Quality and shelf life of strawberry fruit as affected by edible coating by moringa leaf extract, aloe vera gel, oxalic acid, and ascorbic acid

ABSTRACT

In the current study, the most effective concentrations of moringa leaf extract (MLE), aloe vera gel (AV gel), ascorbic acid (1 mM) and oxalic acid (5 mM) were used. Physically mature fruits harvested from a commercial orchard were dipped in aqueous solutions of different edible coatings and kept at ambient conditions (25±2ºC & 55-60% RH) for evaluation of the quality and enzymatic parameters on daily basis till marketability. Current results revealed that 5 mM ascorbic acid treatment resulted in 57% reduced fruit weight loss; while 20% AV gel coating increased firmness by 60% over a 5-day ambient storage period. Application 6% MLE exhibited 50.7% and 49.7% reduced respiration rate and ethylene production respectively. Fruit biochemical quality parameters were also improved with the lowest accumulation of total soluble solids as a result of slow respiration, higher titratable acidity, ascorbic acid contents, carotenoids and anthocyanin contents in all treated fruit. While, total phenolic contents and total antioxidants along with superoxide dismutase, catalase, peroxidase, and anthocyanins were higher in ascorbic acid treated fruits. The activity of poly phenol oxidase enzyme was lowest in fruits treated with 5 mM ascorbic acid solution. Conclusively, the application of different edible coatings was proved to be beneficial as numerous fruit biochemical and antioxidative attributes were improved by ascorbic acid treatment; furthermore, MLE, AV gel and oxalic acid treatments were also helpful in maintaining fruit quality of strawberry fruits over a 5-days ambient storage period.

KEYWORDS:

• Fragaria × ananassa
• edible coating
• antioxidative enzymes
• biochemical fruit quality

Introduction

Strawberry (Fragaria × ananassa) is a false berry that belongs to Rosaceae family^[1] and is cultivated in tropical, subtropical, and temperate areas of the world. It is well known throughout the world because of its high nutritional value, good taste, and flavor,^[2] although North America and France are the origin of strawberries.^[3] Its cultivation area worldwide is 0.255 million ha with production of 7.7 million tonnes. In Pakistan, strawberry cultivation has expanded to Punjab (Gujrat, Lahore, Sheikupura, Muzzafargarh, Multan, and Lodhran districts), Sindh (Khairpur, Dadu, and Sukkur districts), and Balochistan. Strawberry is highly nutritious and refreshing, with increasingly high water contents, 8.4 g carbohydrates, and 59 g vitamin C.^[4]

Strawberry is a highly perishable fruit and, generally, its shelf life ranges from 3 to 4 days. High postharvest losses cause great economic losses to this nutritive fruit crop.^[5] Improper harvest and handling, higher loss of water, more susceptibility to fungal diseases and decay, physiological deterioration, and high respiration rate are the main causes of reduced shelf life and deterioration of fruit quality.^[6] The edible coating improves strawberry fruit quality and shelf life,^[7] by providing a semi-permeable barrier that decreases microbial attack, moisture and solute movement, gas exchange, and oxidative reaction rates.^[8,9] The application of edible coatings increases the esthetic appearance and shelf life of fruits by reducing the decay and physiological disorders and ripening processes.^[10,11] Aloe vera gel (AV gel), moringa leaf extract, oxalic acid, and ascorbic acid are naturally extracted and used commercially to improve the shelf life and maintain the postharvest fruit quality. AV gel potentially increases the shelf life of fruits by reducing respiration rate and moisture loss.^[12] It creates a protective layer that inhibits the action of micro-organisms such as fungi (Botrytis cinerea) and bacteria.^[7,13] Previously, AV gel reduced moisture loss, respiration, and softening process and maintained firmness, in fruits like sweet cherries,^[14] papayas,^[12,15] and table grapes.^[16] Whereas, moringa leaves are also used as an edible coating due to their antifungal properties; additionally, moringa leaf extract (MLE) is also a rich source of plant growth regulators such as cytokinins, abscisic acid, auxins, and antioxidants.^[17,18] The application of MLE is a cheap method for improving the quality and shelf life of fruits.^[19] Previous studies suggest that postharvest application of MLE delayed senescence and improved fruit quality attributes in Kinnow fruits.^[20] Furthermore, foliar spray of MLE (6%) has been reported to reduce weight loss, and maintain fruit quality (color and firmness) in plum fruit.^[21]

On the other hand, oxalic acid being an organic acid plays an important role in delaying senescence and has an anti-browning effect in apples, bananas, and litchi.^[22,23] Studies have shown that postharvest treatment with oxalic acid increased the shelf life of several fruits including mango and sweet cherry and reduced chilling injury occurrence in pomegranate during storage at 2°C.^[24–26] Likewise, ascorbic acid has been reported to control enzymatic browning in different fruits.^[27,28] Postharvest application of ascorbic acid effectively controlled fungus attacks in nuts and fruits and improved the physical characteristics of grapevine,^[29] while the combined application of ascorbic acid and AV gel improves postharvest life and maintains the quality of strawberry fruit.^[5] Moreover, postharvest ascorbic acid application has also been reported to reduce weight loss, increase fruit firmness and overall quality of plum fruit^[30]; furthermore, ascorbic acid application has also been reported to reduce peel browning and maintain fruit quality litchi fruit.^[28]

Scattered information on the impact of AV gel, MLE, ascorbic acid, and oxalic acid on fruit quality has been documented in the literature. However, from the above information, it is also evident that there is still a gap regarding the best suitable coating material. Therefore, this study aimed to explore the potential of AV gel, MLE, ascorbic acid, and oxalic acid, coatings on the quality and shelf life of strawberry cv. “Chandler.”

