IJFS#699_bozza Ital. J. Food Sci., vol. 30, 2018 - 61 PAPER EFFECT OF GELATIN-BASED EDIBLE COATINGS INCORPORATED WITH ALOE VERA AND GREEN TEA EXTRACTS ON THE SHELF-LIFE OF FRESH-CUT APPLE S. AMIRI1,2, H.R. AKHAVAN3,4, N. ZARE2 and M. RADI*1,2 1Young Researchers and Elite Club, Yasooj Branch, Islamic Azad University, Yasooj, Iran 2Department of Food Science and Technology, Yasooj Branch, Islamic Azad University, Yasooj, Iran 3Department of Food Science and Technology, Faculty of Agriculture, Shahid Bahonar University of Kerman, Kerman, Iran 4Research and Technology Institute of Plant Production (RTIPP), Shahid Bahonar University of Kerman, Kerman, Iran *Corresponding author. Tel.: + 98 9177007984 E-mail address: s.amiri@iauyasooj.ac.ir ABSTRACT The objective of the present study was to evaluate the combined effect of edible coatings (gelatin, citric acid, ascorbic acid, and calcium chloride) incorporated with Aloe vera (50, 100, and 150%) and green tea (5, 10, and 15%) extracts on physicochemical, microbial, and sensorial properties of fresh-cut apples at 4ºC for 16 days. Significant differences in terms of quality parameters were observed between the control and coated apple slices. The highest variation in quality parameters was observed in the control, while the least variations were observed in coated slices with 150% Aloe vera. Also, the softening trend was slowed down by edible coatings. Furthermore, in basic coatings, the microbial growth inhibition was a function of the Aloe vera and green tea extract concentrations. Generally, the higher concentrations of Aloe vera and green tea extracts were found to maintain the quality parameters of apple slices for a longer time during the storage period. Keywords: apple slice, edible coating, Aloe vera, green tea extract Ital. J. Food Sci., vol. 30, 2018 - 62 1. INTRODUCTION In order to preserve freshness and to control spoilage and pathogenic bacteria growth, it is recommended to use edible coatings on fresh-cut fruit to extend their shelf life. For this purpose, natural polysaccharides, proteins, and antioxidants are used as raw materials for edible coatings and films (DHALL, 2013). Edible coatings can also be used as carriers of antimicrobials, antioxidants, anti-browning, flavouring, and colouring agents that improve the nutritional, sensorial, and microbiological properties of fresh-cut fruit (OMS-OLIU et al., 2010; VALENCIA-CHAMORRO et al., 2011). A dip treatment of fresh-cut fruit in organic acids (such as citric acid and ascorbic acids) and calcium salts as an alternative to sulphites were used to prevent enzymatic browning after fruits peeling and/or cutting (OMS-OLIU et al., 2010). Also, calcium treatments can maintain or improve the tissue firmness and crispness (OMS-OLIU et al., 2010). In this regards, edible coatings containing Aloe vera and green tea extracts are well documented in the literature. Aloe vera gel and gelatin have been used as edible coatings in fruit storage technology (ANDRADE et al., 2014; DANG et al., 2008). The barrier properties of Aloe gel coatings towards respiratory gases (CHAUHAN et al., 2011), as well as its antimicrobial functions (MARTÍNEZ- ROMERO et al., 2006) in coated fruit and fresh-cut fruit are reported. Besides, gelatin coatings show good barrier characteristics against oxygen and aroma transfers at low and intermediate relative humidity. However, gelatin has poor barrier properties against water vapour transfer due to its hydrophilic nature (ANDRADE et al., 2014). In recent years, the Aloe vera gel has been used as an edible coating for sweet cherries (MARTÍNEZ-ROMERO et al., 2006), mangoes (DANG et al., 2008), apples (CHAUHAN et al., 2011; SONG et al., 2013), papayas (MARPUDI et al., 2011), fresh-cut kiwifruit (BENÍTEZ et al., 2015; BENÍTEZ et al., 2013), and fresh-cut orange (RADI et al., 2017). Besides, the effect of Aloe vera coating, containing anti-browning solution, on apples slices (SONG et al., 2013) has been published in literature. Furthermore, tea (Camellia sinensis), is a good source of polyphenolic compounds, which have strong antioxidant properties. The high antioxidant capacity and overall antimicrobial activity of green tea have been attributed to catechins and their oxidized condensation products (MARTÍN-DIANA et al., 2008; MATAN et al., 2015). Coating with gelatin incorporated with green tea extract successfully retarded the microbial growth and therefore extended the shelf life of fresh-cut orange during cold storage (RADI et al., 2017). Such properties made us use green tea as our coating alternative. The aim of the present study was to investigate the combined effects of edible coatings containing gelatin, calcium chloride, ascorbic acid, and citric acid as well as various concentrations of Aloe vera and green tea extracts on physicochemical and microbial characteristics of fresh-cut apples during storage. 