Browning Potential, Phenolic Composition, and Polyphenoloxidase Activity of Buffer Extracts of Peach and Nectarine Skin Tissue

The relationship of phenolic composition and polyphenoloxidase activity (PPO, E.C. 1.14.18.1) to browning potential (BP) was studied in buffer extracts of peach [Prunuspersica L. Batsch) and nectarine [P. persica var. nectarine (L.) Batsch] fruit skin. The BP varied among cultivars with ‘Flavorcrest’ having the highest value and ‘Maycrest’ the lowest. On average, over 83% of the browning measured at the end of the 5-hour incubation occurred during the first hour. The total soluble phenolics (TSP), the total anthocyanin (TA), and glutathione content (GLU) varied among cultivars, but were not significantly correlated to the BP. Of the phenolics determined by HPLC, only chlorogenic acid had a significant positive correlation and epicatechin a significant negative correlation with BP by the first hour of incubation. The PPO activity, ranging from 4 to 11 optical density units per gram dry weight per minute among peaches and nectarines, was not significantly correlated with BP. However, no browning was detected if the buffer extract was previously boiled. These results indicated that browning in the buffer extracts of peach and nectarine skin tissue depends on the presence of PPO activity and chlorogenic acid, which are major contributors to enzymatic browning. Fruit browning, as aconsequence of bruising, is due to phenolic oxidation (Mathew and Parpia, 1971; Mayer and Harel, 1979; Vamos-Vigyazo, 1981). The destruction of fruit cellular compartmentation allows the phenolic substrates to be accessible to PPOs which catalyze the phenolic oxidation (Mayer and Harel, 1979). The concentration and composition of phenolic compounds andor activity of PPOs are often the major factors determining tissue browning development and intensity (Mathew and Parpia, 1971; Mayer and Harel, 1979). Furthermore, natural chemicals with antioxidant properties such as thio compounds may also play an important role in fruit browning development (Liyanage et al., 1993; Singleton et al., 1985). Due to the importance of visual appearance as a produce cosmetic quality parameter, tissue browning has long gained attention from horticultural researchers (Vamos-Vigyazo, 198 1). While numerous studies have used juice, whole fruit and fruit flesh systems, few of them have used skin tissue. Research on fruit skin browning, as aresult of friction damage, has only been reported for pears (Mellenthin and Wang, 1974; Ranadive and Haard, 1971). The potential for skin browning on other species including peach and nectarine have not been studied at all. Peach and nectarine skin discoloration is becoming an important problem for the California stone fruit industry especially on peach cultivars such as ‘Flavorcrest’, ‘Elegant Lady’, and ‘O’Henry’, and nectarine cultivars such as ‘Fantasia’, ‘Royal Giant’, and ‘Flaming Red’. This skin cosmetic disorder is characterized by the formation of dark brown or black spots and/or light brown spots on the surface of the fruit. Dark brown and black spots commonly called black staining or inking occur as a consequence of abrasion damage to epidermal cells in combination with contaminants (Cheng and Crisosto, 1994; Crisosto et al., 1992, 1993), but light brown spots commonly called skin browning may simply Receivedforpublication2Nov. 1994. Accepted forpublication 10Apr. 1995. This study was funded by a grant from the Califomia Tree Fruit Agreement. We thank Adel A. Kader and Betty Hess for their cooperation. The cost of publishing this paper was defrayed in part by the payment of page charges. Under postal regulations, this paper therefore must be hereby marked advertisement solely to indicate this fact. ‘Postdoctoral research associate. ?Assistant pomologist. be a result of epidermal cell abrasion (Crisosto et al., 1993). Although black staining is the main contributor to the incidence of skin discoloration on peach and nectarine fruit, skin browning may also be a potential cause of the disorder. As