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Total polyphenols, total flavonoids, allicin and antioxidant capacities in garlic scape cultivars during controlled atmosphere storage
Postharvest Biology and Technology 131 (2017) 39–45
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Total polyphenols, total flavonoids, allicin and antioxidant capacities in garlic scape cultivars during controlled atmosphere storage
MARK
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Zobia Naheeda,b, Zhihui Chenga, , Cuinan Wua, Yanbin Wena, Haiyan Dinga a b
College of Horticulture, Northwest A & F University, Yangling, Shaanxi 712100, China Agricultural Research Station, Baffa, Mansehra, Khyber Pakhtunkhwa, Pakistan
A R T I C L E I N F O
A B S T R A C T
Keywords: Garlic scape Quality Polyphenol Flavonoids Allicin Antioxidant
Five commercial cultivars of garlic scapes were subjected to a controlled atmosphere (O2 = 2-5%, CO2 = 3–6%) at temperature = 0 ± 0.5 °C, RH = 85-95%, for 140–224 d to document the quality and related changes in components during storage in two consecutive years. Polyphenols, flavonoids, allicin, and 2,2-diphenyl-1-picrylhydrazl (DPPH) as well as the ferric ion reducing antioxidant power (FRAP), metal chelating capacity (MCC), and hydroxyl radical scavenging activity (HRSC) were analyzed to study overall antioxidant properties of garlic scapes in storage. The storage life was 224, 196, 196, 168, and 140 days for G025, G107, G2011-04, G110, and G064. G025 had the highest total polyphenol concentrations at 140 and 168 days in 2014 and 2015, respectively, whereas G2011-4 had the lowest concentrations of total polyphenols. The highest total polyphenols, total flavonoids and allicin concentrations were observed in G025, whereas G2011-04 displayed the lowest concentrations of total polyphenols and allicin in both years. For all cultivars, total flavonoid concentrations decreased with time. The highest weight loss was observed in G064 both in 2014 and 2015. The antioxidant capacity of G025 and G110 was higher than that of the other cultivars. DPPH and HRSC were highest in G025, and MCC and FRAP were high in G110 and G107 in both years. These results demonstrate that cultivar influences the rate of garlic scape deterioration, chemical composition properties and antioxidant activities.
1. Introduction Garlic scape is the flower stalk of garlic plant. The hardneck type of garlic scape is thin and has a slight garlicky scent, a moderately spicy flavor and an especially fresh and pleasant taste. Garlic flower stalk is widely consumed in various parts of Asia, especially in China, as it is a common ingredient in Chinese cuisine (Simon and Jenderek, 2003). Garlic is a rich source of health-promoting phytochemicals including antioxidants such as phenolics, flavonoids, and allicin (Lanzotti, 2006). Garlic bulbs are good source of natural antioxidants and possess potential health-promoting effects due to their high phenolic phytochemical concentrations (Nuutila et al., 2003). Phenolic concentrations in garlic are affected by agronomic and environmental factors (Waterer and Schmitz, 1994), but cultivar is the primary factor that determine this variation. Bulb firmness, pH, total soluble solids, moisture concentration, and sugar concentration have been reported to differ across 14 garlic cultivars (Volk and Stern, 2009). Most research on garlic has focused on the bulbs, while scapes of garlic receive very little attention, which have gained popularity in recent years. Garlic scapes are used for strong desirable fresh flavor in various recipes and stir-fry preparations
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Corresponding author. E-mail address: [email protected] (Z. Cheng).
http://dx.doi.org/10.1016/j.postharvbio.2017.05.002 Received 16 February 2017; Received in revised form 2 May 2017; Accepted 2 May 2017 0925-5214/ © 2017 Published by Elsevier B.V.
as well as an additive for meat products (Rekowska and Skupien, 2009). Despite its popularity as a high-value vegetable and as a source of flavorful ingredient, very limited research on the antioxidant components of garlic scapes has been carried out (González et al., 2012). The quality of garlic scapes after long-term storage affects consumer acceptance. Many factors have to be considered for optimum quality, such as genotype and pre- and post-harvest conditions. Curing methods, environmental conditions and genotype can affect maximum quality, and post-harvest handling (storage temperature and relative humidity) is essential for maintaining high quality of Allium crops (Brewster, 2008; Gubb and MacTavish, 2002). Variation in allicin, allyl methyl thiosulfinate, and allyl trans-1propenyl thiosulfinate concentrations was observed in 93 garlic cultivars (Kamenetsky, 2007). Allicin (diallylthiosulfinate), an active compound in garlic, represents approximately 70% of the overall thiosulfinate concentrations formed upon crushing of cloves (Kim et al., 2013; Lanzotti, 2006). Garlic cloves contain a large amount of alliinase in sheath cells and alliin in storage cells; these compounds interact once garlic is damaged and alliin is chemically converted to alkenyl sulfinyl compounds (Ellmore and Feldberg, 1994). Allicin has antimicrobial,
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2.2. Determination of total polyphenols, total flavonoids, antioxidant capacity and weight loss
anti-inflammatory, antithrombotic, and anticancer properties (Beato et al., 2011; Bommareddy et al., 2016; Kim et al., 2013; Lanzotti, 2006). Garlic scape can be stored in controlled atmosphere for 10 months at a temperature of 0 ± 0.5 °C and a relative humidity of 85–95% to fill supply gaps in the market, but the consumers complain about the deterioration of quality compared with freshly harvested scapes (Pers.com.). With respect to the quality of green onion cultivars in a modified atmosphere, the total polyphenol concentration was reduced by 6.6% after 20 days of storage (Viškelis et al., 2012). The objective of this study was to investigate the deterioration of quality in different garlic scape cultivars during long-term storage.
50 g of scapes were subsampled from each replicate, homogenized and extracted in 200 mL of ethanol: acetone (7:3, v/v) for 1 h at 37 °C (Lee and Wicker, 1991). The extract was filtered through Whatman No. 41 paper and rinsed with 50 mL of ethanol: acetone (7:3, v/v). Extraction of the residue was repeated using the same procedure. The two combined filtrates were stored at −20 °C until total polyphenol, total flavonoid and antioxidant capacity analyses were carried out. Total polyphenols (TP) were determined using the Folin-Ciocalteu method (Jayaprakasha et al., 2001) with minor modifications. In a 10mL Eppendorf tube, 7.9 mL of distilled water, 0.1 mL of garlic scape extract and 0.5 mL of Folin-Ciocalteu reagent (1:1 with water) were added and mixed. After 1 min, 1.5 mL of sodium carbonate (1.8 mol L−1) was added. The prepared mixture sat at room temperature in the dark for 2 h. The absorbance was measured at 765 nm; the total polyphenol concentration was calculated from the calibration curve using gallic as the standard. All concentrations were expressed as grams per kilogram on a fresh weight basis. Total flavonoids (TF) were determined according to the methods of Yong et al. (2008). In a 10-mL Eppendorf tube, 0.3 mL of garlic scape extract, 3.4 mL of 30% ethanol, 0.15 mL of 0.5 mol L−1 NaNO2 and 0.15 mL of 0.3 mol L−1 AlCl3 were added and gently mixed. After 5 min, 1 mL of 1 mol L−1 NaOH was added. The absorbance was measured at 506 nm. The total flavonoid concentration was calculated from a calibration curve using rutin as the standard. All concentrations were expressed as grams per kilogram on a fresh weight basis. Weight loss of each replicate was measured every 28 d in 2014 and 56 d in 2015. Weight loss was calculated on the basis of differences between the initial and final weights recorded for each sample. Cumulative weight loss was expressed as the percentage loss of the initial total weight.
2. Materials and methods 2.1. Samples and storage conditions Garlic scape cultivars G110, G107, G2011-4, G025, and G064 were obtained from the germplasm program of the Horticultural experimental station of Northwest A & F University, Yangling, China. Garlic scapes were harvested during March and April in 2014 and 2015 respectively, at the commercial ripening stage with no disease and pest damage, without aging of the base and bracts. The scapes had fresh green color and crisp texture, and were harvested at the proper maturity stage (floral axis with bending hooks), without deformities. The garlic scapes were harvested on sunny days, transported to lab immediately in wellventilated conditions and precooled at 5 °C before being kept in cold storage. The base of the scape was trimmed, cleaned and cured in precooling room for 24 h. In 2014, data were recorded at 28 d intervals, whereas in 2015, data were recorded at 56-d intervals. The cultivars were stored at 0 ± 0.5 °C (RH = 85-95%, O2 = 2-5%, CO2 = 3-6%) for a maximum of 224 d, following the commercial storage recommendations (SBT 10887/2012). The fresh garlic scapes were stored at the eating-quality stage. Three boxes each of 360 L had two nozzles, one for injecting O2 and the other for CO2. After taking samples, the lid was closed, and nitrogen gas was slowly injected until the O2 reached the desired level through one nozzle. Then, the nozzle was quickly closed, and CO2 was slowly injected through the second nozzle, after which it was closed upon reaching the desired limit. A general analyzer CYCK401 (Yantai Venture of Measurement and Control Engineering limited, Xian, China) was used to measure temperature, relative humidity, O2 and CO2 concentrations. The same procedure was repeated every time in sampling. Commercial garlic scapes are usually stored in the cold storage room, and the boxes were used only for experiment. Three replicate samples of 1 kg each, were placed on the shelves in 3–4 layers, 300 mm apart. The boxes were kept open for ventilation 2–3 times per month for 15–20 mins each time. O2 and CO2 were maintained at 2–5% and 3–6%, respectively, to avoid sulfur-containing volatiles. Refrigeration time varied for different cultivars. The refrigeration time was 150–300 days. 150 days, the CO2 was kept at the high limit index and O2 at the low limit index; after 150 days, the CO2 was lowered to the low limit, and O2 was raised to the high limit index. Quality evaluations were performed on each sampling day. A panel of five experts subjectively assessed the samples for each quality characteristic (appearance, taste and texture) using a 1–5 scale. In the case of appearance, a scale composed of pictures and a brief description for each score value was used as follows: 5 = excellent, no defects; 4 = very good, minor defects; 3 = fair, moderate defects; 2 = poor; major defects; and 1 = inedible. A score of 3 was considered the limit of marketability and a score of 2 as the limit of edibility. Due to the small size of our taste test panel and the difficulty in tasting several garlic scape samples at one time, only generalizations from their records are presented.