2. MATERIALS AND METHODS 2.1. Materials Gelatin, sodium hydroxide, calcium chloride, citric acid, ascorbic acid, and plate count agars (PCA) were purchased from Merck (Darmstadt, Germany). Aloe vera leaves and green tea were purchased from a local wholesale market (Yasooj, Iran). Red apples (Red Delicious) grown at Semirom Orchard (Isfahan, Iran) were freshly harvested at a commercially mature stage, sorted to eliminate the damaged ones, and selected for uniform size and colour. Ital. J. Food Sci., vol. 30, 2018 - 63 2.2. Preparation of the film-forming solutions for coating the apple slices Aloe vera extract was obtained from fresh Aloe vera leaves according to the method described by NAVARRO et al. (2011). The extract was used intact (for Aloe vera 100% treatment) or was diluted 50:50 with distilled water (for Aloe vera 50% treatment). Moreover, the Aloe vera gel was concentrated to 150% using a rotary evaporator (Heidolph, Germany) at 45°C. Moreover, green tea extract was prepared, based on the SIRIPATRAWAN and HARTE (2010) method. The total solid content (TSC) of tea extract was determined by the air oven method at 105°C. According to the TSC of tea, the final concentration of extracts was adjusted at 5, 10, and 15% TSC using a Rotary evaporator (Heidolph, Germany) at 45°C. Gelatin powder was dissolved in distilled water or concentrated-adjusted Aloe vera and tea extracts by stirring and heating to 50°C under nitrogen gas atmosphere to form 1% gelatin solution. The following coating solutions were assigned: (A) basic formula 1 (BF1): gelatin (1%), citric acid (0.1%), and calcium chloride (0.5%); (B) basic formula 2 (BF2): gelatin (1.0%), citric acid (0.1%), calcium chloride (0.5%), and ascorbic acid (0.5%); (C) the basic formula 1 and 2 with Aloe vera extract at three levels (50, 100, and 150%) that was abbreviated as BF1 or BF2+50, 100, and 150% Aloe; (D) the basic formula 1 and 2 with green tea extract at three levels (5, 10, and 15%) that was abbreviated as BF1 or BF2+5, 10, and 15% GT; and (E) coated with water which served as control. 2.3. Coating the apple slices Apples of uniform size and shape, and without any signs of mechanical damage, were selected, washed with chlorinated water (50 mg Cl2/kg H2O) and manually sliced in chilled water (5–6°C). Apple slices were dipped in the above-mentioned coating solutions for 1 min. and then drained for 30 min. The prepared apple slices were placed in polyethylene terephthalate (PET) clamshells (140 × 128 × 30 mm3) (Pars Plastic Khuzestan, Ahwaz, Iran), and stored at 4°C for 16 days. 2.4. Measurement of titratable acidity (TA) and total soluble solids (TSS) The apple slices were homogenized in a blender (Moulinex, Barcelona, Spain) and centrifuged at 2000 rpm for 1 min. to obtain a clear juice. The titratable acidity and total soluble solid of clear juice were measured (RADI et al., 2010). 2.5. Weight loss determination The weight loss in the samples was calculated as loss in weight of the apple slices in each container during storage and the values were reported on a percentage basis (RADI et al., 2010). 2.6. Firmness measurement The firmness of the apple slices was measured using a Texture Analyzer (CT3, Brookfield Engineering Laboratories, Stoughton, MA, USA) with a uniaxial penetration test. A stainless steel flat-end probe of 4 mm diameter was used to evaluate the firmness of the apple slices. The test conditions used for the measurement were pre-test speed 2mm/s; test speed 1 mm/s; post-test speed 10 mm/s; penetrating distance of 10 mm into the fruit, and a trigger force of 5 g (BENÍTEZ et al., 2013). Ital. J. Food Sci., vol. 30, 2018 - 64 2.7. Measurement of colour The surface colour of the samples was measured using a Hunter colorimeter (Colorflex, Virginia, USA). Hunter CIE L* for lightness, a* for redness, and b* for yellowness were determined (RADI et al., 2017). 2.7. Microbiological evaluation The microbiological analysis of the apple slices was carried out for standard plate counts in accordance with CHAUHAN et al. (2011) procedures. The results were expressed as log CFU/g of sample. 2.8. Sensory analysis Sensory evaluation was performed immediately after the apple slices were prepared at storage times of 0, 8, and 16 days. Twelve panellists were asked about the different quality attributes (colour, aroma and flavour, texture or firmness, and overall acceptance) of the apple slices using a scale with anchors at 0 and 5 as follows: colour, ranging from dark (0) to colour normal (5); aroma and flavour of apple, from weak (0) to strong (5), texture from soft (0) to hard (5). A final, overall preference test was also performed with a hedonic scale from dislike extremely (0) to like extremely (5). Scores from 2.5 to 5 were considered acceptable. 2.9. Statistical analysis All the experiments were run in triplicate. Statistics on a completely randomized design were performed with the analysis of variance (ANOVA) procedure in SAS (Release 9.1, SAS Institute Inc., Cary, NC) software and mean comparisons were carried out by Duncan’s multiple range test (p<0.05). 