Materials and methods

Strawberry (Fragaria × ananassa Duch.) cv. “Chandler,” fruits uniform in size, shape, color, disease, and mechanical damage were harvested at 70% red color stage from the commercial farm located at Adda Band Bosan (300 22’ 24.0” N710 30’ 53.7” E), Multan. Harvested strawberry fruits were washed for removal of dust particles and shifted immediately to the Horticultural Laboratory at MNS-University of Agriculture Multan for the application of different coating treatments and storage.

Gel preparation

Aloe vera gel (AV gel) was prepared by using the technique outlined by Sogvar et al.^[5] with some modifications. Freshly detached AV leaves were washed with a chlorine solution of 0.03% (v/v) to extract transparent hydroparenchyma which was mixed thoroughly in a blender and further pasteurized at 65°C for 30 min, and then immediately cooled in ice to make 20% AV gel concentration.

Moringa leaf extract (MLE) preparation

Disease-free leaves of moringa were harvested from trees cultivated in the nursery of MNS- University of Agriculture Multan. Fresh moringa leaves were water extracted (10 kg fresh material per liter) as described by Yasmeen et al.^[31] which was further filtered through Whatman No. 1 filter paper followed by centrifuging at 8000 × g for 15 min.

Oxalic and ascorbic acid preparation

Ascorbic acid (5 mM) and oxalic acid (1 mM) were prepared by calculating the amount of susceptive chemicals using their molecular weight.

Coating and storage conditions

Harvested strawberries were dipped in chosen concentration from previous studies 6% MLE,^[21] 20% AV gel,^[32] 1 mM oxalic acid,^[33] and 5 mM ascorbic acid concentrations for 5 min using tween-20 as a surfactant, while control fruits were dipped in tap water. After dipping fruits were air dried packed in a plastic punnet and placed at ambient conditions (25 ± 2°C and 55–60% RH) in the Postharvest Laboratory MNS-University of Agriculture, Multan. All treatment samples were subjected to different physicochemical and shelf-life analyses daily till the marketable stage. Each replication consisted of 50 strawberry fruits, and various fruit quality parameters were estimated.

Fruit weight loss (%), fruit color variables (L*, a*, and b*), and fruit firmness (N)

The fruit weight loss of strawberry fruits placed at ambient temperature was determined by comparing initial weight by using an electronic balance (PA4102 OHAUS Corporation, USA) and expressed in percentage (%). The color of freshly harvested strawberry fruits was determined by using a handheld Chroma meter (KONICA MINOLOTA, CR-400, Tokyo, JAPAN) from the polar sides of 10 fruits at random from each replication. The values were L*, a*, and b* to classify the color space of three dimensions. The firmness of strawberry fruits subjected to different coating treatments was measured using a digital penetrometer (Fruit Hardness Tester, Lutron, and FR- 5120), equipped with a plunger of 8 mm and determined as Newton (N) as the unit of force.

Ethylene (µmol kg⁻¹ h⁻¹) and respiration rate (µmol kg⁻¹ h⁻¹)

To determine the ethylene production of strawberry fruit, five fruits per replication were placed in a sealed airtight plastic box for 1 h. An ethylene meter (model ICA-56, International Controlled Atmosphere Ltd., UK) was used to note ethylene production and also calculated as µmol kg⁻¹ h⁻¹. Respiration rate was observed using a CO₂ analyzer (Vaisala MI 70, Vaisala Inc., Helsinki, Finland) from the same jar used for determination of ethylene production and was calculated as µmol CO₂ kg⁻¹ h⁻¹.

Total soluble solids (°Brix), titratable acidity (TA %), and TSS:TA ratio

The total soluble solids (°Brix) of strawberry fruit were determined by putting one to two drops of juice on the prism of hand digital refractometer. Titratable acidity percentage (citric acid) was calculated by the protocol as explained by Ali et al.^[34] in which 10 ml freshly extracted juice added with phenolphthalein indicator was titrated against 0.1 N NaOH till the appearance of light pink color. The ripening index (TSS:TA ratio) was calculated by dividing total soluble solids (TSS) with the respective TA (titratable acidity) of strawberry fruits.