2.3. Determination of allicin using HPLC For allicin analysis, we followed the procedure of Tan (2008). 10 g of scapes were subsampled from each replicate and crushed in 50 mL of methanol. The samples were then centrifuged at 4 °C at 12,000g for 20 min, after which 1 mL of supernatant was filtered through 0.45-μm filter paper. An allicin standard was obtained from Sigma International Co., China. For HPLC, we followed the procedure of Fujisawa et al. (2008), with some modifications. Quantitative analyses were carried out on an HPLC machine (Waters 600E, Milford, USA). The column dimensions were 150 × 4.6 mm, with a 5-μm thickness (C18 Aeris Peptide, Dikma Technologies, China) operating at a constant flow rate of 1.0 mL min−1 at 25 °C. An injection of 10 μL was used for the standard and for all samples. Detection was carried out using a UV detector at 220 nm. The mobile phase was as follows: methanol (9); water (41); acetonitrile (50). Quantification of allicin was performed by comparing the peak area produced by crushed garlic scape extracts with that of authentic allicin. The results of allicin were expressed as milligrams per kilogram on a fresh weight basis. 2.4. Measurements of antioxidant enzyme activity The ability to scavenge 2,2-diphenyl-1-picryl-hydrazl (DPPH) was determined using the method of Hatano et al. (1988). One milliliter of each extract was added to 1 mL of the DPPH radical solution in methanol (the final concentration of DPPH was 0.1318 mmol L−1). The mixture was then stirred vigorously and sat for 30 min; the absorbance of the resultant solution was measured at 517 nm with a spectrophotometer. DPPH was calculated as follows: Scavenging% = (1– Asample)/Acontrol) × 100 40
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cultivar G2011-4 (4.39–3.75 g kg−1). The experiment in 2015 showed the same trends.
Ferric ion reducing antioxidant power (FRAP) was measured as described by Prakash et al. (2007). First, 100 μL of the sample, 2.5 mL of phosphate buffer (0.2 mol L−1, pH 6.6), and 2.5 mL of potassium ferricyanide solution (1%) were sequentially mixed. The mixture was then incubated in a 50 °C water bath for 20 min before cooling. After cooling, 2.5 mL of 3-chloroacetic acid (10%) was added to the solution and mixed thoroughly. The 2.5-mL mixture was extracted into a new tube, and 2.5 mL of distilled water was added. Finally, 300 μL of FeCl3 (0.1%) was added to the mixture, and the reaction was allowed to proceed for 5 min at room temperature in a completely dark room. The absorbance of the product was measured at 700 nm, and FRAP was expressed as this absorbance. The hydroxyl radical scavenging activity (HRSC) was estimated following the methods of Prasad et al. (1996). Briefly, 3 mL of distilled water and 100 μL (0.02 mol L−1) of FeSO4 were added to a microfuge tube. Then, 45 μL of H2O2 (0.15% solution) and 1 mL (8 mmol L−1) of salicylic acid were added. The final volume of the reaction mixture was then added to 100 μL of the sample solution and kept in the dark for 50 min at 20 °C. The absorbance at 510 nm was recorded, and the HRSC was calculated as follows:
3.2. Total flavonoids TF in garlic scapes also varied among the cultivars (Fig. 1). Total flavonoids in the cultivars ranged from 0.927 to 0.357 g kg−1 in 2014 at 140 d (Fig. 1) and from 0.678 to 0.226 g kg−1 in 2015 at 168 d (Fig. 1). In 2014, the highest TF were observed in G025, ranging from 0.927 to 0.538 g kg−1 during 140 d of storage, whereas the lowest TF were observed in G107 (0.585 to 0.357 g kg−1) after the 140-d cold storage period. In 2015, TF were highest in G025 (0.678–0.237 g kg−1) and G107 (0.678 to 0.237 g kg−1) but lowest in G2011-4 (0.474–0.227 g kg−1). Our results in two consecutive years revealed that TF declined throughout the storage period. 3.3. Allicin concentrations In 2014, the highest allicin concentration was observed in G025, increasing from 2.80–13.75 mg kg −1 after 140 d of storage, whereas the lowest allicin concentration was observed in G107 (2.08–6.84 mg kg −1) after 140 d of storage (Fig. 1). In 2015 (Fig. 1), G025 again had the highest allicin concentration (from 3.05–8.01 mg kg−1). The lowest concentration was observed in G2011-4 (1.45–10.02 mg kg−1). A similar result of G025 was obtained in 2015, in which the allicin concentrations increased as the storage period increased.
Scavenging% = (1– Asample)/Acontrol) × 100 The metal chelating capacity (MCC) was measured using the method of Prakash et al. (2007), in which 100 μL of extract was mixed with 3.9 mL of distilled water, 100 μL (2 mmol L−1) of FeCl2, and 50 μL (5 mmol L−1) of ferrozine. The reaction mixture was kept in the dark for 10 min, and the Fe2+ concentration was monitored at 562 nm. The percentage of chelating capacity was expressed as follows:
3.4. Antioxidant capacities
Scavenging% = (1– Asample)/Acontrol) × 100
The highest FRAP (2.60 to 1.10) was observed in G107 and G064, whereas the lowest FRAP was observed in G110 (2.30 to 1.10) after 140 d (Fig. 2). DPPH quickly decreased in descending order; the highest DPPH was observed in G025 (20 to 8%). The MCC exhibited a slight decrease among days; the MCC before treatment was high in G110 (85 to 5%), and the lowest value was observed in G025 (39 to 20%) after 140 d. The HRSC exhibited a gradual decrease; the highest HRSC was noted in G025 (from 86 to 38%), whereas the lowest value was estimated in G110 (68 to 16%) at 140 d. The highest FRAP was observed in G110 at the beginning from 0.18 to 0.13, whereas the lowest was noticed in G2011-4 (0.16–0.11) and G025 (0.16–0.11) at 168 d (Fig. 2). The highest DPPH was observed in G025 (15 to 5%), whereas the lowest was observed in G064 (10 to 5%) at 168 d. The highest MCC was noted in G110 (44 to 11%), with the lowest value recorded in G2011-04 (32 to 11%) at 168 d. G025 showed the highest level of HRSC (25 to 9%), whereas G110 displayed the lowest level of HRSC (18 to 2%) at 168 d. The experiments in 2015 provided further evidence for the same result.
where Acontrol is the absorbance of the blank control (containing all reagents except the extract solution) and Asample is the absorbance of the test sample. 2.5. Statistical analysis This research was conducted with three replicates for each cultivar. Statistical analyses were carried out with the help of SPSS Statistics 17.0 software (SPSS Inc., Chicago, IL, USA). The data were subjected to one-way ANOVA, and the means were separated by Duncan’s multiple range tests. All tests were carried out in triplicate, and the results were presented as the means ± SE. Differences at P < 0.05 were considered significant. Correlations were estimated with the Pearson test at the P < 0.01 and P < 0.05 highly significant and significant levels, respectively. Correlation analysis was performed after 140 days in 2014 and after 168 days in 2015. 3. Results
3.5. Weight loss 3.1. Total polyphenols The highest weight loss was determined in G064 (0 to 10%) at 140 d. This loss was closely followed by G025 at 140 d, while the remaining cultivars showed the same value in 2014 (Fig. 3). The highest weight loss was observed in G064 (0 to 13%), and the rest of the cultivars showed the same values in 2015 (Fig. 3). A similar result was obtained in 2015 in which G064 showed the highest weight loss.