3. RESULTS AND DISCUSSIONS 3.1. Titratable acidity (TA) and total soluble solids (TSS) The effects of coating treatments on the TA and TSS parameters during cold storage are shown in Tables 1 and 2. The TA levels in the control and coated samples gradually decreased during the storage period, and the difference was significant in the control sample only on day 16. But, the decreasing trends of TA in coated samples were not significant during the storage period (data not shown for apple slices coated with BF1 and BF2 containing Aloe vera and green tea extracts). A further reduction of acidity in the control sample in comparison with trehalose/NaCl/sucrose-coated apple slices on the eighth day (ALBANESE et al., 2007) and gel-coated apple slices with cysteine, citric acid, ascorbic acid, and Aloe vera during the 16th day (SONG et al., 2013) were also reported. This phenomenon was linked to the malic acid decrease due to an increase in the respiration rate following peeling and cutting (ALBANESE et al., 2007). The higher acidity of coated apple slices could be attributed to the barrier properties of Aloe gel coatings towards respiratory gases (CHAUHAN et al., 2011; RADI et al., 2017). It seems that during storage, organic acids are used as substrates in respiration metabolism, thereby decreasing the TA and increasing the TSS (BENÍTEZ et al., 2013). Ital. J. Food Sci., vol. 30, 2018 - 65 Table 1. Titratable acidity changes in the control and coated apple slices with basic formulas (BF1 and BF2) during the 16 days of storage at 4°C. Treatment Storage time (day) 0 4 8 12 16 Control 0.37±0.02Aa* 0.35±0.03Aa 0.34±0.03Aa 0.35±0.03Aa 0.27±0.02Bb BF1 0.37±0.05Aa 0.36±0.02Aa 0.33±0.04Aa 0.31±0.01Aa 0.31±0.03Aa BF2 0.34±0.04Aa 0.38±0.05Aa 0.35±0.03Aa 0.35±0.03Aa 0.32±0.02Aa *Mean ± standard deviation (n = 3); Means followed by the different small letter within the same row or by the different capital letter within the same column are statistically different (p<0.05). Table 2. TSS changes in the control and coated apple slices with basic formulas (BF1 and BF2) incorporated Aloe vera and green tea extracts during the 16 days of storage at 4°C. Group Treatment Storage time (day) 0 4 8 12 16 1 Control 16.03±0.02Ae* 16.08±0.01Ad 16.18±0.02Ac 16.31±0.02Ab 16.36±0.02Aa BF1 16.01±0.02Ad 16.03±0.01Bd 16.08±0.02Bc 16.17±0.02Bb 16.22±0.02Ba BF2 16.02±0.01Ad 16.02±0.02Bd 16.08±0.01Bc 16.15±0.02Bb 16.23±0.02Ba 2 BF1 16.01±0.02Ad 16.03±0.01Ad 16.08±0.02Ac 16.17±0.02Ab 16.22±0.02Aa BF1+50% Aloe 16.01±0.01Ae 16.04±0.01Ad 16.07±0.02Ac 16.15±0.02Ab 16.23±0.02ABa BF1+100% Aloe 16.02±0.01Ad 16.04±0.01Ad 16.06±0.01Ac 16.13±0.01Bb 16.19±0.02BCa BF1+150% Aloe 16.02±0.02Ad 16.03±0.02Ad 16.05±0.01Ac 16.11±0.01Cb 16.17±0.02Ca 3 BF2 16.02±0.01Ad 16.02±0.02Ad 16.08±0.01ABc 16.15±0.02ABb 16.23±0.02Aa BF2+50% Aloe 16.02±0.01Ad 16.03±0.02Ad 16.09±0.02Ac 16.16±0.02Ab 16.20±0.01Aa BF2+100% Aloe 16.01±0.01Ad 16.03±0.01Ad 16.06±0.02Bc 16.13±0.01BCb 16.18±0.02Ba BF2+150% Aloe 16.01±0.01Ae 16.04±0.01Ad 16.05±0.01Bc 16.11±0.02Cb 16.15±0.01Ca 4 BF1 16.01±0.02Ad 16.03±0.01Ad 16.08±0.02Ac 16.17±0.02Ab 16.22±0.02Aa BF1+5% GT 15.99±0.02Ae 16.03±0.02Ad 16.08±0.02Ac 16.18±0.01Ab 16.21±0.01ABa BF1+10% GT 16.02±0.01Ad 16.04±0.01Ad 16.07±0.02ABc 16.16±0.02ABb 16.20±0.02ABa BF1+15% GT 16.00±0.05Ad 16.02±0.01Adc 16.05±0.01Bc 16.14±0.02Bb 16.19±0.01Aa 5 BF2 16.02±0.01Ad 16.02±0.02Ad 16.08±0.01ABc 16.15±0.02ABb 16.23±0.02Aa BF2+5% GT 15.99±0.02Ae 16.03±0.01Ad 16.08±0.01Ac 16.15±0.02Ab 16.20±0.01Ba BF2+10% GT 15.98±0.03Ad 16.03±0.02Ac 16.06±0.01Bc 16.13±0.02Ab 16.18±0.01Ca BF2+15% GT 15.99±0.03Ad 16.01±0.01Ad 16.05±0.02Bc 16.12±0.02Ab 16.18±0.01Ca *Mean ± standard deviation (n = 3); Means followed by the different small letter within the same row or by the different capital letter within the same column of each group are statistically different (p<0.05). The TSS of the control and coated samples significantly increased with storage time, while the coated samples showed a slight increase compared to the control sample (Table 2). In this regard, there was a significant difference between the control and the coated samples with the basic formulas (BF1 and BF2 without Aloe vera and green tea extracts) only after four days of storage. But, no difference was observed between the BF1 and BF2, which indicated the same effect of BF1 and BF2 treatments on apple slices during storage time. Increasing the concentration of Aloe vera and green tea extracts in the basic formulas (BF1 and BF2) increased the TSS significantly only after eight days of storage, and especially at Ital. J. Food Sci., vol. 30, 2018 - 66 the end of the storage period. Furthermore, no significant differences were found between BF1+Aloe and BF2+Aloe treatments in similar concentrations of the Aloe