Bazı Bitki Ekstreleri ve Kimyasal Bileşiklerin Lipoksijenaz İnhibitör Aktiviteleri

IUFS Journal of Biology  IUFS J Biol 2014, 73(2): 47-52 Research Article Lipoxygenase Inhibitory Activities of Some Plant Extracts and Chemical Compounds Ozlem Sacan*, Emine Yildiz Turhan Istanbul University, Faculty of Engineering, Department of Chemistry, 34320, Avcilar, Istanbul, Turkey 1 Abstract Lipoxygenases (LOXs, linoleate: oxygen reductase, E.C. 1.13.11.12) are a family of non-heme ironcontaining dioxygenases. LOXs are associated with several inflammation-related diseases such as arthritis, asthma, cardiovascular, kidney, skin and allergic diseases, neurodegenerative disorders, cancer and metabolic syndrome. In this study, the inhibitory effects of ethanolic extracts prepared from 12 different plants and 12 different chemical compounds were investigated on the activity of LOX which has an important value in the health area. All the plant extracts and chemical substances used in our study showed LOX inhibitory effect. The enzyme inhibitory activities of the extracts and chemical compounds were increasing in a dose-dependent manner. The results obtained in the present study indicate that plant extracts and chemical compounds examined can be a potential source of novel antiinflammatory therapeutics. Keywords: Lipoxygenase inhibitory activity, plant extracts, chemical compounds. *Corresponding Author: Ozlem Sacan () (Received: 05.12.2014 Accepted: 12.12.2014) Bazı Bitki Ekstreleri ve Kimyasal Bileşiklerin Lipoksijenaz İnhibitör Aktiviteleri Özet Lipoksijenazlar (LOXs, linoleat: oksijen redüktaz, E.C.1.13.11.12) hem içermeyen, demir içeren dioksijenaz ailesidir. Lipoksijenazlar, artrit, astım, kalp damar, böbrek, deri ve alerjik hastalıklar, nörodejeneratif hastalıklar, kanser ve metabolik sendrom gibi çeşitli inflamasyon ile ilişkili hastalıklar ile ilgilidir. Bu çalışmada, farklı bitkilerden hazırlanan etanollü ekstrelerin ve bazı kimyasal bileşiklerin sağlık alanında önemli bir alana sahip olan LOX aktivitesi üzerine inhibe edici etkileri incelenmiştir. Çalışmada kullanılan tüm bitki ekstreleri ve kimyasal maddelerin LOX inhibitör etkisine sahip olduğu belirlenmiştir. Bitki ekstreleri ve kimyasal bileşiklerin enzim inhibitör aktiviteleri, doza bağlı olarak artmaktadır. Bu çalışmadan elde edilen sonuçlara göre, incelenen bitki ekstreleri ve bazı kimyasal bileşikler antiinflamatuar tedavide yeni bir potansiyel kaynak olabilirler. Anahtar kelimeler: Lipoksijenaz inhibitör aktivite, bitki ekstreleri, kimyasal bileşikler. 48 Introduction Sacan and Turhan / IUFS Journal of Biology 2014, 73(2): 47-52 Lipoxygenases (LOXs, linoleate: oxygen reductase, E.C. 1.13.11.12) are a large monomeric protein family of non-sulphur nonheme, and iron-containing dioxygenases (Brash 1999). LOXs are widely distributed in plants, animals and fungi. LOXs are associated with several inflammation-related diseases such as arthritis, allergic asthma, psoriasis, inflammatory bowel disease, cardiovascular, kidney, metabolic syndrome, skin diseases (Iversen and Kragbella 2000) and neurodegenerative disorders such as Alzheimer’s disease (Khanna et al. 2003; Moreno 2009; Dobrian et al. 2011). In addition, LOXs and their metabolites are involved in many human cancer types such as prostate, lung, breast, colon and other cancer cell lines (Samuelsson 1987). It is very important to find new LOX inhibitors for the therapy of human diseases such as cancer, cardiovascular and neurodegenerative disorders (Kelavkar et al. 2000). According to the site of oxygen insertion within arachidonic acid, LOXs are classified as 5-, 8-, 11-, 12- and 15-LOX. These 5-, 8-, 12-, and 15-LOX isoenzymes are important for mammalian organisms. Also, 5- and 15- LOXs are important for plants. The substrate for mammalian LOXs are arachidonic acid while, for plants LOXs are linoleic and α-linoleic acids (Aparoy et al. 2008). The arachidonic acid pathway is a source of powerful bioregulators such as prostaglandins, thromboxanes, leukotrienes and lipoxins (Parker 1987; Samuelsson et al. 1987; Funk 1996; Yamamato et al. 1997). This pathway is initiated by mammalian LOXs. The lipoxygenase pathway of arachidonic acid metabolism occur reactive oxygen species (ROS). Other arachidonic acid metabolites formed in this way and ROS might play a role in tumour formation and inflammation. (Juntachote and Berghofer 2005). There are many LOX inhibitors for therapeutic application available in pharmacies such as Zileuton and Minocyclines (Misra et al. 2013). Due to their side effects, these inhibitors have been forbidden or limited (Charlier and Michaux 2003). Due to such side effects, developing of new LOX inhibitor drug with minimum side effects is important (Tomy et al. 2014). In this study, in vitro enzyme inhibition potential of ethanolic extracts prepared from different plants and some chemical compounds for LOX were investigated. Materials and methods Plant material and chemical compounds All the plants were purchased from an Istanbul Local Market. Chemical compounds were obtained from Merck, Sigma–Aldrich and were of analytical grade. Preparation of the plant extracts The plant extracts were prepared by refluxing 10 g of the dried leaves with 100 ml 70% ethanol for four hours using a Soxhlet apparatus and then filtering through linen cloth at room temperature. The filtrates were evaporated to dryness in a rotary evaporator. The extracts were kept in - 20˚C until use. All the extracts were dissolved in 70% ethanol before use. Chemical compounds were also dissolved in water. In vitro lipoxygenase-inhibitory assay The inhibition of lipoxygenase activity was determined by a spectrophotometric method reported by Yawer et al. (2007). The reaction mixture, containing test compound solution (inhibition solution), lipoxygenase solution in 0.1 M phosphate buffer (pH 8.0) was incubated for 10 min at 25 ˚C. Then, the reaction was initiated by addition of a solution substrate. After 6 min, absorbans value was measured at 234 nm. Quercetin was used as standard inhibitor. The percent inhibition of lipoxygenase activity was calculated as: Inhibition (%) = (1-A/B)x100 Where A is the enzyme activity without inhibitor, B is the activity in presence of inhibitor. The IC50 was determined as the concentration of plant extracts and chemical drugs required to inhibit lipoxygenase activity by 50%. Sacan and Turhan / IUFS Journal of Biology 2014, 73(2): 47-52 Results The effect of the ethanolic extracts of plants on lipoxygenase activity was shown in Table 1. The enzyme inhibitory activities of the extracts were increasing in a dose-dependent manner. Higher inhibitory activity is associated with a lower IC50 value. Persimmon was found to have the highest LOX inhibitor activity. IC50 value was found 1.2 ± 0.7 ng/mL. On the other hand, grape was found to have the lowest LOX inhibitor activity (102.0 ± 9.4 ng/mL) (Table 1). Quercetin was used as standard. IC50 value was found 7.9± 0.5 ng/mL. This plant extract was the most active compound against t (...truncated)