141 1. Introduction Kiwifruit (Actinidia deliciosa) is cultivated mainly due to its sensory properties and its ability for prolonged cold storage. This latter attribute enables consumption of kiwi- fruit throughout the year in many parts of the world (Minas et al., 2010). The most widely grown Actinidia cultivar is the A. deliciosa cv. Hayward. Commercial production of this variety has spread to many countries because of its distinctive characteristics, including fruit size, high pro- ductivity, and sufficient storageability (Ferguson, 1999). Kiwifruit is a good source of natural antioxidant sub- stances, in particular vitamin C (Nishiyama et al., 2004). In fact, the content of vitamin C in kiwifruit ranges be- tween 25 and 155 mg/100 g of fresh weight (FW) of fruit (Tavarini et al., 2009), making it higher than that deter- mined in orange, strawberry, lemon and grapefruit. Beever and Hopkirk (1990) showed that vitamin C content in ki- wifruit was tenfold higher than the same content found in apple and peach. Esti et al. (1998) have observed that the vitamin C content of kiwifruit depends on genotype, ripen- ing degree, storage and the analysis method utilized. Kiwi- fruit’s strong antioxidant capacity is due to a wide number of phytonutrients including carotenoids, lutein, phenolics, flavonoids and chlorophyll (Kaya et al., 2008). In fact, during fruit ripening, several biochemical, physiological and structural modifications occur and these changes de- termine the final fruit quality attributes (Lee and Kader, 2000; Ayala-Zavala et al., 2004). During postharvest storage of horticultural crops, im- portant changes in antioxidant status can occur (Ayala-Za- vala et al., 2004). Kiwifruit is rich in bioactive compounds, especially polyphenols (Park et al., 2009). Polyphenolic compounds are a complex group of substances that have gained enormous attention in recent years, especially within the analytical chemistry field because they exhibit important quality properties and antioxidant activity (Es- carpa and González, 2001). The antioxidant activity of phenolic compounds is mainly due to their redox proper- ties, which can play an important role in absorbing and neutralizing free radicals, quenching singlet and triplet oxygen, or decomposing peroxides (Osawa, 1994). The dark green color of kiwifruit, due to the chloro- phylls in plastids in the pericarp cells of the flesh (Talens et al., 2002), can be modified during storage. Most fleshy fruits are green only during the earlier stages of develop- ment: they undergo dramatic changes in chemical compo- sition and ultra-structure during maturation and ripening. Associated with tissue softening and the changes in carbo- hydrate and organic acid metabolism there is conversion of chloroplasts into chromoplasts and a concomitant loss of chlorophyll, often accompanied by accumulation of carot- enoids (Montefiori et al., 2009). The pigments responsible for the flesh color in ripe fruit of A. deliciosa have already been described (McGhie and Ainge, 2002; Nishiyama et Analytical statistical interpretation of relationship between different parameters of kiwifruit (Actinidia deliciosa cv. Hayward) during cold storage Sh. Shahkoomahally (1), A. Ramezanian Department of Horticultural Science, Faculty of Agriculture, Shiraz University, Shiraz, Iran. Key words: ascorbic acid, correlation, kiwifruit, total antioxidant activity, total phenolic compounds. Abstract: Physicochemical and metabolic changes of kiwifruit characteristics were studied during storage. Parameters, such as total phenolic compounds (TPC), color (L*, hue, chroma), total antioxidant activity (TAA) and ascorbic acid (AA) content, were evaluated at harvest and every 15 days during fruit storage at 0ºC and 90% ± 5 RH. Correlations between different parameters were also evaluated. The results of this study suggest that high antioxidant capacity in kiwifruit is due to a strong association (r2˃0.90) between AA and TPC, and that phenolic compounds and AA are the major con- tributors to the antioxidative activities of Actinidia deliciosa cv. Hayward. Color parameters were found to have weak correlation coefficients with TPC, TAA and AA. It is worth noting that the physical-chemical parameters do not have a relationship with the chromatic parameters. Adv. Hort. Sci., 2014 28(3): 141-145 (1) Corresponding author: shirin.shakoomahally@yahoo.com Received for publication 25 June 2014 Accepted for publication 17 September 2014 142 al., 2005); the color is mainly due to chlorophylls a and b (Nishiyama et al., 2005). The changes in bioactive compounds of kiwifruit that occur during cold storage have not previously been stud- ied, thus in this paper we describe changes in pericarp color, ascorbic acid (AA), total antioxidant activity (TAA) and total phenolic contents (TPC) in the fruits with dif- ferent storage times. The differences and correlations be- tween these parameters are also examined. 