vera extract (50, 100, and 150%). Without considering the BF1 and BF2 coatings, samples coated with higher concentrations of Aloe vera and green tea extracts showed a lower increase in TSS at the end of the storage periods. The highest increase of TSS was observed in the control (~ 2.5%, TSS increased from 16.0 to 16.4 after 16 days of storage at 4°C), while the lowest increase was observed in samples coated with BF2+150% Aloe (~ 1.3%, TSS increased from 16.0 to 16.2). The findings of this study were similar to the results of AHMED et al. (2009), MARPUDI et al. (2011), and RADI et al. (2017), who reported that TA decreased and TSS increased with increasing storage time in nectarines, papaya and fresh-cut orange, respectively. During ripening, organic acids are used as substrates in respiration metabolism, thereby resulting in an increase in TA and decrease in TSS. In general, it seems that the total soluble solid content tends to increase over the storage period as a consequence of the ripening process (BENÍTEZ et al., 2013). A reduction in the respiration rate has been observed in sweet cherries (MARTÍNEZ-ROMERO et al., 2006) and kiwifruits (BENÍTEZ et al., 2013) coated with Aloe vera gel. Furthermore, softening occurs primarily because of an enzymatic degradation (pectin methylesterase and polygalacturonase) of the cell wall, which is mainly composed of cellulose, hemicelluloses, and pectins (OMS-OLIU et al., 2010). This may affect some physicochemical characteristics such as pH, TA, TSS, etc., of fresh-cut fruit. In this regard, the increasing trend of TSS in apple slices can be attributed to the softening and may, therefore, be associated with ripening (BENÍTEZ et al., 2013). 3.2. Weight loss The weight loss is mainly associated with moisture evaporation through the surface of fruit slices (OLIVAS et al., 2007). All samples demonstrated a gradual weight loss during storage (Table 3). The weight loss of uncoated fruit (25.10 %) was significantly greater than those of coated fruits during storage time. The weight loss of apple slices coated with BF1 and BF2 treatments was significantly lower than the control (p<0.05). In this regard, no significant difference was observed between BF1 and BF2 treatments until the eighth day, but the difference was significant on the 12th and the 16th days, and also the weight loss of BF1was significantly lower than BF2 at the end of the storage time. The least rate of weight loss (11.18 % and 11.52 %) was observed, respectively, in the samples coated with BF1+150% Aloe and BF2+150% Aloe treatments. Consequently, the weight loss of apple slices coated with BF+Aloe was significantly lower than other samples (p<0.05). Accordingly, the BF+Aloe coating was more effective than the BF+Green tea coatings. In the case of BF+15% GT weight loss was ~18.5%. Furthermore, weight loss of samples coated with both basic coatings (BF1 and BF2) containing 150% Aloe and 15% GT was significantly lower than of those coated with a lower level of extracts at the end of the storage periods. Similar results were obtained by SONG et al. (2013) and RDAI et al. (2017). These authors reported that the weight loss increased during storage, but the weight loss of the Aloe vera gel-coated apple slices was significantly (p<0.05) reduced compared to the control during storage. The binding of Aloe vera gel molecules to the surface of apple slices may have reduced the porosity of apple slices, resulting in lower water loss (SONG et al., 2013). It is reported that Aloe vera gel reduces the respiration rate, ethylene production, weight loss and, therefore, the softening of fresh-cut fruit textures (BENÍTEZ et al., 2013). Ital. J. Food Sci., vol. 30, 2018 - 67 Table 3. Weight loss changes in the control and coated apple slices with basic formulas (BF1 and BF2) incorporated Aloe vera and green tea extracts during the 16 days of storage at 4°C. Group Treatment Storage time (day) 0 4 8 12 16 1 Control 0.5±0.2Ae* 12.13±0.26Ad 19.68±0.14Ac 23.32±0.26Ab 25.10±0.37Aa BF1 0.3±0.1Ae 8.34±0.36Bd 12.19±0.29Bc 16.5±0.7Bb 18.41±0.19Ba BF2 0.3±0.2Ae 7.7±0.43 Bd 12.63±0.23Bc 15.59±0.26Cb 19.55±0.3Ca 2 BF1 0.3±0.1Ae 8.34±0.36Ad 12.19±0.29Ac 16.5±0.7Ab 18.41±0.19Aa BF1+50% Aloe 0.43±0.15Ae 7.9±0.58ABd 11.69±0.17Ac 14.90±0.28Bb 17.20±0.2Ba BF1+100% Aloe 0.23±0.15Ae 7.23±0.32Bd 10.3±0.35Bc 13.09±0.24Cb 15.41±0.2Ca BF1+150% Aloe 0.2±0.10Ae 6.2±0.2Cd 8.15±0.39Cc 9.50±0.22Db 11.18±0.19Da 3 BF2 0.3±0.2Ae 7.7±0.43Ad 12.63±0.23Ac 15.59±0.26Ab 19.55±0.3Aa BF2+50% Aloe 0.27±0.15Ae 7.78±0.14Ad 12.40±0.25Ac 14.59±0.27Bb 17.51±0.29Ba BF2+100% Aloe 0.23±0.15Ae 7.24±0.39Ad 10.69±0.17Bc 12.50±0.17Cb 15.51±0.34Ca BF2+150% Aloe 0.37±0.21Ae 5.99±0.32Bd 8.72±0.21Cc 10.20±0.17Db 11.52±0.23Da 4 BF1 0.3±0.1Ae 8.34±0.36BCd 12.19±29Bc 16.5±0.7Ab 18.41±0.19Aa BF1+5% GT 0.27±0.15Ae 8.10±0.14Cd 11.72±0.20Cc 14.87±0.21Bb 17.23±0.25Ba BF1+10% GT 0.37±0.15Ae 8.84±0.33ABd 11.81±0.25BCc 13.27±0.36Cb 17.50±0.32Ba BF1+15% GT 0.17±0.12Ae 9.2±0.36Ad 12.68±0.18Ac 14.70±0.23Bb 18.4±0.22Aa 5 BF2 0.3±0.2Ae 7.7±0.43Bd 12.63±0.23Bc 15.59±0.26Bb 19.55±0.3Aa BF2+5% GT 0.23±0.15Ae 10.09±0.29Ad 12.19±0.4Bc 14.56±0.21Cb 17.51±0.40Ca BF2+10% GT 0.17±0.12Ae 9.65±0.34Ad 12.69±0.18Bc 15.84±0.24ABb 18.11±0.21Ba BF2+15% GT 0.27±0.15Ae 9.81±0.28Ad 13.31±0.28Ac 16.09±0.24Ab 18.50±0.20Ba *Mean ± standard deviation (n = 3); Means followed by the different small letter within the same row or by the different capital letter within the same column of each group are statistically different (p<0.05). 