Qualitative evaluations were performed on garlic scape samples of five cultivars each after 28 days in 2014 and 56 days in 2015. The five tasting panel members agreed that G025 retained high-quality characteristics (appearance, taste, and texture) for 224 days. In 2014, G025, G107, G2011-04, G110 and G064 retained quality characteristics for 224, 196, 196, 168 and 140 days, respectively. TP varied among the different cultivars (Fig. 1). TP in different garlic scapes decreased from 3.98 to 2.22 g kg−1 at 140 d in 2014 (Fig. 1) and from 4.78 to 3.39 g kg−1 at 168 d in 2015 (Fig. 1). In 2014, the most extreme changes were recorded in cultivar G025, in which TP decreased during 140 d of storage from 3.98 to 2.59 g kg−1. The lowest TP were observed in cultivar G2011-4, decreasing from 2.91–2.22 g kg−1 during the 140 d storage period. In 2015, the highest TP was noted in cultivar G025 (4.78–3.80 g kg−1), whereas the lowest value was observed in
3.6. Correlation among total polyphenols, total flavonoids, allicin and antioxidant capacity Correlation analysis was used to explore the relationships among the different antioxidant variables measured for five garlic cultivars (Tables 1 and 2). Strong correlations were observed between TP and TF and among DPPH, FRAP, MCC, and HRSC in both years. TF were positively and highly correlated with TP. FRAP showed strong, positive 41
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Fig. 1. Total polyphenols, total flavonoids, allicin are represented in figure A, B, and C for the year 2014 while in figure D, E and F for the year 2015. Vertical bars represent the standard error of the means (n = 3).
ment with the results of Kevers et al. (2007), who concluded that polyphenol compounds in leeks decreased during initial stages of storage at 4 °C but stabilized afterwards. Petropoulos et al. (2016) reported that total phenol concentration decreased at 5 °C and 25 °C with 60–70% relative humidity in Greek garlic. In onion, flavonoid concentration increased to 64% when stored for 168–196 d in cold storage, with 58% increase noted after the first 84 d (Leja et al., 2003; Rodrigues et al., 2010). It was noted in most cases that the increase of total phenols concentrations during storage was accompanied by a decrease of anthocyanins. Our results are in agreement with those of Viškelis et al. (2012), who reported that total polyphenol concentrations of green onion after 20 days of modified atmosphere (10% CO2, temperature of 0 ± 0.5 °C and relative humidity of 95%) decreased by 6.6%. However, there was no significant difference observed in radical scavenging activity of green onion before and after the storage.
correlations with TP and TF. DPPH was highly correlated with TP, TF and FRAP. The MCC showed strong, positive correlations with TP, TF, FRAP and DPPH. Last, the HRSC was highly correlated with TP, TF, FRAP, DPPH and MCC. 4. Discussion The main quality parameters of garlic scapes that are considered to be necessary for human consumption are water content, nutrients and antioxidants. This study discussed the impact of a controlled atmosphere on the health-promoting compounds of garlic scapes, including total polyphenols, total flavonoids, antioxidant capacity, and allicin concentrations. The decreasing trend noted in total polyphenol compounds after postharvest processing of the garlic scape samples is in partial agree42
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Fig. 2. FRAP, DPPH, MCC, and HRSC are represented in figure G, H, I and J for the year 2014 while K, L, M and N for the year 2015. Vertical bars represent the standard error of the means (n = 3).
cultivar, and garlic ecotype on the concentrations of individual organosulfur compound exists. The quality parameter of weight loss is quite crucial, as this parameter has a strong impact on the appearance and shape of produce when it deteriorates or becomes disfigured due to water loss. In our study it was noted that significant weight loss occurred when garlic scapes were stored for 84 d; this weight loss may be due to the drying activity starting from the exterior surface and extending to the interior core area (Bloem et al., 2011). The metabolism process may also have a role in moisture loss in garlic cloves. The changes in water may be related to substance metabolism in the garlic scapes. In a comparative study in asparagus, it was noted that asparagus becomes unsalable for consumption when 8% weight loss occurs (Siomos, 2003). The weight loss increased progressively upon storage in asparagus and was mainly attributed to transpiration due to differences between the atmosphere
Flavor compounds that are important quality parameters are affected by many factors, such as storage temperature, cultivar, and storage duration (Bloem et al., 2011; Randle and Lancaster, 2002). Allicin concentrations were increased after 56 d of controlled atmosphere storage, the first time that this effect has been shown, although concentrations of sulfur compounds in other Allium species have been described by many researchers. Sukkaew and Tira-umphon (2012) reported increased allicin concentrations when garlic was stored at 4–6 °C with 80–90% relative humidity. Some management practices, such as cutting the basal portion during the storage period, had a negative influence on allicin concentration. In a study by Tsouvaltzis et al. (2007), removal of the basal portion of leeks showed a significant decrease of thiosulfinate concentrations during the storage period of 7 d at a temperature of 10 °C. Montaño et al. (2011) argued that a significant effect of location, 43
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Fig. 3. Weight loss is represented in figure O for the year 2014 while in figure P for the year 2015. Vertical bars represent the standard error of the means (n = 3).
and antioxidant capacity parameters and between each phenolic compound and antioxidant capacity measurement for 5 garlic scape cultivars in 2014 and 2015. The analyses showed highly significant correlations between total polyphenols and total flavonoids and among DPPH, FRAP, MCC and HRSC. There were also highly significant correlations among DPPH, FRAP, MCC and HRSC antioxidant assay measurements in 2014 and 2015. These findings indicated that the DPPH, FRAP, MCC and HRSC methods are stable and reliable for measuring antioxidant capacities of garlic scape cultivars.
and the asparagus surface Park et al. (1994). In this study, 4 in vitro assays (DPPH, HRSC, FRAP, and MCC) were used as complementary methods to evaluate the potential antioxidant activity of garlic cultivars. Significant differences were observed among different cultivars in these assays. The DPPH-scavenging activity of garlic scapes ranged from 3.41 to 20.81% in 2014 and from 3.41 to 15.10% in 2015 studies, exceeding the initial range (from 5.07 to 11.36%) in Allium species reported by Nencini et al. (2011). Queiroz et al. (2009) reported contrasting results: DPPH values were much higher in fried garlic than fresh garlic (21.68%). Decreased antioxidant capacity results is in agreement with the results of Kevers et al. (2007), who reported a decrease in antioxidant capacity measured with a DPPH assay in the white stem portion of leeks during storage for 23 d. Our results are also in line with those of Petropoulos et al. (2016), who reported that DPPH radical scavenging activity decreased significantly in three genotypes of Greek garlic when stored at 5 °C and 25 °C with 60 to 70% relative humidity for 196 d. Chen et al. (2013) reported contrasting results in which values of antioxidant capacities measured by DPPH, FRAP, MCC and HRSC methods were low compared to our results. In addition to the differences resulting from method use, the observed differences in the reported data can be explained by extraction procedure, cultivar and weather conditions of the production season (Michiels et al., 2012). The antioxidant activity was reduced by 29% after 6 weeks at 5 °C and by 36% when onion bulbs were stored in warm ambient conditions (Gennaro et al., 2002). The results and studies discussed above were conducted using shorter storage period in comparison to our studies; this shorter period may result in higher losses of antioxidant capacity. Contrasting results were reported by Leja et al. (2001), in which an increase in antioxidant capacity was observed in broccoli flower buds (Brassica oleracea var. italica cv. Lord) stored at 20 °C and at 5 °C for 3 and 7 d, respectively. Correlation analysis was performed among the phenolic compounds
5. Conclusion The variation in chemical composition in garlic scapes among different cultivars was investigated for the first time. G025 had the longest shelf life and the highest total polyphenol, flavonoid, and allicin concentrations, and DPPH and HRSC were observed with the lowest weight loss, along with MCC. Controlled atmosphere storage for 224 d did not negatively affect the antioxidant capacity and total phenolic concentrations. However, controlled atmosphere storage was associated with an increase in allicin concentrations. High amounts of natural antioxidants such as phenolic compounds in all cultivars makes garlic scape even more important for daily consumption. During controlled atmosphere storage, antioxidant properties were highly correlated with the presence of phenolic compounds.
Acknowledgments This research was supported by the National Natural Science Foundation of China (Project #31471865) and the China Scholarship Council.