Ascorbic acid, total antioxidants (%), and total phenolic contents (GAE mg/100 g)

Ascorbic acid contents were measured using a sample of juice extracted from treated strawberry fruit as per the method given by Razzaq et al.^[35] The pH of strawberry juice was calculated by using the digital pH meter (Starter 3100 OHASU Corporation, USA). The total antioxidant contents of strawberry fruit were determined by following the procedure described by Razzaq et al.^[35] using the 1-diphenyl-2-picrylhydrazyl (DPPH) method. The 1 g strawberry pulp was homogenized in 5 ml extraction mixture of Methanol:Acetone: The supernatant (sample) was taken in a test tube having 5 ml of 0.004% DPPH, and absorbance was read at 520 nm on Epoch, Eliza Microplate Reader (Bio- Tek Instruments, Inc., Vermont, USA). Total phenolic contents (TPC) were expressed in milligram gallic acid equivalent per 100 g of fruit pulp and were estimated by adopting the Folin-Ciocalteu procedure adopted by Razzaq et al.^[35] The collected supernatant was reacted with 200 µL F-C reagent (10%) and 800 µL of Na2CO3 (700 mM) solution before reading the absorbance at 765 nm with Epoch Eliza Reader.

Anthocyanin (mg⁻¹ 100 g FW), carotenoid contents (µg/g F.W.), and malondialdehyde (MDA)

Anthocyanin contents were determined as mg-1 100 g FW by the method adopted by Razzaq et al.^[35] where 1 g frozen pulp was extracted with HCl-methanol and absorbance was read at 530, 620 and 650 nm. Carotenoid pigments from strawberry samples were determined µg/g FW by centrifuging the homogenized mixture at 9000 rpm and subsequently reading the absorbance at 662 nm, 645 nm, and 470 nm as determined by Razzaq et al.^[35]

Antioxidative enzymes activity

Strawberry fruit samples of strawberry pulp (1 g) were homogenized in a 2 mL phosphate buffer (pH 7–7.8) with pestle and mortar. Samples were centrifuged at 12,000 rpm at 4°C for 5 min. The supernatant was separated for use in antioxidative enzyme activity determination using Epoch, Eliza Microplate Reader (Bio-Tek Instruments, Inc., Vermont, USA).

Activities of superoxide dismutase (SOD), catalase (CAT) activity, peroxidase (POX) activity, and polyphenol oxidase (PPO) activity

SOD activity of strawberries was analyzed by estimating the 50% inhibition of the photochemical reduction of nitro blue tetrazolium (NBT). CAT activity (U/mg protein) was determined by mixing 100 μL enzyme extract with 100 μL of freshly prepared H2O2 (5.9 mM) for initiation of enzyme reaction at 240 nm absorbance.^[35] POX activity of strawberries was determined by adding (50 mM) 800 μL phosphate buffer (5pH) into (40 mM) 100 μL H₂O₂ and (20 mM) 100 μL guaiacol, mixed with 100 μL enzyme extract and absorbance was taken at 470 nm. PPO activity of strawberry enzyme extract was determined by reading an absorbance at 412 nm using an ELX800 Microplate Reader.^[36] PPO activity was expressed as U mg⁻¹ protein. MDA contents were expressed as nmol kg⁻¹ fresh weight from strawberry fruit pulp and was determined as per the method given by Zheng and Tian.^[37]

Statistical analysis

The experimental data were evaluated under a complete randomized design with two-factor factorial arrangements (coating treatments and days at shelf) and analyzed using the analysis of variance technique. The least significant difference (LSD) was performed 5% level of significance using a computer-based software Statistix®8.1.

Results

Fruit weight loss (%) and fruit firmness (N)

Weight loss of strawberry fruits increased during the shelf storage period regardless of treatment, with the maximum weight of strawberry fruits on Day 1 and the lowest weight recorded on Day 5 (Table 1). All treatments, i.e. MLE, AV gel, oxalic acid, and ascorbic acid, were able to reduce weight loss significantly compared to uncoated strawberry fruit during the storage at ambient period. Minimum fruit weight loss was recorded in strawberry fruits dipped in 1 mM oxalic acid and 5 mM ascorbic acid solution followed by AV gel and MLE. However, maximum fruit weight loss percentage was noted in control fruit. Likewise, firmness of strawberries cv. “Chandler” was decreased significantly throughout the shelf period regardless of treatments applied as the lowest firmness was recorded on Day 5. Application of naturally extracted MLE, AV gel, oxalic acid, and ascorbic acid treatments significantly reduced the loss in firmness of strawberry fruit during the whole shelf period. Strawberry fruits coated with 20% AV gel solution proved to be the best treatment with a 2.1 N firmness value compared to oxalic acid and ascorbic acid dipping that resulted in 1.75 N and 1.50 N on Day 5 of the shelf period. Significant loss in firmness was noted in control fruit followed by 6% MLE treatment on Day 5 (Table 1).

Table 1. Effect of edible coatings and shelf days on fruit color (L, a, b) of strawberry cv. “Chandler” at ambient conditions (25 ± 2°C and 55–60% RH).