this disorder is critical for consumer acceptance, even when the taste of the fruit is not affected, we decided to investigate fruit skin phenolic composition and study its phenolic oxidation to enhance the understanding of stone fruit skin browning. In addition, skin tissue glutathione level was determined to examine its potential effect on browning development. Materials and Methods Plant materials and chemicals. Fruit of ‘Maycrest’ (MC), ‘Flavorcrest’ (FC), ‘Elegant Lady’ (EL), and ‘O’Henry’ (OH) peaches and ‘Mayglo’ (MG) and ‘Flaming Red’ (FR) nectarines were collected from the orchard at the Univ. of California, Kearney Agricultural Center at commercial maturity based on ground color. Among these cultivars, only MG nectarine and MC peach have a low incidence of skin browning as reported commercially under California conditions (Crisosto et al., 1992). Samples of five fruit were collected from five randomly marked trees per cultivar with four replications. Fruit were carefully harvested, washed with tap water, rinsed with distilled water, and air dried. The skin was peeled off, the flesh tissue scraped off, then frozen immediately in liquid nitrogen, freeze-dried, ground into a powder, and stored at 4OC. All of the chemicals were purchased from Sigma Co. Browning potential (BP). Skin powder (400 mg) was homogenized under a nitrogen atmosphere in 15 ml of chilled 0.1 M sodium phosphate buffer (pH 4.0) (degassed with N2) with an Ultra-Turrax homogenizer (IKA Labortechnik, Cincinnati, Ohio) for 1 min. The homogenate was centrifuged at 15 ,000~ g at 4C for 10 min. The absorbance at 420 nm (A,,,,) of the warmed-up supernatant (BP extract) was determined using a spectrophotometer (UV160; Shimadzu) at time zero and after incubation for 1 h and 5 h in a covered water bath at 30C with slow swirling. Analysis of phenolics. Soluble phenolics were extracted with methanol (methanol extract) three times and the total soluble J. AMER. SOC. HORT. Scr. 120(5):835-838. 1995 835 phenolics (TSP) determined according to Slinkard and Singleton (1977). The total phenolics (TSP) were expressed as gallic acid equivalents (CAE). Total anthocyanins (TA) were estimated as previously described (Cheng and Crisosto, 1994) and expressed as cyanadin-3-glucoside equivalents (CGE). For high-performance liquid chromatography (HPLC) analysis of phenolics, skin powder (200 mg) was homogenized as above in 5 ml 100% HPLC-grade methanol followed by centrifugation at 18,OOOx g for 20 min. Methanol of the supernatant was evaporated with a speed vac concentrator (RC1010; Jouan, Winchester, Va.). The resultant residue was dissolved in 160 pl methanol followed by 840 p1 phosphate buffer (0.1 M, pH 2.8), filtered (Millipore, 0.45-pm), and collected into an auto sampler vial for HPLC analysis according to Gorse1 et al. (1992). Phenolics were identified and concentration estimated by retention time and co-chromatography of authentic standards. PPO activity measurements. Skin powder (300 mg) was homogenized as above in 15 ml chilled phosphate buffer (0.2 M, pH 6.2). Immediately after homogenization, 5% PVPP (w/v), 2% wet Amberlite XAD-4 (w/v), and 2% Triton X-100 (by volume) were added and vortexed. After 5 min in an ice bath, the homogenate was filtered through four layers of cheesecloth and centrifuged at 20,000~ g for 20 min at 4C. The PPO activity was measured by the change in A420nm of the assay mixture (30C) that contained 0.5 ml supernatant (enzyme extract), 2.3 mlO.1 M phosphate buffer (pH 6.2), and 0.2 m10.2 M catechol (in buffer) which was added after a 5 min preincubation. The PPO activity is presented as the change in unit of OD at 420 nm per gram dry weight per minute. Analysis of glutathione. The glutathione content of 400 mg skin powder was assayed according to Liyanage et al. (1993) after extraction with 15 mlO.5% perchloric acid (v/v). Statistical analysis. The Statistical Analysis System (SAS) for the personal computer program (SAS Inst., 1988) was used for the ANOVA, LSD means separation, single, Pearson and stepwise regression analyses.