Kinetics of the transformation of n-propyl gallate and structural analogs in the perfused rat liver Gabrielle Jacklin Eler, Israel Souza Santos, Amarilis Giaretta de Moraes, Márcio Shigueaki Mito, Jurandir Fernando Comar, Rosane Marina Peralta, Adelar Bracht ⁎ Department of Biochemistry, University of Maringá, 87020900 Maringá, Brazil abstract article info Article history: Received 6 May 2013 Revised 8 August 2013 Accepted 26 August 2013 Available online 5 September 2013 Keywords: Liver Gallic acid esters Transport Transformation Distribution n-Propyl gallate and its analogs are used in foods and other products to prevent oxidation. In the liver the com- pound exerts several harmful effects, especially gluconeogenesis inhibition. The mode of transport and distribu- tion of n-propyl gallate and its kinetics of biotransformation have not yet been investigated. To fill this gap the transformation, transport and distribution of n-propyl gallate and two analogs were investigated in the rat liver. Isolated perfused rat liver was used. n-Propyl gallate, methyl gallate, n-octyl gallate and transformation products were quantified by high pressure-liquid chromatography coupled to fluorescence detection. The inter- actions of n-propyl gallate and analogs with the liver presented three main characteristics: (1) the hydrolytic re- lease of gallic acid from n-propyl gallate and methyl gallate was very fast compared with the subsequent transformations of the gallic acid moiety; (2) transport of the esters was very fast and flow-limited in contrast to the slow and barrier-limited transport of gallic acid; (3) the apparent distribution volume of n-propyl gallate, but probably also of methyl gallate and n-octyl gallate, greatly exceeded the water space in the liver, contrary to the gallic acid space which is smaller than the water space. It can be concluded that at low portal concentrations (b 50 μM) the gallic acid esters are 100% extracted during a single passage through the liver, releasing mainly gal- lic acid into the systemic circulation. For the latter a considerable time is required until complete biotransforma- tion. The exposure of the liver to the esters, however, is quite prolonged due to extensive intracellular binding. © 2013 Elsevier Inc. All rights reserved. Introduction n-Propyl-gallate is an ester formed by gallic acid and n-propanol (Fig. 1). It has been used in foods (mainly oils and fats) cosmetics, hair products, adhesives, and lubricants to prevent oxidation. It is also an im- portant component of natural products such as, for example, green tea. The compound protects against oxidation by hydrogen peroxide and oxygen free-radicals via a catalytic mechanism similar to that of the superoxide dismutase (Nakagawa et al., 1996a; Reddan et al., 2003). n-Propyl gallate presents a certain degree of toxicity, including hepato- toxicity as suggested by increased levels of circulating transaminases (Galati et al., 2006). In hepatocytes, cell death, acute inhibition of oxy- gen consumption, and ATP depletion have been observed. For these rea- sons the toxic actions of n-propyl gallate have been attributed to an action on the mitochondrial energy metabolism, but the conclusion was based on experiments done with quite high concentrations of the compound (1 to 2 mM, Nakagawa et al., 1996a, 1996b). More recently, experiments with isolated perfused rat liver revealed that n-propyl gal- late is able to affect metabolism without ATP depletion and at much smaller concentrations than those reported for the inhibition of energy metabolism in isolated hepatocytes (Eler et al., 2009). The most impor- tant effect is inhibition of glucose output, but the compound also causes other alterations. Among these is the stimulation of oxygen uptake that occurs in both mitochondria and microsomal electron transport chains. The most important cause for the diminished glucose output is likely to be the inhibition of gluconeogenesis as a consequence of the inhibition of pyruvate carboxylation (Eler et al., 2009). The latter, in turn, seems to be the consequence of an inhibition of pyruvate transport across the mi- tochondrial membrane. At least two of the observations mentioned above, namely inhibition of pyruvate transport and stimulation of the microsomal respiration, suggest complex and intense interactions of n-propyl gallate with the cellular membranes. These interactions are closely linked to transport, distribution and transformation of the compound in the liver. Although it is not difficult to solubilize n-propyl gallate in an aqueous environment at alkaline pH (Eler et al., 2009), it is evident that the compound bears a relatively strong lipophilic character, a fact that has been stressed in sev- eral studies (Frey et al., 2007; Nakagawa et al., 1996a,b). This lipophilicity must confer to the compound a certain degree of affinity for biological membranes, which could also explain its actions on mitochondria and microsomes (Eler et al., 2009; Nakagawa et al., 1996a,b). On the other hand, the metabolic transformation of n-propyl gallate seems to be quite complex and its transformation products may also exert metabolic effects, especially on n-propanol. These and other facts raise several Toxicology and Applied Pharmacology 273 (2013) 35–46 ⁎ Corresponding author at: Department of Biochemistry, University of Maringá, Avenida Colombo 5790, 87020900 Maringá, Brazil. E-mail address: adebracht@uol.com.br (A. Bracht). 0041-008X/$ – see front matter © 2013 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.taap.2013.08.026 Contents lists available at ScienceDirect Toxicology and Applied Pharmacology journal homepage: www.elsevier.com/locate/ytaap