Treatments	Shelf Period (Days)
	0	1	2	3	4	5	Means
	Fruit weight loss (%)
Control	0 o	13.26 m	28.16 jk	41.69 f-h	51.05 c	62.57 a	32.79 A
MLE 4%	0 o	8.47 n	23.72 l	32.48 i	43.38 ef	55.52 b	27.26 C
AV gel 20%	0 o	9.21 mn	24.52 kl	39.07 gh	43.04 efg	58.26 b	29.02 B
Oxalic acid 1 mM	0 o	5.86 n	21.87 l	30.26 ij	37.58 h	46.38 de	23.66 D
Ascorbic acid 5 mM	0 o	6.74 n	21.15 l	30.45 ij	38.27 h	49.27 cd	24.31 D
Means (T)	0.00 F	8.71 E	23.88 D	34.79 C	42.66 B	54.40 A
	Fruit firmness (N)
Control	5.99 a	5.07 c-e	3.77 g-i	2.78 j-l	2.14 lm	1.25 n	3.50 B
MLE 4%	5.99 a	4.43 e-g	3.96 f-h	3.59 hi	2.53 kl	1.39 n	3.65 B
AV gel 20%	5.99 a	5.55 a-c	4.47 ef	3.69 hi	3.24 ij	2.10 lm	4.17 A
Oxalic acid 1 mM	5.99 a	5.88 ab	5.22 b-d	3.71 hi	2.55 j-l	1.75 mn	4.18 A
Ascorbic acid 5 mM	5.99 a	4.72 de	4.57 d-f	3.14 i-k	2.18 lm	1.48 mn	3.68 B
Means (T)	5.99 A	5.13 B	4.40 C	3.38 D	2.53 E	1.59 F

LSD (P ≤ .05). For color fruit weight loss: T = 1.6798, SP = 1.8401, T×SP = 4.1147; for fruit firmness: T = 0.2835, SP = .3106, T×SP = .6945; Where T = Treatment and SP = Shelf Period.

Fruit color (L*, a*, and b*)

AV gel coating, MLE, oxalic acid, and ascorbic acid solutions significantly affected the change in strawberry color during the shelf-life period. The lightness value (L*) of strawberry fruits was decreased continuously from Day 0 to Day 5; however, all treated strawberries prolonged their freshness as maximum lightness value (L*) was recorded in strawberries treated with 6% MLE solution followed by strawberries that were coated with 20% AV gel, whereas fruit dipping in oxalic acid and ascorbic acid resulted in significantly lower lightness (Table 2). Chroma value a* deteriorated during the whole shelf period irrespective of treatments applied. However, chroma value a* was higher in untreated strawberries as compared to treated strawberries. Among the treated strawberries, naturally extracted 20% AV gel and 6% MLE solutions retained significantly low chroma a* value by retaining 6.78 and 7.5 chroma after 5 days of ambient period which was significantly lower than other treatments including control (Table 2). The color (b) describes the yellowness in the fruit. All the treatments exhibited a significant change in b* index from day 1 to day 5 with the highest b* index recorded in uncoated strawberry fruit by exhibiting 8.1 b*, while the lowest was recorded in 20% AV gel coated strawberry fruit with 5.9 b* index (Table 2).

Table 2. Effect of edible coatings and shelf days on fruit weight loss (A) and firmness (b) in strawberry cv. “Chandler” at ambient conditions (25 ± 2°C and 55–60% RH).

Treatments	Shelf Period (Days)
	0	1	2	3	4	5	Means
	L*
Control	27.473 a-c	23.072 e-i	20.345 i-k	17.805 kl	16.887 l	12.124 m	19.7 C
MLE 4%	27.473 a-c	28.136 ab	27.126 a-c	25.323 c-e	23.687 d-g	20.85 h-j	25.4 A
AV gel 20%	27.473 a-c	26.073 bcd	23.859 d-g	23.149 e-h	21.267 g-j	20.075 jk	25.7 B
Oxalic acid 1 mM	27.473 a-c	26.828 bc	25.355 c-e	22.983 e-i	21.799 g-j	15.271 l	23.3 B
Ascorbic acid 5 mM	27.473 a-c	29.606 a	24.763 c-f	22.083 f-j	21.173 g-j	15.959 l	23.5 B
Means (T)	27.473 A	26.743 A	24.289 B	22.269 C	20.963 D	16.856 E
	a*
Control	23.156 a	16.991 bc	16.5 b-d	14.933 d-f	15.109 c-f	10.95 j-l	16.272 A
MLE 4%	23.156 a	14.992 d-f	13.815 e-h	13.399 f-h	11.06 j-l	10.78 j-l	14.534 B
AV gel 20%	23.156 a	15.669 b-e	14.642 d-g	14.345 e-g	11.407 i-k	6.469 m	14.281 B
Oxalic acid 1 mM	23.156 a	17.389 b	14.427 e-g	13.293 f-i	12.187 h-j	9.553 kl	15.001 B
Ascorbic acid 5 mM	23.156 a	14.797 d-f	14.399 e-g	13.802 e-h	12.709 g-j	9.427 l	14.715 B
Means (T)	23.156 A	15.968 B	14.757 C	13.954 C	12.494 D	9.435 E
	b*
Control	15.729 a	11.573 b	9.568 c	8.772 c-f	8.667 d-f	7.797 g-j	10.30 A
MLE 4%	15.729 a	9.153 c-e	8.186 f-i	7.531 h-j	6.647 kl	5.915 lm	8.86 B
AV gel 20%	15.729 a	9.551 c	8.788 c-f	7.715 g-j	7.081 jk	6.247 l	9.18 B
Oxalic acid 1 mM	15.729 a	9.278 cd	8.305 f-g	8.246 f-i	7.476 ij	5.107 m	9.02 B
Ascorbic acid 5 mM	15.729 a	9.517 c	8.387 e-g	7.738 g-j	7.125 jk	6.179 l	9.11 B
Means (T)	15.729 A	9.814 B	8.647 C	8.000 D	7.399 E	6.249 F

