide a systematic and organized survey of C-dots as nanozymes with POD-like activities as well as their applications in biosensing. We anticipate that the years' efforts in developing C-dots as potential POD-mimic have not been covered by any research group in a review format and yet to be published. The scope intends to cover not only the C-dots but also graphene quantum dots (GQDs), doped C-dots/GQDs, C-dots/GQDs nanocomposites, and carbon nitride dots (CN-dots). However, as the synthesis and properties of C-dots have been comprehensively reported elsewhere [20; 21; 22; 23; 25], we will just briefly discuss the fundamentals of the same. Furthermore, readers are recommended to consult other potential review articles for the corresponding unique and productive details on POD enzymes as well as nanozymes [5; 8; 16; 26; 27; 28]. It is our hope that this mini review will provide a valuable insight and stimulate researchers for the development of better performing C-dots (in terms of good water dispersibility, (photo)stability and cell permeability) as nanozymes.