Analytica Chimica Acta 572 (2006) 25–31 Preventing inhibition of tyrosinase with modified electrodes Isıl Narlı, Senem Kiralp, Levent Toppare ∗ Department of Chemistry, Middle East Technical University, 06531 Ankara, Turkey Received 25 January 2006; received in revised form 22 April 2006; accepted 29 April 2006 Available online 9 May 2006 Abstract Wines, especially red wines, contain numerous biologically active compounds, the most important of which are polyphenols, whose nutritional importance is attributed to their antioxidant power. Because of this, the detection of the amount of phenolic compounds in red wines becomes extremely important. However, using free enzyme in the determination of phenolic compounds in wines cannot reflect the actual values since there are also naturally found inhibitors in red wines. In this study, benzoic acid, cinnamic acid, and sorbic acid were utilized to understand the behavior of immobilized polyphenol oxidase in the conducting polymer matrices toward inhibition. Cinnamic acid was found to be the most powerful inhibitor for both free and immobilized enzyme in copolymer matrix of poly(terephthalic acid bis-(2-thiophen-3-yl-ethyl) ester) (PTATE) with polypyrrole (PPy). In the case of immobilized enzyme in PPy matrix, it was observed that sorbic acid is a stronger inhibitor than cinnamic acid. The inhibitory effects of these inhibitors on PPO were compared with respect to both the structural differences of inhibitors and conducting polymer matrices. © 2006 Elsevier B.V. All rights reserved. Keywords: Enzyme; Tyrosinase; Biosensor; Inhibition; Immobilization; Wine 1. Introduction Tyrosinase (EC 1.14.18.1) is a copper containing oxidoreduc- tase enzyme which catalyzes two different reactions via separate active sites: (a) the orthohydroxylation of monophenols, com- monly referred to cresolase activity and (b) the oxidoreduction of orthodiphenols to orthoquinones, commonly referred to as catecholase activity. It is commonly found in yeast, mushroom, apples, and potatoes. Tyrosinase has widespread applications in medical and indus- trial fields. Tyrosinase catalyzes the synthesis of melanin through the hydoxylation of tyrosine to dihydoxyphenylalanine (DOPA) and the subsequent oxidation of DOPA to dopaquinone. The unstable dopaquinone will polymerize and precipitate into melanin. The cresolase activity of tyrosinase is of particular importance because it synthesizes DOPA. DOPA is a precursor of dopamine, an important neural message transmitter. Patients who suffer from Parkinson’s disease show a significant decrease in the concentration of dopamine found in the substantial nigra of the brain [1,2]. Also, the production of epidermal hyperpigmen- tation of melanin causes some dermatological disorders such ∗ Corresponding author. Tel.: +90 312 2103251; fax: +90 312 2101280. E-mail address: toppare@metu.edu.tr (L. Toppare). as melasma, freckles, ephelide, senile, lentignes, etc. [3]. It has been used as part of an enzyme–electrode system to detect cat- echols and assess catecholamines in the urine of patients with neural crest tumors [4]. As to the industrial aspects, tyrosinase is used in the deter- mination of phenols and its derivatives, especially for cleaning surface water and the effluent of industrial discharges. Some of the industrial sources of phenol discharge include oil refiner- ies, coke and coal conversion plants, plastics and petrochemical companies, dyes, textiles, timber, mining, and the pulp and paper industries. Virtually all phenols are toxic. Moreover, they have a high oxygen demand and can deplete oxygen [5]. As a conse- quence, this may affect the ecosystem of water sources where phenols are discharged. Tyrosinase causes the precipitation of phenols, which can then be filtered out from surface waters and industrial discharge sources. The enzyme has also been used as a sensor to detect the concentration of phenols in waste water [6,7]. Tyrosinase is responsible for enzymatic browning of some fruits and vegetables during handling and storage by affect- ing the taste and nutrition value of them. Browning has been attributed to a rapid degradation of the red pigments by tyrosi- nase, producing brown-colored byproducts [8]. Enzyme immobilization has become an important aspect of biotechnology. The usage of tyrosinase is limited because of 0003-2670/$ – see front matter © 2006 Elsevier B.V. All rights reserved. doi:10.1016/j.aca.2006.04.088