LSD (P ≤ .05). For color L*: T = 1.1285, SP = 1.2362, T×SP = 2.7643; for color b*: T = 0.3362, SP = .3683, T×SP = .8236; for color a*: T = 0.7916, SP = .8672, T×SP = 1.9391; Where T = Treatment and SP = Shelf Period.

Ethylene (nmol C₂H₄/kg h) and respiration rate (mmol CO₂/kg h)

Although strawberries are non-climacteric and produce a limited amount of ethylene after harvest, postharvest application of natural edible coatings significantly influenced ethylene production and respiration rate of the strawberry fruit. Ethylene release in strawberry fruits was significantly affected by various edible coatings, shelf days, and their interaction at ambient conditions as the significantly highest amount of ethylene was produced from untreated fruits which was almost twofold higher than other treatments, i.e. 31.2 µmol kg⁻¹ h⁻¹. The lowest ethylene production obtained in treated fruit was about 15.5 µmol kg⁻¹ h⁻¹ with MLE (6%) which was statistically at par with AV gel, ascorbic acid, and oxalic acid treatments that exhibited 17.1, 17.3, and 17.1 µmol kg⁻¹ h⁻¹ ethylene, respectively, after 5-days of shelf period. However, minimum ethylene production was recorded in treated fruits with edible coatings as compared to control (Figure 1a). Likewise, the respiration rate was also significantly high with 28.283 µmol CO₂ kg⁻¹ h⁻¹ in untreated strawberry fruit throughout the shelf period, whereas almost twofold lowest respiration was noted in MLE treatment exhibiting about 14.3 µmol CO₂ kg⁻¹ h⁻¹ as other treatments exhibiting similar respirations throughout the shelf period (Figure 1b).

Figure 1. Impact of edible coatings (a), shelf days (b), and their interaction (c) on ethylene production (nmol C₂H₄/kg h) in strawberry cv. “Chandler” at ambient conditions (25 ± 2°C and 55–60% RH). Vertical bars Indicated ± SE of means, n = 15 replicates. Means not sharing same letters differ significantly from each other; P ≤ .05.

Total soluble solid (°Brix), titratable acidity (TA %), and TSS:TA ratio

A significant impact was recorded on °Brix of strawberry fruit treated with an organic acid, MLE, and AV gel coatings. An increasing trend was noted in the TSS of strawberry fruit with an increase in shelf period as the lowest TSS was recorded on Day 0 and the maximum was noted on Day 5 of the shelf period. All dipping and coating treatments resisted sharp increments in TSS of strawberry fruits until the end of the shelf period. The lowest TSS was recorded in treated fruit with 6% MLE and 1 mM oxalic acid treatments with 9.4 °Brix each which was at par with TSS contents of strawberries coated with 20% AV gel and 5 mM ascorbic acid treatments till the end of shelf period (Figure 2a). Likewise, the lowest titratable acidity, i.e. 0.153% TA, was recorded in untreated strawberry fruits compared to all treatments at the end of the shelf period. Higher TA percentage was recorded in strawberry fruits treated with 6% MLE and 1 mM oxalic acid treatments with 0.21% and 0.28% TA, respectively. Lowest TA percentage among treated fruits was recorded in strawberry fruit treated with AV gel and ascorbic acid showing about 0.19% and 0.18% TA, respectively, at the end of the storage period (Figure 2b). Similarly, the TSS:TA ratio was significantly higher in the control fruit with a 65.3 TSS:TA ratio followed by the MLE treatment that exhibited a 51.6 TSS:TA ratio, whereas all other treatments showed at par TSS:TA ratio (Figure 2c).

Figure 2. Impact of edible coatings (a) and shelf days (b) on total soluble solid contents (°Brix) in strawberry cv. “Chandler” at ambient conditions (25 ± 2°C and 55–60% RH). Vertical bars Indicated ± SE of means, n = 15 replicates. Means not sharing same letters differ significantly from each other; P ≤ .05.