2. Materials and Methods Plant material Mature, unripe kiwifruit (Actinidia deliciosa cv. Hay- ward) of medium size (80-120 g), free from visible defects or decay, were harvested from a commercial kiwifruit orchard in Gorgan, Iran with average firmness of 10 (kg/ cm2) and 7% °Brix. Fruits were immediately transferred to the postharvest laboratory at Shiraz University. Kiwifruits were individually labeled and packaged into ventilated bags, then stored for four months at 0±1°C and 90±5% relative humidity (RH). Samples were taken at monthly in- tervals during storage for quality evaluation and analyses. Physical and physicochemical assays Total phenolic contents.The total phenolic contents of each extract were determined according to the method of Gutfinger (1981). Extracts (1 ml) at 1 mg/ml concentration were mixed with 1 ml of 2% Na 2 CO 3 . After standing for 3 min, 0.2 ml of 50% Folin-Ciocalteu reagent was added to the mixture and left to stand for 30 min. The mixture was centrifuged at 13 400×g for 5 min. The absorbance was measured at 750 nm and TPC are expressed as gallic acid quivalents (GAE). Color. Color was determined using digital imaging (Afshari-Jouybari and Farahnaky, 2011). Fruit was photo- graphed in a chamber; angle light with the horizontal sur- face of the images was 45 degrees. After transferring the images to a computer, Photoshop image processing soft- ware was performed. Individual L*, a* and b* parameters were recorded: L* is lightness, a* [-greenness to +redness) and b* (-blueness to + yellowness) are chromacity coordi- nates. The a* and b* values were converted to chroma (C* = (a*2 + b*2)1/2) and hue angle (h° = tan-1(b*/a*)]. DPPH radical scavenging activity (RSA). The free radical scavenging activity was measured by 2,2-diphe- nyl-2- picrylhydrazil (DPPH) on the basis of the method of Brand-Williams et al. (1995) with minor modifications. For this determination, aliquots (0.1 mL) of the extract were mixed with 1 mL of a DPPH solution (500 μM) in 80% ethanol. The mixture was incubated at room tempera- ture for 30 min. Solution absorbance was determined at λ =515 nm. The DPPH radical concentration was calculated using the following equation: scavenging effect (TAA %) = (1 - Af/Ao) × 100; Ao stands for the absorbance of the control sample and Af for the absorbance in the presence of the sample. L-ascorbic acid was used for the calibration curve, and the results are expressed as mg L-ascorbic acid equivalent. 100 g-1 fw (fresh weight). Ascorbic acid. Ascorbic acid was measured by the oxi- dation of ascorbic acid with 2, 6-dichlorophenol endophe- nol and the results are expressed as mg/100 g fresh weight (Rangana, 1977). Statistical analysis. Four replicates of each sample were used for statistical analysis. Correlation analyses between different parameters were carried out using the correlation and regression programme in MINITAB 16. Correlations were obtained by Pearson’s correlation coefficient (r) in bivariate linear correlations. All statistical analyses and correlations were carried out with SAS software package v. 9.1 for Windows. Differences at P<0.05 were consid- ered to be significant. 3. Results and Discussions Ascorbic acid Ascorbic acid (AA) content first increased up to 30 days (47.74 mg/100 g-1) and then decreased (Table 1) and could be a result of its synthesis during the initial storage period. The observed variation (increase) of AA content is due to fruit weight loss by dehydration (hence due to a higher concentration) or actually to AA ex novo synthesis. Utilization of AA during later storage periods may be the reason for its decreased amounts. The accumulation of AA during ripening depends on the type of fruit. Lee and Kad- er (2000) reported that AA content increased with ripening in apricot, peach and papaya, but decreased in apple and mango. Generally, when fruits become overripe, vitamin C Table 1 - Changes in AA, TPC, TAA and chromacity of kiwifruit during storage Storage time (day) Ascorbic acid (mg/100 g FW) Total phenolic compounds (mg/100 g DW) Total antioxidant activity (mg/ 100 g FW) Color parameters L* Hue Chroma 0 40.17±1.53 b 45.75 ±0.99 c 489.17±42.01 bc 51.9 ± 0.83 a 66.92±1.01 a 48.42±1.75 a 30 47.74 ±1.38 a 57.9±1.91 a 602.17±43.97 a 46.07±0.55 b 64.63±2.95 ab 45.49±1.19 b 60 40.97±2.17 b 50.65±3.35 b 556.20±70.33 ba 41.34±0.78 c 62.20±0.73 bc 39.61±0.77 c 90 35.68±2.11 c 43.3±2.56 c 434.34±47.60 dc 35.35±1.50 d 61.82±1.33 bc 35.68±1.16 d 120 27.44±1.89 d 30.45±1.65 d 354.18±40.84 d 39.92±0.82 c 59.76±1.41 c 32.62±1.73 e Means within each column with different superscript letters are significantly different (p < 0.05) for each sampling. 