3.3. Texture evaluation The texture degradation and softening trend continued through the storage time, but its rate was slowed down by the BF1 and BF2 coating compared to the control sample (Table 4). The maximum firmness was observed in BF1+150% Aloe (firmness decreased from 10.51 to 9.82 N after 16 days of storage at 4°C), while the least firmness was observed in control (firmness decreased from 10.44 to 5.62 N after 16 days of storage at 4°C). In addition, similar concentrations of Aloe vera (50, 100, and 150%) and green tea (5, 10, and 15%) extracts used in the BF1 and BF2 coatings had no significant effect on firmness, which indicated the same effects of BF coatings on apple slices during storage. Regardless of the BF1 and BF2 coatings, samples coated with the higher concentration of Aloe vera and green tea extracts had higher firmness. There was also no significant difference between coated samples with green tea and Aloe vera extracts. The lower firmness of the control sample than the coated samples was probably due to the growth of spoilage microorganisms in the sliced apple, but which was limited in coated samples due to the antimicrobial properties of Aloe vera and green tea extracts (BENÍTEZ et al., 2013; MATAN et al., 2015). Softening occurred primarily due to the enzymatic degradation (pectin methylesterase and polygalacturonase) of the cell wall. Calcium is reported to maintain firmness by cross-linking with pectins to form insoluble calcium pectates, which strengthen the structure of the cell wall (OMS-OLIU et al., 2010). In this Ital. J. Food Sci., vol. 30, 2018 - 68 regard, the fresh-cut apples, treated with calcium, showed no significant differences throughout the three weeks of storage (ALANDES et al., 2006). Table 4. Apple slices firmness changes (N) in the control and coated apple slices with basic formulas (BF1 and BF2) incorporated Aloe vera and green tea extracts during the 16 days of storage at 4°C. Group Treatment Storage time (day) 0 4 8 12 16 1 Control 10.44±0.25Aa* 8.70±0.13Cb 6.34±0.18Cc 6.02±0.08Bd 5.62±0.11Be BF1 10.48±0.24Ab 10.88±0.09Aa 9.68±0.20Ac 9.13±0.08Ad 8.87±0.06Ad BF2 10.24±0.08Aa 10.23±0.06Ba 9.33±0.11Bb 9.10±0.07Ac 8.86±0.11Ad 2 BF1 10.48±0.24Ab 10.88±0.09Aa 9.68±0.20Bc 9.13±0.08Cd 8.87±0.06Cd BF1+50% Aloe 10.43±0.09Aa 10.50±0.12Ba 9.91±0.08ABb 9.52±0.11Bc 8.98±0.08Cd BF1+100% Aloe 10.34±0.11Aa 10.26±0.06Ca 9.96±0.06Ab 9.73±0.09Ac 9.25±0.11Bd BF1+150% Aloe 10.51±0.13Aa 10.41±0.08BCa 10.05±0.10Ab 9.86±0.06Ac 9.82±0.10Ac 3 BF2 10.24±0.08Ca 10.23±0.06Ba 9.33±0.11Cb 9.10±0.07Cc 8.86±0.11Cd BF2+50% Aloe 10.35±0.13BCa 10.16±0.05Bb 9.87±0.08Bc 9.45±0.10Bd 9.09±0.14BCe BF2+100% Aloe 10.52±0.09ABa 10.27±0.09ABb 10.06±0.09Ab 9.71±0.17Ac 9.32±0.12ABd BF2+150% Aloe 10.55±0.08Aa 10.43±0.12Aa 10.04±0.07Ab 9.87±0.06Ab 9.47±0.14Ac 4 BF1 10.48±0.24Ab 10.88±0.09Aa 9.68±0.20Bc 9.13±0.08Cd 8.87±0.06Dd BF1+5% GT 10.27±0.08Aa 10.21±0.02Ba 9.99±0.04Ab 9.46±0.12Bc 9.05±0.10Cd BF1+10% GT 10.34±0.14Aa 10.22±0.04Bab 10.03±0.15Ab 9.63±0.12Bc 9.23±0.08Bd BF1+15% GT 10.47±0.03Aa 10.34±0.03Ba 10.13±0.06Ab 9.83±0.07Ac 9.50±0.07Ad 5 BF2 10.24±0.08Ba 10.23±0.06Aa 9.33±0.11Bb 9.10±0.07Cc 8.86±0.11Cd BF2+5% GT 10.33±0.09ABa 10.27±0.05Aa 9.93±0.07Ab 9.64±0.12Bc 9.34±0.07Bd BF2+10% GT 10.34±0.08ABa 10.23±0.08Aa 9.96±0.09Ab 9.85±0.10Ab 9.46±0.11ABc BF2+15% GT 10.44±0.06Aa 10.24±0.14Aab 10.08±0.14Ab 9.86±0.12Ac 9.53±0.10Ad *Mean ± standard deviation (n = 3); Means followed by the different small letter within the same row or by the different capital letter within the same column of each group are statistically different (p<0.05). The results of studies have indicated that Aloe vera reduces the respiration rate and ethylene production, weight loss, and softening (BENÍTEZ et al., 2013). In this regard, CHAUHAN et al. (2011) showed that Aloe gel coating alone or in combination with shellac, preserves the firmness in apple slices. Further, the Aloe vera edible coating application generally resulted in harder kiwifruit slices (BENÍTEZ et al., 2013). In addition to the antimicrobial effects, the improvement in mechanical properties of the films incorporating green tea extracts may be responsible for the interaction between polymeric matrix and polyphenolic compounds from green tea extracts (SIRIPATRAWAN and HARTE, 2010). 