Ascorbic acid contents, total phenolic contents (GAE mg/100 g), and total antioxidative activity (% inhibition)

The ascorbic acid contents, total phenolic contents, and total antioxidative activity of strawberry fruits treated/coated with different coating treatments varied significantly throughout the storage period at ambient temperature. An overall decreasing trend was observed in the concentration of ascorbic acid, total phenolic contents, and antioxidative activity of strawberries throughout the shelf period with maximum drop noted in control fruits. The concentration of ascorbic acid decreased significantly from Day 0 to Day 5; however, there was no significant difference among treatments on Day 5 for ascorbic acid contents (Figure 3a). Significantly higher TPC throughout the shelf period was recorded in strawberries dipped in 5 mM ascorbic acid solution followed by 6% MLE treatment. At the end of the shelf period, 5 mM ascorbic acid treatment exhibited 193.0 GAE mg/100 gm TPC which was 1.5-fold higher than the control fruit (Figure 3b). Significantly higher total antioxidants were noted in 5 mM ascorbic acid treated strawberry fruits throughout the shelf period followed by 6% MLE; whereas, the significantly lowest total antioxidative activity was noted in control fruits. At the end of the 5-day shelf period, 5 mM ascorbic acid treated with 55.9% antioxidative activity showed about 1.88-fold more antioxidants than untreated strawberry fruits (Figure 3c).

Figure 3. Impact of edible coatings (a), shelf days (b), and their interaction (c) on ascorbic acid contents in strawberry cv. “Chandler” at ambient conditions (25 ± 2°C and 55–60% RH). Vertical bars Indicated ± SE of means, n = 15 replicates. Means not sharing same letters differ significantly from each other; P ≤ .05.

Anthocyanins, carotenoid, and malondialdehyde

A significant increase in anthocyanin and carotenoid contents was recorded throughout the shelf storage period regardless of the treatment applied. All postharvest dipping treatments of strawberry fruits resulted in significantly higher anthocyanin contents compared to control fruits. However, anthocyanin contents of all treatments were at par with each other at the end of the 5-day shelf period with relatively higher anthocyanin contents (2.6 ∆Ag ⁻¹FW) observed in 5 mM ascorbic acid treated strawberry fruits (Figure 4a). Meanwhile, carotenoid showed a significant increase in control fruits with significantly higher carotenoid content, i.e. 3.1 Ug g⁻¹ β-carotene, at the end of the 5-day shelf storage period. Carotenoid contents in all treated/coated strawberry fruits showed a slow increment with a significantly lowest value (1.9 Ug g⁻¹ β-carotene) which was 0.63-fold less than control fruits at the end of the 5-day shelf period (Figure 4b). On the other hand, all postharvest dipping treatments of strawberry fruit significantly reduced peel leakage and maintained significantly reduced MDA through the shelf period, while control fruits showed significantly higher MDA (52.8 nmol g⁻¹FW). Among all treatments, 6% MLE-treated strawberry fruits exhibited the lowest leakage by exhibiting 36.25 nmol g⁻¹FW which was 0.68-fold lower than control fruits at the end of the 5-Day shelf storage period (Figure 4c).

Figure 4. Impact of edible coatings (a) and shelf days (b) on anthocyanin, carotenoid, and malondialdehyde contents in strawberry cv. “Chandler” at ambient conditions (25 ± 2°C and 55–60% RH). Vertical bars Indicated ± SE of means, n = 15 replicates. Means not sharing same letters differ significantly from each other; P ≤ .05.

Superoxide dismutase (U mg protein⁻¹), catalase (U mg protein⁻¹), peroxidase (U mg protein⁻¹), and polyphenol oxidase (U mg protein⁻¹)

A significant impact of edible coatings and shelf life was recorded on superoxide dismutase (SOD) and catalase (CAT) activities of strawberry fruit. Results revealed that the shelf period considerably influenced SOD and CAT activities with the lowest SOD activity recorded at 0-Day and which kept increasing gradually up to the end of the shelf period. Naturally extracted 6% MLE coating exhibited 117.14 U mg protein⁻¹ SOD which was 1.2-fold higher than uncoated strawberries followed by 1% oxalic acid and 5% ascorbic acid coatings which showed at par value for SOD activity after 5-days of shelf period (Figure 5a). Significantly higher CAT activity was recorded in strawberry fruits coated with 5 mM ascorbic acid coating which retained 1.69-fold higher CAT activity than uncoated strawberry fruits. Naturally extracted 6% MLE treatment exhibited the highest CAT activity up to the 4^th day of the shelf period which was decreased drastically on Day 5 of the shelf period (Figure 5b).

Figure 5. Impact of edible coatings (a), shelf days (b), and their interaction (c) on catalase in strawberry cv. “Chandler” at ambient conditions (25 ± 2°C and 55–60% RH). Vertical bars Indicated ± SE of means, n = 15 replicates. Means not sharing same letters differ significantly from each other; P ≤ .05.

Likewise, a gradual increase in POX and PPO activities was noticed throughout the shelf period with maximum enzymatic activity recorded on Day 5 in strawberry fruits regardless of the coating treatment applied. Application of 1% oxalic acid and 5% ascorbic acid maintained at par but significantly higher POX activity than naturally extracted 6% MLE and 20% AV gel coating treatments. Furthermore, 1% oxalic acid and 5% ascorbic acid exhibited about 1.68-fold higher POX activity than uncoated strawberry fruits (Figure 5c). Contradictory results were recorded for PPO activity with maximum PPO activity recorded in control/uncoated followed by 20% AV gel coating. The lowest PPO activity was recorded in strawberries treated with 5 mM ascorbic acids which exhibited 4.5 U mg protein⁻¹ followed by 1 mM oxalic acid treatment that exhibited 6.5 U mg protein⁻¹ PPO enzyme activity on Day 5 of the shelf period (Figure 5d).