Table 1 Correlation coefficients among antioxidant capacity (FRAP, DPPH, MCC, and HRSC), total polyphenols (TP), total flavonoids (TF) and allicin in garlic scapes (2014). Index
Total polyphenols
Allicin
Total flavonoids
FRAP
DPPH
MCC
HRSC
Total polyphenols Allicin Total flavonoids FRAP DPPH MCC HRSC
1 −0.574** 0.488** 0.509** 0.704** 0.520** 0.547**
1 −0.494** −0.719** −0.685** −0.687** −0.636**
1 0.611** 0.773** 0.449** 0.716**
1 0.721** 0.623** 0.687**
1 0.700** 0.809**
1 0.823**
1
** Correlation is significant at the 0.01 level.
44
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Table 2 Correlation coefficients among antioxidant capacity (FRAP, DPPH, MCC, and HRSC), total polyphenols (TP), total flavonoids (TF) and allicin in garlic scapes (2015). Index
Total polyphenols
Allicin
Total flavonoids
FRAP
DPPH
MCC
HRSC
Total polyphenols Allicin Total flavonoids FRAP DPPH MCC HRSC
1 −0.723** 0.539* 0.639** 0.691** 0.645** 0.655**
1 −0.743** −0.887** −0.773** −0.774** −0.730**
1 0.711** 0.784** 0.781** 0.640**
1 0.851** 0.867** 0.701**
1 0.919** 0.819**
1 0.766**
1
* Correlation is significant at the 0.05 level. ** Correlation is significant at the 0.01 level. and environmental factors on organosulfur compounds in garlic (Allium sativum L.) grown in Andalusia, Spain. J. Agric. Food Chem. 59, 1301–1307. Nencini, C., Menchiari, A., Franchi, G.G., Micheli, L., 2011. In vitro antioxidant activity of aged extracts of some italian allium species. Plant Foods Hum. Nutr. 66, 11–16. Nuutila, A.M., Puupponen-Pimiä, R., Aarni, M., Oksman Caldentey, K.M., 2003. Comparison of antioxidant activities of onion and garlic extracts by inhibition of lipid peroxidation and radical scavenging activity. Food Chem. 81, 485–493. Park, H.J., Chinnan, M.S., Shewfelt, R.L., 1994. Edible coating effects on storage life and quality of tomatoes. J. Food Sci. 59, 568–570. Petropoulos, S.A., Ntatsi, G., Fernandes, Â., Barros, L., Barreira, J.C.M., Ferreira, I.C.F.R., Antoniadis, V., 2016. Long-term storage effect on chemical composition, nutritional value and quality of Greek onion landrace Vatikiotiko. Food Chem. 201, 168–176. Prakash, D., Singh, B.N., Upadhyay, G., 2007. Antioxidant and free radical scavenging activities of phenols from onion (Allium cepa). Food Chem. 102, 1389–1393. Prasad, K., Laxdal, V.A., Yu, M., Raney, B.L., 1996. Evaluation of hydroxyl radicalscavenging property of garlic. Mol. Cell. Biochem. 154, 55–63. Queiroz, Y.S., Ishimoto, E.Y., Bastos, D.H.M., Sampaio, G.R., Torres, E.A.F.S., 2009. Garlic (Allium sativum L.) and ready-to-eat garlic products: in vitro antioxidant activity. Food Chem. 115, 371–374. Randle, W.M., Lancaster, J.E., 2002. Sulphur compounds in Alliums in relation to flavour quality. In: Brewster, J.L. (Ed.), Onions and Other Vegetable Alliums. Commonwealth Agricultural Bureau, International, Wallingford, pp. 329–356. Rekowska, E., Skupien, K., 2009. The influence of selected agronomic practices on the yield and chemical composition of winter garlic. Veg. Crops Res. Bull. 70, 173–182. Rodrigues, A.S., Pérez-Gregorio, M.R., García-Falcón, M.S., Simal-Gándara, J., Almeida, D.P.F., 2010. Effect of post-harvest practices on flavonoid content of red and white onion cultivars. Food Control. 21, 878–884. Simon, P.W., Jenderek, M.M., 2003. Flowering, seed production, and the genesis of garlic breeding. In: In: Janick, J. (Ed.), Plant Breeding Reviews, vol. 23. John Wiley and Sons. Oxford, UK, pp. 211–244. Siomos, A.S., 2003. Quality, handling and storage of white asparagus. In: Dris, R., Niskanen, R., Jain, S.M. (Eds.), Crop Management and Post Harvest Handling of Horticultural Crops. Science Publishers, Inc, Enfield, New Hampshire, USA, pp. 65–88. Sukkaew, P., Tira-umphon, A., 2012. Effects of storage conditions on allicin content in garlic (Allium sativum). Acta Hortic. 969, 209–212. Tan, Z.j., 2008. Determination of Allicin and Garlicin in garlic by HPLC. Gu Izhou Sci. 26, 76–79. Tsouvaltzis, P., Gerasopoulos, D., Siomos, A.S., 2007. Effects of base removal and heat treatment on visual and nutritional quality of minimally processed leeks. Postharvest Biol. Technol. 43, 158–164. Viškelis, Pranas, Bobinaitė, Ramunė, Lepse, Liga, Lepsis, Janis, Viškelis, Jonas, 2012. Quality changes of green onions stored in modifed atmosphere. Scientific works of the institute of Horticulture, lithuanian research centre for Agriculture and Forestry and aleksandras stulginskis university Sodininkystė ir daržininkystė 31, 1–2. Volk, G.M., Stern, D., 2009. Phenotypic characteristics of ten garlic cultivars grown at different North American locations. HortScience 445, 1238–1247. Waterer, D., Schmitz, D., 1994. Influence of variety and cultural practices on garlic yields in Saskatchewan. Can. J. Plant Sci. 743, 611–614. Yong, S.P., Soon, T.J., Seong, G.K., Buk, G.H., Patricia, A.A., Fernando, T., 2008. Antioxidants and proteins in ethylene-treated kiwifruits. Food Chem. 107, 640–648.
References Beato, V.M., Orgaz, F., Mansilla, F., Montaño, A., 2011. Changes in phenolic compounds in garlic (Allium sativum L.) owing to the cultivar and location of growth. Plant Foods Hum. Nutr. 66, 218–223. Bloem, E., Haneklaus, S., Schnug, E., 2011. Storage life of field-grown garlic bulbs (Allium sativum L.) as influenced by nitrogen and sulfur fertilization. J. Agric. Food Chem. 59, 4442–4447. Bommareddy, A., VanWert, A.L., McCune, D.F., Brozena, S.L., Witczak, Z., Singh, S.V., 2016. The role of organosulfur compounds derived from allium vegetables in cancer prevention and therapy. In: Ullah, F.M., Ahmad, A. (Eds.), Critical Dietary Factors in Cancer Chemoprevention. Springer International Publishing, Switzerland, pp. 111–152. Brewster, J.L., 2008. Onions and Other Vegetable Alliums, 2nd edition. Commonwealth Agricultural Bureau International, Wallingford, UK. Chen, S., Shen, X., Cheng, S., Li, P., Du, J., Chang, Y., 2013. Evaluation of garlic cultivars for polyphenolic content and antioxidant properties. PLoS One 8, 1–12. Ellmore, G.S., Feldberg, R.S., 1994. Alliin lyase localization in bundle sheaths of the garlic clove (Allium sativum). Am. J. Bot. 81, 89–94. Fujisawa, H., Suma, K., Origuchi, K., Kumagai, H., Seki, T., Ariga, T., 2008. Biological and chemical stability of garlic-derived allicin. J. Agric. Food Chem. 56, 4229–4235. Gennaro, L., Leonardi, C., Esposito, F., Salucci, M., Maiani, G., Quaglia, G., Fogliano, V., 2002. Flavonoid and carbohydrate contents in tropea red onions: effects of homelike peeling and storage. J. Agric. Food Chem. 50, 1904–1910. González, R.E., Sance, M., Soto, V.C., Galmarini, C.R., 2012. Garlic scape, an alternative food with human health benefits. Acta Hortic. 969, 233–237. Gubb, I.R., MacTavish, H.S., 2002. Onion pre- and post-harvest considerations. In: In: Rabinowitch, H.D., Currah, L. (Eds.), Allium Crop Science: Recent Advances 3. pp. 233–265. Hatano, T., Kagawa, H., Yasuhara, T., Okuda, T., 1988. Two new flavonoids and other constituents in licorice root—their relative astringency and radical scavenging effects. Chem. Pharm. Bull. 36, 2090–2097. Jayaprakasha, G.K., Singh, R.P., Sakariah, K.K., 2001. Antioxidant activity of grape seed (Vitis vinifera) extracts on peroxidation models in vitro. Food Chem. 73, 285–290. Kamenetsky, 2007. Garlic: botany and horticulture. Hortic. Rev. 33, 123–172. Kevers, C., Falkowski, M., Tabart, J., Defraigne, J.O., Dommes, J., Pincemail, J., 2007. Evolution of antioxidant capacity during storage of selected fruits and vegetables. J. Agric. Food Chem. 55, 8596–8603. Kim, J.S., Kang, O.J., Gweon, O.C., 2013. Comparison of phenolic acids and flavonoids in black garlic at different thermal processing steps. J. Funct. Foods 5, 80–86. Lanzotti, V., 2006. The analysis of onion and garlic. J. Chromatogr. 1112, 3–22. Lee, H.S., Wicker, L., 1991. Anthocyanin pigments in the skin of lychee fruit. J. Food Sci. 56, 466–468. Leja, M., Mareczek, A., Starzynska, A., Rozek, S., 2001. Antioxidant ability of broccoli flower buds during short-term storage. Food Chem. 72, 219–222. Leja, M., Mareczek, A., Ben, J., 2003. Antioxidant properties of two apple cultivars during long-term storage. Food Chem. 80, 303–307. Michiels, J.A., Kevers, C., Pincemail, J., Defraigne, J.O., Dommes, J., 2012. Extraction conditions can greatly influence antioxidant capacity assays in plant food matrices. Food Chem. 130, 986–993. Montaño, A., Beato, V.M., Mansilla, F., Orgaz, F., 2011. Effect of genetic characteristics
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Contents lists available at ScienceDirect
Postharvest Biology and Technology journal homepage: www.elsevier.com/locate/postharvbio
Total polyphenols, total flavonoids, allicin and antioxidant capacities in garlic scape cultivars during controlled atmosphere storage
MARK
⁎
Zobia Naheeda,b, Zhihui Chenga, , Cuinan Wua, Yanbin Wena, Haiyan Dinga a b
College of Horticulture, Northwest A & F University, Yangling, Shaanxi 712100, China Agricultural Research Station, Baffa, Mansehra, Khyber Pakhtunkhwa, Pakistan
A R T I C L E I N F O
A B S T R A C T
Keywords: Garlic scape Quality Polyphenol Flavonoids Allicin Antioxidant
Five commercial cultivars of garlic scapes were subjected to a controlled atmosphere (O2 = 2-5%, CO2 = 3–6%) at temperature = 0 ± 0.5 °C, RH = 85-95%, for 140–224 d to document the quality and related changes in components during storage in two consecutive years. Polyphenols, flavonoids, allicin, and 2,2-diphenyl-1-picrylhydrazl (DPPH) as well as the ferric ion reducing antioxidant power (FRAP), metal chelating capacity (MCC), and hydroxyl radical scavenging activity (HRSC) were analyzed to study overall antioxidant properties of garlic scapes in storage. The storage life was 224, 196, 196, 168, and 140 days for G025, G107, G2011-04, G110, and G064. G025 had the highest total polyphenol concentrations at 140 and 168 days in 2014 and 2015, respectively, whereas G2011-4 had the lowest concentrations of total polyphenols. The highest total polyphenols, total flavonoids and allicin concentrations were observed in G025, whereas G2011-04 displayed the lowest concentrations of total polyphenols and allicin in both years. For all cultivars, total flavonoid concentrations decreased with time. The highest weight loss was observed in G064 both in 2014 and 2015. The antioxidant capacity of G025 and G110 was higher than that of the other cultivars. DPPH and HRSC were highest in G025, and MCC and FRAP were high in G110 and G107 in both years. These results demonstrate that cultivar influences the rate of garlic scape deterioration, chemical composition properties and antioxidant activities.
