ANTIOXIDANT POTENTIAL OF HYDROLYZED POLYPHENOLIC EXTRACTS FROM TARA (Caesalpinia spinosa) PODS Flor Chambi a,b, Rosana Chirinos a, Romina Pedreschi c, Indira Betalleluz-Pallardel a, Frédéric Debaste b, David Campos a (*) a Instituto de Biotecnología (IBT), Universidad Nacional Agraria La Molina-UNALM. Av. La Molina s/n, Lima, Perú b Transfers, Interfaces and Processes (TIPs), Université Libre de Bruxelles-ULB. 50 avenue F.D. Roosevelt, C.P. 165/67, 1050 Brussels, Belgium c Fresh, Food & Chains. Wageningen UR Food & Biobased Research. Bornse Weilanden 9, 6708WG, The Netherlands (*) address: INTRODUCTION Tara (Caesalpinia spinosa), a leguminous tree from Peru has traditionally and extensively been used by Peruvian folk medicine. Especifically tara pods have been consumed and used as infusions. Tara pods contain gallotannins (~ 50% w/w), which are hydrolysable tannins and are considered to be one of the most potent antioxidants known from plant sources. Gallotannins hydrolysis yields either gallic acid or tannin oligomers, which display even higher antioxidant capacity than tannins. This study aims: (1) to study the chemical hydrolysis of gallotannin extracts from tara, (2) to evaluate the hydrolsis degree (HD) of tara extracts regarding in vitro antioxidant capacity and (3) to evaluate the performance of these extracts by differential scanning calorimetry to retard soybean oil oxidation. MATERIALS AND METHODS Fresh tara pods were purchased from a local market in Caraz (Ancash, Peru). Tara pods (without seeds) were washed and air-dried at 55 ºC until a final humidity of ~ 3 % was reached. Tara pods minced Extraction Hydrolysis whole extract (WE) 80 % (v/v) acetone/water QUANTITATIVE ANALYSIS RESULTS Fig. 1. Total phenolics ( ) and gallic acid ( ) (a), and hydrolysis degree (b) evolution during chemical hydrolysis of tara extracts (with 2N H2SO4, 20 mg of GAE/mL and 100ºC). Different capital and short letters on each curve indicate significant differences (p < 0.05) as revealed by a Duncan test Fig. 2. Specific antioxidant capacity by ABTS (a), FRAP (b) and ORAC (c) assay and liphophility (d) evolution during chemical hydrolysis of tara extracts (with 2N H2SO4, 20 mg of GAE/mL and 100ºC). Different capital and short letters on each curve indicate significant differences (p < 0.05) as revealed by a Duncan test. CONCLUSIONS This study provides strong evidence about the antioxidant potential of TPHEs with different hydrolysis degree. An increase in vitro antioxidant capacity for the different TPHEs was observed as the hydrolysis degree increased, especially from 4 h de hydrolysis. At a concentration of 100 ppm TPHEs at 9 and 20 h hydrolysis significantly better antioxidant protection against soybean oil oxidation was observed than with 100 ppm TBHQ using the scanning calorimetry assay. These results indicate that 4 and 9 h of chemical hydrolysis of tara extract under the tested conditions are sufficient to obtain a product with good antioxidant properties to be utilized as an alternative source of natural antioxidants. This research was supported by the Peru Biocomercio project (Peru) and by the PIC project of the Belgian Coopération Universitaire au Développement (CUD, Belgium). REFERENCES - Arnao, M., Cano, A., & Acosta, M. (2001). Food Chemistry, 73, Benzie, I., & Strain, J Analytical Biochemistry, 239, Chirinos, R., Campos, D., Costa, N., Arbizu, C., Pedreschi, R., & Larondelle, Y. (2008). Food Chemistry, 106, 1285– Inoue, K.H., & Hagerman, A.E. (1988). Analytical Biochemistry, 169, Ou, B., Hampsch-Woodill, M., & Prior, R. (2001). J. Agric. Food Chem., 49, Salminen, J. P. (2003). Journal of Chemical Ecology, 29, Singleton, V., & Rossi, J. (1965). Am. J. Enol. Vitic., 16, Takacs-Novak K, Avdeef A (1996).. J Pharm Biomed Anal 14: 1405– Tan, C., Chen, Y., & Selamat, J. (2002). Food Chemistry, 76, ACKNOWLEDGMENTS H 2 SO 4 final 2N H 2 SO 4 concentration; 100ºC 0-28 hours Liquid-liquid partition ethyl acetate aqueous phase hydrolyzed extracts (HE) Vacuum concentration organic phase ethanol tara purified and hydrolyzed extract (TPHE) EXTRACTS PREPARATION - Total phenolics compounds. Method of Folin Ciocalteau ( Singleton and Rossi,1965) -Gallic acid and gallotannin contents. Rhodanine assay (Inoue and Hagerman,1988; Salminen, 2003). - In vitro Antioxidant Capacity. ABTS, FRAP and ORAC assays (Arnao et al., 2001; Benzie and Strain,1996; Ou et al., 2001). - Differential Scanning Calorimetry (DSC) Assay. Isothermal temperature at 140 ºC (Tan et al., 2002). - Lipophicity was examined by measuring the partition coefficients using an n-octanol/aqueous system (Takàcs- Novàk and Avdeef, 1996). -HPLC-PAD Analysis of Phenolic Compounds. Phenolic profiles were determined (Chirinos et al.,2008). (a) Fig. 3. Chromatograms of hydrolyzed tara extracts at different hydrolysis degrees: 0 h (non hydrolyzed extract) (a), 4 h (38.8% HD) (b), 9 h (93.7% HD) and 20 h (100% HD). (c) (d) Table 1. Induction periods (IP) and stabilization factors (SF) for TPHEs at different hydrolysis times using differential scanning calorimetry a Results are means  SD (n=3). Means within a column with the same letter are not significantly different (p >0.05) as revealed by a Duncan test. Hydrolysis Time (h) Induction period (min) Stabilization factor ± 0.28 b 1.07 ± 0.01 b ± 0.37 c 1.12 ± 0.01 c ± 0.36 d 1.16 ± 0.01 d ± 0.29 e 1.26 ± 0.01 e ± 0.29 e 1.27 ± 0.00 e ± 0.13 f 1.29 ± 0.00 f ± 0.04 g 1.31 ± 0.00 g ± 0.18 h 1.35 ± 0.01 h ± 0.41 h 1.34 ± 0.01 h TBHQ49.45 ± 0.25 g 1.32 ± 0.00 g Control37.35 ± 0.19 a 1.00 ± 0.00 a