Discussion

Fruit weight loss is considered a major factor in determining fruit quality and marketability. Fruit weight loss increased due to different metabolic processes including respiration and loss of transpiration.^[38] A continuous decline in weight loss was noted through the ambient storage duration, although strawberry fruit treated with naturally extracted coatings significantly checked the increase in fruit weight loss with the highest weight loss recorded in uncoated fruits. The lowest fruit weight loss was recorded in strawberry fruits treated with 1 mM oxalic and 5 mM ascorbic acid treatments than MLE or AV gel solutions (Table 1). Weight loss is principally caused by transpiration and loss of carbon reserves during respiration.^[39] Our results for the reduction of weight loss in oxalic acid-treated strawberry fruits are consistent with previous findings reported in lemon, pomegranate, and mango fruits.^[33,40,41] Firmness is an important indicator of strawberry quality that influences its marketability and acceptability. The reduction of firmness during the ripening process might be owed to the hydrolyzing of protopectin into pectin using enzyme activities and microbial infection. Our result revealed the highest firmness in AV gel-coated strawberry fruits (Table 1) which is in line with the findings of Martinez-Romero et al.,^[14] where AV gel reduced the activity of cell wall degrading enzymes, maintained firmness, reduced metabolic activity, and delayed ripening of strawberry fruit. Similar results were reported in strawberry fruit^[14,42] and in sweet cherry.^[43]

The visual appeal of a strawberry is thought to be directly related to its pleasant red color. In our study, hue angle was significantly influenced as all treatments maintained significantly higher L* values compared to uncoated strawberry fruits. A decreased L* index indicates that the strawberry color was changed to a darker color at ambient storage duration. On the other hand, all coating treatments were maintained at par L* values with AV gel exhibiting a slightly higher value at the end of the shelf period (Table 2). Our results are in agreement with the findings of Hosseinifarahi et al.,^[42] Nasrin et al.,^[44] and Martínez-Romero et al.^[14] where AV gel-coated strawberry fruits were lighter than uncoated fruits. The a* index is a good parameter for indicating ripening and expanding fruit red color.^[45] Strawberry fruits coated with naturally extracted AV gel exhibited significantly lower a* index followed by 6% MLE extract (Table 2). Therefore, the treatment with AV gel resulted in lower a* values, which showed that the ripening/senescence processes were retarded, as previously reported by Hosseinifarahi et al.^[42] in strawberry fruits. A similar observation has been recorded with other natural coatings like Arabic gum in “Grand nain” banana.^[46] All coating treatments maintained a significantly lower b* value than uncoated strawberry fruits, with AV gel-coated fruit exhibiting the lowest b* index through the ambient storage period (Table 2). Rapid color change in uncoated strawberry fruits could be due to an increased respiration rate that might have hastened the ripening/metabolic processes that were slowed down in coated/treated fruits. Our results for the b* index are consistent with the findings of Hosseinifarahi et al.^[42] who reported little change b* index in AV gel-coated strawberry fruit over the storage period. Respiration rate along with ethylene release indicates the longevity of acceptance. All coating treatments significantly inhibited ethylene release and respiration rate compared to untreated strawberries (Figure 1a,b). Our results for a gradual increase in respiration during storage and slow respiration due to MLE and other coating treatments are in line with the findings of Tesfay and Magwaza^[19] in avocado fruit. An increase in respiration rate can be associated with fruit senescence during ambient storage. Coating applications generally delay fruit ripening by modifying endogenous carbon dioxide, oxygen, and ethylene.^[26] Likewise, postharvest treatment of AV gel significantly delayed ethylene production and respiration rate of peach and plum fruits during ambient storage,^[47] and similar observations were reported in AV gel-coated cherry fruit over long-term cold storage conditions.^[48]

Total soluble solids (TSS) and titratable acidity determine the edible quality and overall acceptability of fruit. The current study showed a steady increase in TSS of all coated and uncoated strawberry fruit; however, all coating treatments maintained less increase in TSS with MLE and oxalic-treated strawberry fruits maintaining the lowest TSS (6.4) after 5 days of ambient storage period (Figure 2a). Strawberries coated with 6% MLE and 1 mM oxalic acid might have developed resistance to moisture loss, whereas a continuous increase in TSS can be associated with starch hydrolysis. Similar results were reported by Shafique et al.^[28] in litchi fruit, where the TSS increment rate was slow in ascorbic acid-treated litchi fruits; likewise, Sogvar et al.^[5] reported that TSS of strawberry ascorbic acid + AV gel coated cv. “Chandler” fruit was lower than the control fruit during the storage. In another study, sweet cherries subjected to oxalic acid treatment maintained lower TSS than control fruits during ambient storage.^[26] Titratable acidity (TA) of strawberry fruits showed an opposite trend with a continuous decrease during ambient storage. Strawberries that were coated with 6% MLE and 1 mM oxalic maintained higher TA compared to coating treatments. Moreover, the ripening index or TSS:TA was increasing during ambient storage which was significantly higher in uncoated fruits followed by 6% MLE and other coating treatments (Figures 2b, 2c). Oxidation of organic acid may have contributed to the depletion of TA which was inhibited due to the application of coating treatments. A similar observation was reported where litchi fruits treated with ascorbic acid and oxalic acid maintained higher TA during long-term low-temperature storage conditions^[23,28]; while, MLE treated peach fruit also maintained higher TA in peach fruit.^[49]