1. Introduction Garlic scape is the flower stalk of garlic plant. The hardneck type of garlic scape is thin and has a slight garlicky scent, a moderately spicy flavor and an especially fresh and pleasant taste. Garlic flower stalk is widely consumed in various parts of Asia, especially in China, as it is a common ingredient in Chinese cuisine (Simon and Jenderek, 2003). Garlic is a rich source of health-promoting phytochemicals including antioxidants such as phenolics, flavonoids, and allicin (Lanzotti, 2006). Garlic bulbs are good source of natural antioxidants and possess potential health-promoting effects due to their high phenolic phytochemical concentrations (Nuutila et al., 2003). Phenolic concentrations in garlic are affected by agronomic and environmental factors (Waterer and Schmitz, 1994), but cultivar is the primary factor that determine this variation. Bulb firmness, pH, total soluble solids, moisture concentration, and sugar concentration have been reported to differ across 14 garlic cultivars (Volk and Stern, 2009). Most research on garlic has focused on the bulbs, while scapes of garlic receive very little attention, which have gained popularity in recent years. Garlic scapes are used for strong desirable fresh flavor in various recipes and stir-fry preparations
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Corresponding author. E-mail address: [email protected] (Z. Cheng).
http://dx.doi.org/10.1016/j.postharvbio.2017.05.002 Received 16 February 2017; Received in revised form 2 May 2017; Accepted 2 May 2017 0925-5214/ © 2017 Published by Elsevier B.V.
as well as an additive for meat products (Rekowska and Skupien, 2009). Despite its popularity as a high-value vegetable and as a source of flavorful ingredient, very limited research on the antioxidant components of garlic scapes has been carried out (González et al., 2012). The quality of garlic scapes after long-term storage affects consumer acceptance. Many factors have to be considered for optimum quality, such as genotype and pre- and post-harvest conditions. Curing methods, environmental conditions and genotype can affect maximum quality, and post-harvest handling (storage temperature and relative humidity) is essential for maintaining high quality of Allium crops (Brewster, 2008; Gubb and MacTavish, 2002). Variation in allicin, allyl methyl thiosulfinate, and allyl trans-1propenyl thiosulfinate concentrations was observed in 93 garlic cultivars (Kamenetsky, 2007). Allicin (diallylthiosulfinate), an active compound in garlic, represents approximately 70% of the overall thiosulfinate concentrations formed upon crushing of cloves (Kim et al., 2013; Lanzotti, 2006). Garlic cloves contain a large amount of alliinase in sheath cells and alliin in storage cells; these compounds interact once garlic is damaged and alliin is chemically converted to alkenyl sulfinyl compounds (Ellmore and Feldberg, 1994). Allicin has antimicrobial,
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2.2. Determination of total polyphenols, total flavonoids, antioxidant capacity and weight loss
anti-inflammatory, antithrombotic, and anticancer properties (Beato et al., 2011; Bommareddy et al., 2016; Kim et al., 2013; Lanzotti, 2006). Garlic scape can be stored in controlled atmosphere for 10 months at a temperature of 0 ± 0.5 °C and a relative humidity of 85–95% to fill supply gaps in the market, but the consumers complain about the deterioration of quality compared with freshly harvested scapes (Pers.com.). With respect to the quality of green onion cultivars in a modified atmosphere, the total polyphenol concentration was reduced by 6.6% after 20 days of storage (Viškelis et al., 2012). The objective of this study was to investigate the deterioration of quality in different garlic scape cultivars during long-term storage.
50 g of scapes were subsampled from each replicate, homogenized and extracted in 200 mL of ethanol: acetone (7:3, v/v) for 1 h at 37 °C (Lee and Wicker, 1991). The extract was filtered through Whatman No. 41 paper and rinsed with 50 mL of ethanol: acetone (7:3, v/v). Extraction of the residue was repeated using the same procedure. The two combined filtrates were stored at −20 °C until total polyphenol, total flavonoid and antioxidant capacity analyses were carried out. Total polyphenols (TP) were determined using the Folin-Ciocalteu method (Jayaprakasha et al., 2001) with minor modifications. In a 10mL Eppendorf tube, 7.9 mL of distilled water, 0.1 mL of garlic scape extract and 0.5 mL of Folin-Ciocalteu reagent (1:1 with water) were added and mixed. After 1 min, 1.5 mL of sodium carbonate (1.8 mol L−1) was added. The prepared mixture sat at room temperature in the dark for 2 h. The absorbance was measured at 765 nm; the total polyphenol concentration was calculated from the calibration curve using gallic as the standard. All concentrations were expressed as grams per kilogram on a fresh weight basis. Total flavonoids (TF) were determined according to the methods of Yong et al. (2008). In a 10-mL Eppendorf tube, 0.3 mL of garlic scape extract, 3.4 mL of 30% ethanol, 0.15 mL of 0.5 mol L−1 NaNO2 and 0.15 mL of 0.3 mol L−1 AlCl3 were added and gently mixed. After 5 min, 1 mL of 1 mol L−1 NaOH was added. The absorbance was measured at 506 nm. The total flavonoid concentration was calculated from a calibration curve using rutin as the standard. All concentrations were expressed as grams per kilogram on a fresh weight basis. Weight loss of each replicate was measured every 28 d in 2014 and 56 d in 2015. Weight loss was calculated on the basis of differences between the initial and final weights recorded for each sample. Cumulative weight loss was expressed as the percentage loss of the initial total weight.
2. Materials and methods 2.1. Samples and storage conditions Garlic scape cultivars G110, G107, G2011-4, G025, and G064 were obtained from the germplasm program of the Horticultural experimental station of Northwest A & F University, Yangling, China. Garlic scapes were harvested during March and April in 2014 and 2015 respectively, at the commercial ripening stage with no disease and pest damage, without aging of the base and bracts. The scapes had fresh green color and crisp texture, and were harvested at the proper maturity stage (floral axis with bending hooks), without deformities. The garlic scapes were harvested on sunny days, transported to lab immediately in wellventilated conditions and precooled at 5 °C before being kept in cold storage. The base of the scape was trimmed, cleaned and cured in precooling room for 24 h. In 2014, data were recorded at 28 d intervals, whereas in 2015, data were recorded at 56-d intervals. The cultivars were stored at 0 ± 0.5 °C (RH = 85-95%, O2 = 2-5%, CO2 = 3-6%) for a maximum of 224 d, following the commercial storage recommendations (SBT 10887/2012). The fresh garlic scapes were stored at the eating-quality stage. Three boxes each of 360 L had two nozzles, one for injecting O2 and the other for CO2. After taking samples, the lid was closed, and nitrogen gas was slowly injected until the O2 reached the desired level through one nozzle. Then, the nozzle was quickly closed, and CO2 was slowly injected through the second nozzle, after which it was closed upon reaching the desired limit. A general analyzer CYCK401 (Yantai Venture of Measurement and Control Engineering limited, Xian, China) was used to measure temperature, relative humidity, O2 and CO2 concentrations. The same procedure was repeated every time in sampling. Commercial garlic scapes are usually stored in the cold storage room, and the boxes were used only for experiment. Three replicate samples of 1 kg each, were placed on the shelves in 3–4 layers, 300 mm apart. The boxes were kept open for ventilation 2–3 times per month for 15–20 mins each time. O2 and CO2 were maintained at 2–5% and 3–6%, respectively, to avoid sulfur-containing volatiles. Refrigeration time varied for different cultivars. The refrigeration time was 150–300 days. 150 days, the CO2 was kept at the high limit index and O2 at the low limit index; after 150 days, the CO2 was lowered to the low limit, and O2 was raised to the high limit index. Quality evaluations were performed on each sampling day. A panel of five experts subjectively assessed the samples for each quality characteristic (appearance, taste and texture) using a 1–5 scale. In the case of appearance, a scale composed of pictures and a brief description for each score value was used as follows: 5 = excellent, no defects; 4 = very good, minor defects; 3 = fair, moderate defects; 2 = poor; major defects; and 1 = inedible. A score of 3 was considered the limit of marketability and a score of 2 as the limit of edibility. Due to the small size of our taste test panel and the difficulty in tasting several garlic scape samples at one time, only generalizations from their records are presented.