3.4. Colour change Colour is an important factor in the perception of the quality of fresh-cut fruit during their shelf-life. The colour indices (L*, a*, and b*) of apple slices stored at 4°C for 16 days were measured and only L* is reported in Table 5. Statistical analysis showed that the L*, a*, and Ital. J. Food Sci., vol. 30, 2018 - 69 b* colour indices significantly changed during storage. A significant increase in colorimetric a* and b* values, and a significant decrease in the L* value were observed in apple slices during storage time. The colour indices of apple slices showed a significant difference (p<0.05) between the uncoated and coated samples. Table 5. L* changes in the control and coated apple slices with basic formulas (BF1 and BF2) incorporated Aloe vera and green tea extracts during the 16 days of storage at 4°C. Group Treatment Storage time (day) 0 4 8 12 16 1 Control 76.00±2.00Aa* 72.33±0.58Ab 69.00±1.00Ac 66.33±1.53Cd 64.67±1.53Bd BF1 74.00±1.00Aa 71.66±0.58Ab 70.00±1.00Ac 68.66±0.58Bcd 68.00±1.00Ad BF2 75.66±1.15Aa 73.00±1.00Ab 71.00±1.00Abc 71.33±1.14Abc 70.00±1.00Ac 2 BF1 74.00±1.00Aa 71.66±0.58Ab 70.00±1.00Bc 68.66±0.58Acd 68.00±1.00Cd BF1+50% Aloe 74.33±1.53Aa 71.66±0.58Ab 70.33±0.56ABbc 68.66±0.55Acd 68.00±1.00Cd BF1+100% Aloe 74.66±1.53Aa 72.33±1.50Aab 71.33±0.57ABcb 70.33±1.00Aa 70.33±0.58Ba BF1+150% Aloe 74.67±0.56Aa 72.67±1.53Aab 72.00±1.00Acb 70.33±1.53Ac 70.67±0.55Ac 3 BF2 75.66±1.15Aa 73.00±1.00Ab 71.00±1.00Abc 71.33±1.14Abc 70.00±1.00Ac BF2+50% Aloe 75.33±1.50Aa 73.00±2.00Aab 70.67±1.15Acb 69.66±1.12Ac 69.00±1.00ABc BF2+100% Aloe 75.34±1.14Aa 73.33±1.50Ab 72.33±0.54Ab 70.33±0.59Ac 70.00±1.00ABc BF2+150%Aloe 76.67±1.55Aa 74.66±1.12Aa 72.00±1.00Ab 71.67.00±1.50Ab 71.33±1.10Bb 4 BF1 74.00±1.00Aa 71.66±0.58Bb 70.00±1.00Bc 68.66±0.58Ccd 68.00±1.00Bd BF1+5% GT 75.00±0.95Aa 72.00±1.00Bb 71.00±1.00ABb 69.00±1.00BCc 68.66±1.12ABc BF1+10% GT 75.33±1.49Aa 74.33±1.50Aa 71.67±1.45ABb 70.66±0.52ABb 69.33±1.50ABb BF1+15% GT 76.33±1.44Aa 74.67±1.10Aab 73.00±1.00Abc 71.66±1.50Acd 70.66±0.51Ad 5 BF2 75.66±1.15Ba 73.00±1.00Bb 71.00±1.00Abc 71.33±1.14ABbc 70.00±1.00Ac BF2+5% GT 75.00±1.00ABa 72.67±1.60Bb 71.00±1.05Acb 69.66±0.62Bc 69.33±0.60Ac BF2+10% GT 76.33±1.61ABa 75.33±1.55Aa 72.66±1.49Ab 71.00±1.00ABb 70.66±1.09Ab BF2+15% GT 77.66±1.55Aa 76.33±0.63Aa 73.34±1.60Ab 72.33±1.17Abc 71.00±1.00Ac *Mean ± standard deviation (n = 3); Means followed by the different small letter within the same row or by the different capital letter within the same column of each group are statistically different (p<0.05). The reduction trend of L* values in coated and uncoated samples occurred at different rates during the storage (p<0.05), showing a darkening tendency in the surface colour of the apple slices. The least reduction trend for L* was observed in the coated samples with both basic coatings (BF1 and BF2) containing 150% Aloe and 15% GT (~ 7.0%), in contrast to the control, which had the highest L* reduction (14.9%). The L* reduction during storage may be related to the occurrence of browning (MARTÍN-DIANA et al., 2008). The least increasing trend for a* and b* was observed in the coated samples with both basic coatings (BF1 and BF2) containing 150% Aloe and 15% GT in contrast to control, which had the highest a* and b* increasing (a* and b* respectively increased from -5.33 and 22.0 to 4.33 and 33.67 after 16 days of storage at 4°C). The variations of L*, a*, and b* in the coated samples were significantly low at the highest concentrations of Aloe vera and green tea extracts at the end of the storage periods, confirming the effect of these coatings in preventing the darkening and browning of apple slices. But at the same concentrations (50, 100, and 150%) of Aloe vera and green tea (5, 10, Ital. J. Food Sci., vol. 30, 2018 - 70 and 15%) extracts, there was no significant difference between BF1 and BF2 treatments in any of the measured colour parameters. Colour is a critical quality property of fresh-cut fruit, since the slicing of fruit may often lead to enzymatic browning by polyphenol oxidases and peroxidases, which react with phenolic compounds and cause surface browning (ALBANESE et al., 2007; OMS-OLIU et al., 2010). Furthermore, the oxidative degradation of ascorbic acid and non-enzymatic browning are reported to be a major deteriorative reactions occurring during storage (WIBOWO et al., 2015). Thus, anti-browning agents such as ascorbic acid, thiol-containing substances, carboxylic acids, and certain phenolic acids have been studied (OMS-OLIU et al., 2010). In this study, citric and ascorbic acids were used as anti-browning agents to inhibit enzymatic browning. Cut-surface colour of apple slices that had been treated with ascorbic acid (in BF2) was well maintained than BF1-coated samples, but this effect was not significant. An increase in the browning reactions in fresh-cut apples during storage was observed after increases in Hunter a* and b* values and a decrease in