143 content declines concurrently with the degradation of fruit tissues (Kalt, 2006). In persimmon fruits, AA and TPC content showed a linear relationship with a positive correlation coefficient of r2=0.976 (Fig. 1). Other authors found strong correlations between AA and TPC in different fruits (Gonçalves et al., 2004; Serrano et al., 2005). Ascorbic acid content was positively correlated with antioxidant activity (r2 = 0.944), suggesting that AA makes a significant contribution to the total antioxidant capacity of kiwifruit (Fig. 2). Total phenolic compounds Total phenolic compounds of fruits increased up to 30 days (57.9 mg/100 g-1) and then decreased (Table 1). Dur- ing storage, TPC increased initially, probably due to syn- thesis from sugars, and decreased later due to its participa- tion or utilization in other metabolic processes. Tavarini et al. (2008) reported that TPC may increase or decrease in fruits and vegetables, depending on the storage conditions. The increase in TPC during storage may be the result of fruit damage and tissue disruption during storage. Pheno- lic compound synthesis in response to wounding has been reported (Saltveit, 2000). All o-quinone molecules are highly reactive and may interact with other phenols or other substances, co-poly- merise and thus produce compounds, which determine undesired fruit browning, or oxidise further compounds, reduce to original phenols, or react with different nucleo- phile compounds such as amines, thiols, imidazole, and indole. Generally there is a positive correlation between TPC availability and vulnerability to PPO attack (Ramírez et al., 2003). Phenolic compounds represent the main sub- strates used by oxidative enzymes, having consequences in terms of color and quality changes, as well as being associated with plant defense mechanisms against stress situations that can affect the postharvest period (Tomás‐ Barberán and Espín, 2001). Good correlation between TPC and AA was observed with a high significance level (P<0.001), and a similar relationship (0.97) was also obtained between TPC and TAA (Table 2). This positive and significant relationship between TPC and TAA was greater compared to AA and TAA. The results indicate strong association between an- tioxidative activities and phenolic compounds, suggesting that the latter are probably responsible for the antioxida- tive activities of kiwifruit. Phenolic compounds are also ef- fective hydrogen donors, making them good antioxidants (Rice-Evans et al., 1995). Reports in literature on the re- lationship between TPC and TAA are contradictory; some authors have observed a high correlation (Proteggente et al., 2002; Tsao et al., 2003; Khanizadeh et al., 2008). Total antioxidant activity TAA increased and peaked over the course of one month (57.9 mg/100 g-1) then decreased toward the end of the storage period (Table 1). There is debate in the literature about the influence of vitamin C on the antioxidant capac- ity of fruits and vegetables (Guo et al., 2003). However, it is also known that fruits with high antioxidant capac- ity generally contain more antioxidants and most of these antioxidants have been shown to be phenolic compounds, in particular flavonoids (Guo et al., 2003). For example in pomegranate, ascorbic acid and phenolic compounds are responsible for the TAA, alone or in combination (Kulkar- ni and Aradhya, 2005). Examining the entire storage pe- riod, it can be observed that changes in antioxidant activity Table 2 - Correlation matrix (Pearson correlation coefficients) TPC AA TAA Chroma Hue L* TPC 1.00 AA 0.98*** 1.00 TAA 0.97** 0.97** 1.00 Chroma ns ns ns 1.00 Hue ns ns ns 0.97** 1.00 L* ns ns ns 0.89* 0.86* 1.00 NS= not significant. * Significant to 0.05 p level. ** Significant to 0.01 p level. *** Significant to 0.001 p level. Fig. 1 - Correlation between TAA and AA in kiwifruit. Fig. 2 - Correlation between TAA and total phenolic compounds or ascorbic acid in kiwifruit. 144 were very similar to phenolic compounds (Lemoine et al., 2009). TAA was highly and positively related to AA (r2 = 0.944) (Fig. 2). However, in the present study, the best cor- relation (r2 > 0.96) was observed between TPC and RSA (Fig. 2). This fact probably indicates that the antioxidant capacity of kiwifruits is primarily due to TPC and AA. Color A significant decrease in hue angle was observed during the storage of fruits at 0°C, indicating continued ripening during cold storage. The initial L* value was 51.9. Internal lightness decreased sharply within 90 days of storage and then increased significantly until the end of storage (Table 1). The chroma (C*) value was initially 48.42 (Table 1). Chroma values (internal) of fruits decreased during the en- tire storage period and reached their minimum values at the end of storage. Changes in C* result principally from a loss of chlorophyll content, mostly chlorophyll a which decreases during storage (Fuke et al., 1985). In kiwifruit, lightness and chroma values significantly decreased dur- ing cold storage and shelf-life, indicating less color in- tensity (Koukounaras and Sfakiotakis, 2007). When fruits darken, skin color becomes less chromatic and surface browning develops. Diminished red skin and darkening due to oxidative browning reactions have been found to be more marked in ripe strawberries that suffer greater mois- ture loss during storage (Nunes et al., 2005). Correlation analyses showed that in the fruit tissues, most of the correlation coefficients were lower, positive and not significant. However, the correlation between chroma of kiwifruit and TPC, TAA and AA was weakly positive and significant (Table 2). 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