2.3. Determination of allicin using HPLC For allicin analysis, we followed the procedure of Tan (2008). 10 g of scapes were subsampled from each replicate and crushed in 50 mL of methanol. The samples were then centrifuged at 4 °C at 12,000g for 20 min, after which 1 mL of supernatant was filtered through 0.45-μm filter paper. An allicin standard was obtained from Sigma International Co., China. For HPLC, we followed the procedure of Fujisawa et al. (2008), with some modifications. Quantitative analyses were carried out on an HPLC machine (Waters 600E, Milford, USA). The column dimensions were 150 × 4.6 mm, with a 5-μm thickness (C18 Aeris Peptide, Dikma Technologies, China) operating at a constant flow rate of 1.0 mL min−1 at 25 °C. An injection of 10 μL was used for the standard and for all samples. Detection was carried out using a UV detector at 220 nm. The mobile phase was as follows: methanol (9); water (41); acetonitrile (50). Quantification of allicin was performed by comparing the peak area produced by crushed garlic scape extracts with that of authentic allicin. The results of allicin were expressed as milligrams per kilogram on a fresh weight basis. 2.4. Measurements of antioxidant enzyme activity The ability to scavenge 2,2-diphenyl-1-picryl-hydrazl (DPPH) was determined using the method of Hatano et al. (1988). One milliliter of each extract was added to 1 mL of the DPPH radical solution in methanol (the final concentration of DPPH was 0.1318 mmol L−1). The mixture was then stirred vigorously and sat for 30 min; the absorbance of the resultant solution was measured at 517 nm with a spectrophotometer. DPPH was calculated as follows: Scavenging% = (1– Asample)/Acontrol) × 100 40
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cultivar G2011-4 (4.39–3.75 g kg−1). The experiment in 2015 showed the same trends.
Ferric ion reducing antioxidant power (FRAP) was measured as described by Prakash et al. (2007). First, 100 μL of the sample, 2.5 mL of phosphate buffer (0.2 mol L−1, pH 6.6), and 2.5 mL of potassium ferricyanide solution (1%) were sequentially mixed. The mixture was then incubated in a 50 °C water bath for 20 min before cooling. After cooling, 2.5 mL of 3-chloroacetic acid (10%) was added to the solution and mixed thoroughly. The 2.5-mL mixture was extracted into a new tube, and 2.5 mL of distilled water was added. Finally, 300 μL of FeCl3 (0.1%) was added to the mixture, and the reaction was allowed to proceed for 5 min at room temperature in a completely dark room. The absorbance of the product was measured at 700 nm, and FRAP was expressed as this absorbance. The hydroxyl radical scavenging activity (HRSC) was estimated following the methods of Prasad et al. (1996). Briefly, 3 mL of distilled water and 100 μL (0.02 mol L−1) of FeSO4 were added to a microfuge tube. Then, 45 μL of H2O2 (0.15% solution) and 1 mL (8 mmol L−1) of salicylic acid were added. The final volume of the reaction mixture was then added to 100 μL of the sample solution and kept in the dark for 50 min at 20 °C. The absorbance at 510 nm was recorded, and the HRSC was calculated as follows:
3.2. Total flavonoids TF in garlic scapes also varied among the cultivars (Fig. 1). Total flavonoids in the cultivars ranged from 0.927 to 0.357 g kg−1 in 2014 at 140 d (Fig. 1) and from 0.678 to 0.226 g kg−1 in 2015 at 168 d (Fig. 1). In 2014, the highest TF were observed in G025, ranging from 0.927 to 0.538 g kg−1 during 140 d of storage, whereas the lowest TF were observed in G107 (0.585 to 0.357 g kg−1) after the 140-d cold storage period. In 2015, TF were highest in G025 (0.678–0.237 g kg−1) and G107 (0.678 to 0.237 g kg−1) but lowest in G2011-4 (0.474–0.227 g kg−1). Our results in two consecutive years revealed that TF declined throughout the storage period. 3.3. Allicin concentrations In 2014, the highest allicin concentration was observed in G025, increasing from 2.80–13.75 mg kg −1 after 140 d of storage, whereas the lowest allicin concentration was observed in G107 (2.08–6.84 mg kg −1) after 140 d of storage (Fig. 1). In 2015 (Fig. 1), G025 again had the highest allicin concentration (from 3.05–8.01 mg kg−1). The lowest concentration was observed in G2011-4 (1.45–10.02 mg kg−1). A similar result of G025 was obtained in 2015, in which the allicin concentrations increased as the storage period increased.
Scavenging% = (1– Asample)/Acontrol) × 100 The metal chelating capacity (MCC) was measured using the method of Prakash et al. (2007), in which 100 μL of extract was mixed with 3.9 mL of distilled water, 100 μL (2 mmol L−1) of FeCl2, and 50 μL (5 mmol L−1) of ferrozine. The reaction mixture was kept in the dark for 10 min, and the Fe2+ concentration was monitored at 562 nm. The percentage of chelating capacity was expressed as follows:
3.4. Antioxidant capacities
Scavenging% = (1– Asample)/Acontrol) × 100
The highest FRAP (2.60 to 1.10) was observed in G107 and G064, whereas the lowest FRAP was observed in G110 (2.30 to 1.10) after 140 d (Fig. 2). DPPH quickly decreased in descending order; the highest DPPH was observed in G025 (20 to 8%). The MCC exhibited a slight decrease among days; the MCC before treatment was high in G110 (85 to 5%), and the lowest value was observed in G025 (39 to 20%) after 140 d. The HRSC exhibited a gradual decrease; the highest HRSC was noted in G025 (from 86 to 38%), whereas the lowest value was estimated in G110 (68 to 16%) at 140 d. The highest FRAP was observed in G110 at the beginning from 0.18 to 0.13, whereas the lowest was noticed in G2011-4 (0.16–0.11) and G025 (0.16–0.11) at 168 d (Fig. 2). The highest DPPH was observed in G025 (15 to 5%), whereas the lowest was observed in G064 (10 to 5%) at 168 d. The highest MCC was noted in G110 (44 to 11%), with the lowest value recorded in G2011-04 (32 to 11%) at 168 d. G025 showed the highest level of HRSC (25 to 9%), whereas G110 displayed the lowest level of HRSC (18 to 2%) at 168 d. The experiments in 2015 provided further evidence for the same result.
where Acontrol is the absorbance of the blank control (containing all reagents except the extract solution) and Asample is the absorbance of the test sample. 2.5. Statistical analysis This research was conducted with three replicates for each cultivar. Statistical analyses were carried out with the help of SPSS Statistics 17.0 software (SPSS Inc., Chicago, IL, USA). The data were subjected to one-way ANOVA, and the means were separated by Duncan’s multiple range tests. All tests were carried out in triplicate, and the results were presented as the means ± SE. Differences at P < 0.05 were considered significant. Correlations were estimated with the Pearson test at the P < 0.01 and P < 0.05 highly significant and significant levels, respectively. Correlation analysis was performed after 140 days in 2014 and after 168 days in 2015. 3. Results
3.5. Weight loss 3.1. Total polyphenols The highest weight loss was determined in G064 (0 to 10%) at 140 d. This loss was closely followed by G025 at 140 d, while the remaining cultivars showed the same value in 2014 (Fig. 3). The highest weight loss was observed in G064 (0 to 13%), and the rest of the cultivars showed the same values in 2015 (Fig. 3). A similar result was obtained in 2015 in which G064 showed the highest weight loss.