L* (PEREZ-GAGO et al., 2006; SONG et al., 2013). The findings of this study indicated that the variations of colour in Aloe-treated apple slices, especially at higher concentration (150%), were significantly lower than BF1- and BF2-coated apple slices. Similarly, CHAUHAN et al. (2011) reported that the L*, a*, and b* values of the Aloe vera gel-coated apple slices showed fewer changes compared to the control during storage for 30 days at 6°C, suggesting the anti-browning functionality of the Aloe vera coating. It is reported that the application of Aloe vera gel coating is an effective method for maintaining the colour of fresh-cut apple slices, as the Aloe vera gel coating can act as an oxygen barrier film, thus reducing enzymatic browning. However, the coating does not completely prevent oxidative browning (SONG et al., 2013). Therefore, to inhibit enzymatic browning, anti-browning agents were added to the Aloe vera gel coating solution (SONG et al., 2013). In this regard, SONG et al. (2013) reported that Aloe vera gel, containing 0.5% cysteine, was most effective in delaying the browning of apple slices during storage. The L* levels of coated apple slices increased with green tea extract concentrations. But the increase was not significant, indicating the anti-browning effects of green tea extract. Conversely, MARTÍN-DIANA et al. (2008) reported that an increase in green tea concentrations decreased the L* values of coated lettuce. 3.5. Microbial analysis Fresh-cut fruit is highly susceptible to pathogenic and spoilage microorganisms during the preparatory steps as a consequence of cross-contamination, the presence of a large area of cut surfaces, and juice and sugar leakage from damaged tissues (OMS-OLIU et al., 2010). The microbial growth on the surface of coated apple slices showed significant differences between the coated and uncoated samples (Table 6). A significant difference was found between the BF1- and BF2-coated samples after 12 days of storage. The total viable counts gradually and significantly increased with storage time in all treatments (Table 6). The microbial population of the control sample was higher than in the other treatments (2.63 and 6.78 log CFU/g at 0 and 16 days of storage, respectively), while samples coated with BF1+15% GT had the lowest microbial count compared to other treatments (2.42 and 3.17 log CFU/g at day 0 and 16 of storage period, respectively). The microbial counts in the Aloe vera- and green tea-coated samples did not exceeded 4.0 log CFU/g during storage time and were significantly lower than the coated samples with BF1 and BF2 (p<0.05). Therefore, the addition of Aloe vera and green tea extracts in BF1 and BF2 coatings reduced the microbial population significantly, and this effect was Ital. J. Food Sci., vol. 30, 2018 - 71 enhanced at higher concentrations of these extracts. In both the BF1 and BF2 coatings, the inhibition of microbial growth was a function of the Aloe vera and green tea extracts. Significant differences were also found between BF1 and BF2 treatments in similar concentrations of Aloe vera (100 and 150%) and also green tea extract (10% and 15%). The results showed that the antimicrobial effect of green tea was more than that of Aloe vera, especially in the higher concentrations of these extracts. Table 6. Microbial growth (log CFU/g) in the control and coated apple slices with basic formulas (BF1 and BF2) incorporated Aloe vera and green tea extracts during the 16 days of storage at 4ºC. Group Treatment Storage time (day) 0 4 8 12 16 1 Control 2.48±0.06Ae* 3.16±0.07Ad 5.25±0.13Ac 6.10±0.04Ab 6.78±0.04Aa BF1 2.63±0.09Ad 2.73±0.09Cd 3.22±0.05Bc 4.02±0.04Bb 4.94±0.07Ba BF2 2.52±0.11Ae 2.92±0.05Bd 3.17±0.06Bc 3.73±0.08Cb 4.65±0.11Ca 2 BF1 2.63±0.09Ad 2.73±0.09Ad 3.22±0.05Ac 4.02±0.04Ab 4.94±0.07Aa BF1+50% Aloe 2.47±0.06Bd 2.56±0.05Bd 3.03±0.08Bc 3.32±0.07Bb 3.91±0.09Aa BF1+100% Aloe 2.45±0.09Bd 2.56±0.07Bd 2.96±0.07Bc 3.22±0.01BCb 3.92±0.05Aa BF1+150% Aloe 2.43±0.05Be 2.56±0.08Bd 2.92±0.06Bc 3.17±0.08Cb 3.95±0.07Aa 3 BF2 2.52±0.11Ae 2.92±0.05Ad 3.17±0.06Ac 3.73±0.08Ab 4.65±0.11Aa BF2+50% Aloe 2.47±0.06Ae 2.62±0.06Bd 2.95±0.09Bc 3.51±0.09Bb 3.99±0.06Ba BF2+100% Aloe 2.49±0.06Ad 2.59±0.02Bd 2.86±0.11Bc 3.19±0.07Cb 3.65±0.10Ca BF2+150% Aloe 2.43±0.05Ad 2.54±0.05Bd 2.80±0.12Bc 3.14±0.11Cb 3.56±0.08Ca 4 BF1 2.63±0.09Ad 2.73±0.09Ad 3.22±0.05Ac 4.02±0.04Ab 4.94±0.07Aa BF1+5% GT 2.47±0.04ABd 2.57±0.04Bd 2.96±0.06Bc 3.29±0.09Bb 4.14±0.10Ba BF1+10% GT 2.43±0.09Bd 2.49±0.02BCcd 2.62±0.04Cc 3.08±0.13Cb 3.64±0.09Ca BF1+15% GT 2.42±0.12ABc 2.47±0.03Cc 2.55±0.06Cc 2.85±0.06Db 3.17±0.06Da 5 BF2 2.52±0.11Ae 2.92±0.05Ad 3.17±0.06Ac 3.73±0.08Ab 4.65±0.11Aa BF2+5% GT 2.51±0.05Ad 2.59±0.05Bd 2.89±0.08Bc 3.26±0.10Bb 4.10±0.08Ba BF2+10% GT 2.42±0.03Ad 2.46±0.06Cd 2.58±0.02Cc 3.11±0.09Bb 3.85±0.10Ca BF2+15% GT 2.48±0.03Ac 2.44±0.03Cc 2.52±0.03Cc 2.86±0.09Cb 3.30±0.07Da *Mean ± standard deviation (n = 3); Means