Qualitative evaluations were performed on garlic scape samples of five cultivars each after 28 days in 2014 and 56 days in 2015. The five tasting panel members agreed that G025 retained high-quality characteristics (appearance, taste, and texture) for 224 days. In 2014, G025, G107, G2011-04, G110 and G064 retained quality characteristics for 224, 196, 196, 168 and 140 days, respectively. TP varied among the different cultivars (Fig. 1). TP in different garlic scapes decreased from 3.98 to 2.22 g kg−1 at 140 d in 2014 (Fig. 1) and from 4.78 to 3.39 g kg−1 at 168 d in 2015 (Fig. 1). In 2014, the most extreme changes were recorded in cultivar G025, in which TP decreased during 140 d of storage from 3.98 to 2.59 g kg−1. The lowest TP were observed in cultivar G2011-4, decreasing from 2.91–2.22 g kg−1 during the 140 d storage period. In 2015, the highest TP was noted in cultivar G025 (4.78–3.80 g kg−1), whereas the lowest value was observed in
3.6. Correlation among total polyphenols, total flavonoids, allicin and antioxidant capacity Correlation analysis was used to explore the relationships among the different antioxidant variables measured for five garlic cultivars (Tables 1 and 2). Strong correlations were observed between TP and TF and among DPPH, FRAP, MCC, and HRSC in both years. TF were positively and highly correlated with TP. FRAP showed strong, positive 41
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Fig. 1. Total polyphenols, total flavonoids, allicin are represented in figure A, B, and C for the year 2014 while in figure D, E and F for the year 2015. Vertical bars represent the standard error of the means (n = 3).
ment with the results of Kevers et al. (2007), who concluded that polyphenol compounds in leeks decreased during initial stages of storage at 4 °C but stabilized afterwards. Petropoulos et al. (2016) reported that total phenol concentration decreased at 5 °C and 25 °C with 60–70% relative humidity in Greek garlic. In onion, flavonoid concentration increased to 64% when stored for 168–196 d in cold storage, with 58% increase noted after the first 84 d (Leja et al., 2003; Rodrigues et al., 2010). It was noted in most cases that the increase of total phenols concentrations during storage was accompanied by a decrease of anthocyanins. Our results are in agreement with those of Viškelis et al. (2012), who reported that total polyphenol concentrations of green onion after 20 days of modified atmosphere (10% CO2, temperature of 0 ± 0.5 °C and relative humidity of 95%) decreased by 6.6%. However, there was no significant difference observed in radical scavenging activity of green onion before and after the storage.
correlations with TP and TF. DPPH was highly correlated with TP, TF and FRAP. The MCC showed strong, positive correlations with TP, TF, FRAP and DPPH. Last, the HRSC was highly correlated with TP, TF, FRAP, DPPH and MCC. 4. Discussion The main quality parameters of garlic scapes that are considered to be necessary for human consumption are water content, nutrients and antioxidants. This study discussed the impact of a controlled atmosphere on the health-promoting compounds of garlic scapes, including total polyphenols, total flavonoids, antioxidant capacity, and allicin concentrations. The decreasing trend noted in total polyphenol compounds after postharvest processing of the garlic scape samples is in partial agree42
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Fig. 2. FRAP, DPPH, MCC, and HRSC are represented in figure G, H, I and J for the year 2014 while K, L, M and N for the year 2015. Vertical bars represent the standard error of the means (n = 3).
cultivar, and garlic ecotype on the concentrations of individual organosulfur compound exists. The quality parameter of weight loss is quite crucial, as this parameter has a strong impact on the appearance and shape of produce when it deteriorates or becomes disfigured due to water loss. In our study it was noted that significant weight loss occurred when garlic scapes were stored for 84 d; this weight loss may be due to the drying activity starting from the exterior surface and extending to the interior core area (Bloem et al., 2011). The metabolism process may also have a role in moisture loss in garlic cloves. The changes in water may be related to substance metabolism in the garlic scapes. In a comparative study in asparagus, it was noted that asparagus becomes unsalable for consumption when 8% weight loss occurs (Siomos, 2003). The weight loss increased progressively upon storage in asparagus and was mainly attributed to transpiration due to differences between the atmosphere
Flavor compounds that are important quality parameters are affected by many factors, such as storage temperature, cultivar, and storage duration (Bloem et al., 2011; Randle and Lancaster, 2002). Allicin concentrations were increased after 56 d of controlled atmosphere storage, the first time that this effect has been shown, although concentrations of sulfur compounds in other Allium species have been described by many researchers. Sukkaew and Tira-umphon (2012) reported increased allicin concentrations when garlic was stored at 4–6 °C with 80–90% relative humidity. Some management practices, such as cutting the basal portion during the storage period, had a negative influence on allicin concentration. In a study by Tsouvaltzis et al. (2007), removal of the basal portion of leeks showed a significant decrease of thiosulfinate concentrations during the storage period of 7 d at a temperature of 10 °C. Montaño et al. (2011) argued that a significant effect of location, 43
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Fig. 3. Weight loss is represented in figure O for the year 2014 while in figure P for the year 2015. Vertical bars represent the standard error of the means (n = 3).
and antioxidant capacity parameters and between each phenolic compound and antioxidant capacity measurement for 5 garlic scape cultivars in 2014 and 2015. The analyses showed highly significant correlations between total polyphenols and total flavonoids and among DPPH, FRAP, MCC and HRSC. There were also highly significant correlations among DPPH, FRAP, MCC and HRSC antioxidant assay measurements in 2014 and 2015. These findings indicated that the DPPH, FRAP, MCC and HRSC methods are stable and reliable for measuring antioxidant capacities of garlic scape cultivars.
and the asparagus surface Park et al. (1994). In this study, 4 in vitro assays (DPPH, HRSC, FRAP, and MCC) were used as complementary methods to evaluate the potential antioxidant activity of garlic cultivars. Significant differences were observed among different cultivars in these assays. The DPPH-scavenging activity of garlic scapes ranged from 3.41 to 20.81% in 2014 and from 3.41 to 15.10% in 2015 studies, exceeding the initial range (from 5.07 to 11.36%) in Allium species reported by Nencini et al. (2011). Queiroz et al. (2009) reported contrasting results: DPPH values were much higher in fried garlic than fresh garlic (21.68%). Decreased antioxidant capacity results is in agreement with the results of Kevers et al. (2007), who reported a decrease in antioxidant capacity measured with a DPPH assay in the white stem portion of leeks during storage for 23 d. Our results are also in line with those of Petropoulos et al. (2016), who reported that DPPH radical scavenging activity decreased significantly in three genotypes of Greek garlic when stored at 5 °C and 25 °C with 60 to 70% relative humidity for 196 d. Chen et al. (2013) reported contrasting results in which values of antioxidant capacities measured by DPPH, FRAP, MCC and HRSC methods were low compared to our results. In addition to the differences resulting from method use, the observed differences in the reported data can be explained by extraction procedure, cultivar and weather conditions of the production season (Michiels et al., 2012). The antioxidant activity was reduced by 29% after 6 weeks at 5 °C and by 36% when onion bulbs were stored in warm ambient conditions (Gennaro et al., 2002). The results and studies discussed above were conducted using shorter storage period in comparison to our studies; this shorter period may result in higher losses of antioxidant capacity. Contrasting results were reported by Leja et al. (2001), in which an increase in antioxidant capacity was observed in broccoli flower buds (Brassica oleracea var. italica cv. Lord) stored at 20 °C and at 5 °C for 3 and 7 d, respectively. Correlation analysis was performed among the phenolic compounds
5. Conclusion The variation in chemical composition in garlic scapes among different cultivars was investigated for the first time. G025 had the longest shelf life and the highest total polyphenol, flavonoid, and allicin concentrations, and DPPH and HRSC were observed with the lowest weight loss, along with MCC. Controlled atmosphere storage for 224 d did not negatively affect the antioxidant capacity and total phenolic concentrations. However, controlled atmosphere storage was associated with an increase in allicin concentrations. High amounts of natural antioxidants such as phenolic compounds in all cultivars makes garlic scape even more important for daily consumption. During controlled atmosphere storage, antioxidant properties were highly correlated with the presence of phenolic compounds.
Acknowledgments This research was supported by the National Natural Science Foundation of China (Project #31471865) and the China Scholarship Council.
Table 1 Correlation coefficients among antioxidant capacity (FRAP, DPPH, MCC, and HRSC), total polyphenols (TP), total flavonoids (TF) and allicin in garlic scapes (2014). Index
Total polyphenols
Allicin
Total flavonoids
FRAP
DPPH
MCC
HRSC
Total polyphenols Allicin Total flavonoids FRAP DPPH MCC HRSC
1 −0.574** 0.488** 0.509** 0.704** 0.520** 0.547**
1 −0.494** −0.719** −0.685** −0.687** −0.636**
1 0.611** 0.773** 0.449** 0.716**
1 0.721** 0.623** 0.687**
1 0.700** 0.809**
1 0.823**
1
** Correlation is significant at the 0.01 level.
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Table 2 Correlation coefficients among antioxidant capacity (FRAP, DPPH, MCC, and HRSC), total polyphenols (TP), total flavonoids (TF) and allicin in garlic scapes (2015). Index
Total polyphenols
Allicin
Total flavonoids
FRAP
DPPH
MCC
HRSC
Total polyphenols Allicin Total flavonoids FRAP DPPH MCC HRSC
1 −0.723** 0.539* 0.639** 0.691** 0.645** 0.655**
1 −0.743** −0.887** −0.773** −0.774** −0.730**
1 0.711** 0.784** 0.781** 0.640**
1 0.851** 0.867** 0.701**
1 0.919** 0.819**
1 0.766**
1
* Correlation is significant at the 0.05 level. ** Correlation is significant at the 0.01 level. and environmental factors on organosulfur compounds in garlic (Allium sativum L.) grown in Andalusia, Spain. J. Agric. Food Chem. 59, 1301–1307. Nencini, C., Menchiari, A., Franchi, G.G., Micheli, L., 2011. In vitro antioxidant activity of aged extracts of some italian allium species. Plant Foods Hum. Nutr. 66, 11–16. Nuutila, A.M., Puupponen-Pimiä, R., Aarni, M., Oksman Caldentey, K.M., 2003. Comparison of antioxidant activities of onion and garlic extracts by inhibition of lipid peroxidation and radical scavenging activity. Food Chem. 81, 485–493. Park, H.J., Chinnan, M.S., Shewfelt, R.L., 1994. Edible coating effects on storage life and quality of tomatoes. J. Food Sci. 59, 568–570. Petropoulos, S.A., Ntatsi, G., Fernandes, Â., Barros, L., Barreira, J.C.M., Ferreira, I.C.F.R., Antoniadis, V., 2016. Long-term storage effect on chemical composition, nutritional value and quality of Greek onion landrace Vatikiotiko. Food Chem. 201, 168–176. Prakash, D., Singh, B.N., Upadhyay, G., 2007. Antioxidant and free radical scavenging activities of phenols from onion (Allium cepa). Food Chem. 102, 1389–1393. Prasad, K., Laxdal, V.A., Yu, M., Raney, B.L., 1996. Evaluation of hydroxyl radicalscavenging property of garlic. Mol. Cell. Biochem. 154, 55–63. Queiroz, Y.S., Ishimoto, E.Y., Bastos, D.H.M., Sampaio, G.R., Torres, E.A.F.S., 2009. Garlic (Allium sativum L.) and ready-to-eat garlic products: in vitro antioxidant activity. Food Chem. 115, 371–374. Randle, W.M., Lancaster, J.E., 2002. Sulphur compounds in Alliums in relation to flavour quality. In: Brewster, J.L. (Ed.), Onions and Other Vegetable Alliums. Commonwealth Agricultural Bureau, International, Wallingford, pp. 329–356. Rekowska, E., Skupien, K., 2009. The influence of selected agronomic practices on the yield and chemical composition of winter garlic. Veg. Crops Res. Bull. 70, 173–182. Rodrigues, A.S., Pérez-Gregorio, M.R., García-Falcón, M.S., Simal-Gándara, J., Almeida, D.P.F., 2010. Effect of post-harvest practices on flavonoid content of red and white onion cultivars. Food Control. 21, 878–884. Simon, P.W., Jenderek, M.M., 2003. Flowering, seed production, and the genesis of garlic breeding. In: In: Janick, J. (Ed.), Plant Breeding Reviews, vol. 23. John Wiley and Sons. Oxford, UK, pp. 211–244. Siomos, A.S., 2003. Quality, handling and storage of white asparagus. In: Dris, R., Niskanen, R., Jain, S.M. (Eds.), Crop Management and Post Harvest Handling of Horticultural Crops. Science Publishers, Inc, Enfield, New Hampshire, USA, pp. 65–88. Sukkaew, P., Tira-umphon, A., 2012. Effects of storage conditions on allicin content in garlic (Allium sativum). Acta Hortic. 969, 209–212. Tan, Z.j., 2008. Determination of Allicin and Garlicin in garlic by HPLC. Gu Izhou Sci. 26, 76–79. Tsouvaltzis, P., Gerasopoulos, D., Siomos, A.S., 2007. Effects of base removal and heat treatment on visual and nutritional quality of minimally processed leeks. Postharvest Biol. Technol. 43, 158–164. Viškelis, Pranas, Bobinaitė, Ramunė, Lepse, Liga, Lepsis, Janis, Viškelis, Jonas, 2012. Quality changes of green onions stored in modifed atmosphere. Scientific works of the institute of Horticulture, lithuanian research centre for Agriculture and Forestry and aleksandras stulginskis university Sodininkystė ir daržininkystė 31, 1–2. Volk, G.M., Stern, D., 2009. Phenotypic characteristics of ten garlic cultivars grown at different North American locations. HortScience 445, 1238–1247. Waterer, D., Schmitz, D., 1994. Influence of variety and cultural practices on garlic yields in Saskatchewan. Can. J. Plant Sci. 743, 611–614. Yong, S.P., Soon, T.J., Seong, G.K., Buk, G.H., Patricia, A.A., Fernando, T., 2008. Antioxidants and proteins in ethylene-treated kiwifruits. Food Chem. 107, 640–648.
References Beato, V.M., Orgaz, F., Mansilla, F., Montaño, A., 2011. Changes in phenolic compounds in garlic (Allium sativum L.) owing to the cultivar and location of growth. Plant Foods Hum. Nutr. 66, 218–223. Bloem, E., Haneklaus, S., Schnug, E., 2011. Storage life of field-grown garlic bulbs (Allium sativum L.) as influenced by nitrogen and sulfur fertilization. J. Agric. Food Chem. 59, 4442–4447. Bommareddy, A., VanWert, A.L., McCune, D.F., Brozena, S.L., Witczak, Z., Singh, S.V., 2016. The role of organosulfur compounds derived from allium vegetables in cancer prevention and therapy. In: Ullah, F.M., Ahmad, A. (Eds.), Critical Dietary Factors in Cancer Chemoprevention. Springer International Publishing, Switzerland, pp. 111–152. Brewster, J.L., 2008. Onions and Other Vegetable Alliums, 2nd edition. Commonwealth Agricultural Bureau International, Wallingford, UK. Chen, S., Shen, X., Cheng, S., Li, P., Du, J., Chang, Y., 2013. Evaluation of garlic cultivars for polyphenolic content and antioxidant properties. PLoS One 8, 1–12. Ellmore, G.S., Feldberg, R.S., 1994. Alliin lyase localization in bundle sheaths of the garlic clove (Allium sativum). Am. J. Bot. 81, 89–94. Fujisawa, H., Suma, K., Origuchi, K., Kumagai, H., Seki, T., Ariga, T., 2008. Biological and chemical stability of garlic-derived allicin. J. Agric. Food Chem. 56, 4229–4235. Gennaro, L., Leonardi, C., Esposito, F., Salucci, M., Maiani, G., Quaglia, G., Fogliano, V., 2002. Flavonoid and carbohydrate contents in tropea red onions: effects of homelike peeling and storage. J. Agric. Food Chem. 50, 1904–1910. González, R.E., Sance, M., Soto, V.C., Galmarini, C.R., 2012. Garlic scape, an alternative food with human health benefits. Acta Hortic. 969, 233–237. Gubb, I.R., MacTavish, H.S., 2002. Onion pre- and post-harvest considerations. In: In: Rabinowitch, H.D., Currah, L. (Eds.), Allium Crop Science: Recent Advances 3. pp. 233–265. Hatano, T., Kagawa, H., Yasuhara, T., Okuda, T., 1988. Two new flavonoids and other constituents in licorice root—their relative astringency and radical scavenging effects. Chem. Pharm. Bull. 36, 2090–2097. Jayaprakasha, G.K., Singh, R.P., Sakariah, K.K., 2001. Antioxidant activity of grape seed (Vitis vinifera) extracts on peroxidation models in vitro. Food Chem. 73, 285–290. Kamenetsky, 2007. Garlic: botany and horticulture. Hortic. Rev. 33, 123–172. Kevers, C., Falkowski, M., Tabart, J., Defraigne, J.O., Dommes, J., Pincemail, J., 2007. Evolution of antioxidant capacity during storage of selected fruits and vegetables. J. Agric. Food Chem. 55, 8596–8603. Kim, J.S., Kang, O.J., Gweon, O.C., 2013. Comparison of phenolic acids and flavonoids in black garlic at different thermal processing steps. J. Funct. Foods 5, 80–86. Lanzotti, V., 2006. The analysis of onion and garlic. J. Chromatogr. 1112, 3–22. Lee, H.S., Wicker, L., 1991. Anthocyanin pigments in the skin of lychee fruit. J. Food Sci. 56, 466–468. Leja, M., Mareczek, A., Starzynska, A., Rozek, S., 2001. Antioxidant ability of broccoli flower buds during short-term storage. Food Chem. 72, 219–222. Leja, M., Mareczek, A., Ben, J., 2003. Antioxidant properties of two apple cultivars during long-term storage. Food Chem. 80, 303–307. Michiels, J.A., Kevers, C., Pincemail, J., Defraigne, J.O., Dommes, J., 2012. Extraction conditions can greatly influence antioxidant capacity assays in plant food matrices. Food Chem. 130, 986–993. Montaño, A., Beato, V.M., Mansilla, F., Orgaz, F., 2011. Effect of genetic characteristics
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