followed by the different small letter within the same row or by the different capital letter within the same column of each group are statistically different (p<0.05). According to Table 6, lower microbial populations in samples coated with Aloe vera and green tea extracts can be attributed to antimicrobial properties of the coated compounds (BENÍTEZ et al., 2013; MATAN et al., 2015; RADI et al., 2017)). Aloe vera extract was reported to have antimicrobial functions, significantly reducing mesophilic bacteria, and especially showing antifungal activity (MARTÍNEZ-ROMERO et al., 2006; VALVERDE et al., 2005). Some individual components found in Aloe vera gel, such as saponins, acemannan, and anthraquinone derivatives, are known to have antibiotic activity and could be responsible for its antibacterial activity (VALVERDE et al., 2005). Green tea, too, is a rich source of polyphenols (mainly catechins and catechin derivatives) and the antimicrobial activity of green tea has been attributed to these compounds (MARTÍN- DIANA et al., 2008; MATAN et al., 2015). MATAN et al. (2015) and RADI et al. (2017) Ital. J. Food Sci., vol. 30, 2018 - 72 confirmed the antimicrobial activity in green tea extracts on fresh-cut dragon and fresh-cut orange, respectively. The reduction of microbial populations presented in this study was in good agreement with the antimicrobial effects of Aloe vera coating on table grape ((VALVERDE et al., 2005), sweet cherry (MARTÍNEZ-ROMERO et al., 2006), apple slices (CHAUHAN et al., 2011; SONG et al., 2013), kiwifruit slices (BENÍTEZ et al., 2015), raspberry fruit (HASSANPOUR, 2015), and fresh-cut orange (RADI et al., 2017) which reduced the aerobic bacteria, as well as yeast and mould counts during storage. 3.6. Sensory analysis The quality attribute scores (colour, aroma and flavour, texture or firmness, and overall acceptance) for the control and coated samples were studied on days 0, 8, and 16 (Fig. 1). Figure 1: Sensory attributes of apple slices coated with basic formulas (BF1 and BF2) incorporated with Aloe vera extracts during the 16 days of storage at 4ºC. The panellists gave greater sensory scores to coated slices than uncoated slices at the three stages of the experiment (Days 0, 8, and 16). Thus, the sensory analyses revealed the beneficial effects of coating in terms of delaying browning and maintaining the sensory quality of the apple slices. All edible coating treatments resulted in higher sensory scores than uncoated apple slices for all quality factors tested. But, except for colour, the other sensory characteristics were not significantly different in control and BF1 and BF2. In this Ital. J. Food Sci., vol. 30, 2018 - 73 regard, the colour score of the BF1 was significantly higher than those of BF2 and control samples. Although increasing the concentration of Aloe vera and green tea extract in the basic formulas led to higher sensory scores, no significant difference was observed between the apple slices coated with different concentrations of Aloe vera and green tea extracts. But, at the end of the storage, the panellists gave greater sensory scores (colour and overall acceptance) to BF2+150% Aloe-coated slices than the other treatments. Unexpectedly, Aloe gel-coated samples showed the lowest firmness at the end of storage even at high concentrations. The higher scores of coated slices compared to the uncoated ones were reported in the Aloe vera gel-coated apple slices (CHAUHAN et al., 2011; SONG et al., 2013) and Aloe-coated orange slices (RADI et al., 2017). 4. CONCLUSIONS In this study, an attempt was made to use Aloe vera and green tea extracts in gelatin-based coating to maintain the freshness of apple slices. Although gelatin-based coatings obtained higher quality attributes than those of control during storage time, the coatings did not completely prevent chemical and biochemical reactions. The addition of Aloe vera and green tea extracts in the gelatin-based coatings maintained the quality parameters of apple slices for a longer time during the storage period. In this regard, the least increasing trend for a* and b* was observed in samples coated with both gelatin-based coatings (BF1 and BF2) containing 150% Aloe vera and 15% green tea extracts. The samples coated with higher concentrations of Aloe vera and green tea extracts had lower increases in TSS at the end of storage periods. In terms of microbial count, the total count gradually and significantly increased with storage time in all treatments. The antimicrobial compounds Aloe vera and green tea extracts contributed to the lower microbial populations in samples coated with them. Slices coated with 150% Aloe vera and 15% green tea extracts obtained higher values for firmness. Moreover, the panellists gave greater sensory scores to coated apple slices than uncoated samples during the storage period. 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