Meanwhile, a gradual decline in ascorbic acid contents was noted from day 0 to day-5 under an ambient storage environment; however, all treated strawberry fruits inhibited sharp decline and maintained significantly higher quantities than uncoated fruits (Figure 3). Ascorbic acid is the key factor in the determination of the freshness and quality of stored strawberries^[50] which is reduced over time due to continuous oxidative processes as a result of rising temperature.^[5] Among all coating treatments, strawberry fruits coated with 5 mM ascorbic acid solution retained significantly higher ascorbic acid, TPC, and total antioxidants through the ambient storage period. The decline in vitamin C during storage could be associated with respiration rate and other metabolic processes that change organic acid into sugars.^[51] The total phenolic content increase or decrease is considered to be dependent on postharvest conditions and practices that may affect certain biological processes in fruits.^[52,53] Ascorbic being itself an antioxidant might have helped in maintaining membrane integrity, deterred polyphenols, and oxidation of phenolic substrates.^[54] Previously similar results have been reported in litchi fruit where postharvest ascorbic acid application maintained higher ascorbic acid contents, TPC, and total antioxidants,^[28,55] while similar findings have also been reported in oxalic-acid-treated peach and sweet cherry fruits that exhibited higher total phenolic contents and antioxidants during long-term cold storage.^[26,56]

Our result showed a gradual increase in anthocyanin and carotenoid contents throughout the shelf storage period regardless of the treatment applied. Anthocyanin contents that are related to the surface color of strawberries were significantly higher in strawberries subjected to different coating treatments (Figure 4a). Fruit color is an excellent indicator of fruit quality and maturity.^[57] Low anthocyanin contents in untreated strawberry fruits could be the result of hastened fruit senescence and breakdown of pigments.^[58] Higher anthocyanin could be the result of higher antioxidative potential and increased enzymatic content in treated strawberries. Our results can be correlated with previous findings where moringa leaf extract-treated strawberries retained higher anthocyanin contents,^[21] while significantly higher anthocyanin contents in ascorbic acid treated lotus root slices have also been reported by Ali et al.^[59] Carotenoid pigments in strawberries were increased during a 5-day ambient storage period. Treated strawberries maintained less increase in carotenoid contents than control strawberry fruits (Figure 4b). All coated strawberries resisted a sharp increase in the carotenoid content which could be owed to delayed ripening and senescence processes. Our results are in line with the previous findings of Jodhani et al.^[60] who reported less increase in carotenoid contents of strawberry fruits coated with different plant gums treatments. MDA is considered to be a good indicator of oxidative stress which increases with lipid peroxidation of membranes. Results of the current study revealed that MDA was significantly lower in treated strawberries, particularly the lowest MDA was noted in strawberries treated with 6% MLE (Figure 4c). Decreased MDA in treated strawberries could be attributed to increased membrane integrity due to enhanced antioxidative potential as shown in our results. Previously chitosan coated guava, longan and strawberry fruits.^[61–63] Edible coatings develop a semipermeable layer that inhibits oxygen intake and alleviates oxidative damage in strawberry tissues,^[64] while ascorbic acid treatment overcomes oxidative stress by enhancing antioxidative potential.^[59] Likewise, MLE has also been reported to maintain membrane stability by inhibiting lipid peroxidation in cut roses,^[65] while MLE-coated avocado and mango fruits also exhibited reduced lipid peroxidation, indicating the potential of moringa leaves in alleviating oxidative damage.^[19,66]

Antioxidant enzymes have been discovered to play a major role in defending cells against the harmful effects of reactive oxygen species. Our results revealed an increasing trend in the activities of CAT, SOD, POX, and PPO during 5-day ambient storage with significantly higher activities of 5 mM ascorbic acid and other coatings treatments (Figure 5). These antioxidative enzymes retard the peroxidation of membrane lipids by scavenging ROS produced as a result of oxidative stress. The increase in SOD, CAT, and POX enzymatic activities illustrated the beneficial aspect of natural coatings are in agreement with the findings reported by Li et al.^[67] in strawberry fruits. The increase in activities of SOD, CAT, and POX could be attributed to protecting role of antioxidative enzymes which were produced in strawberries subjected to 5-days oxidative stress. Our findings for higher activities of antioxidative enzymes in coated strawberries are in line with the outcomes reported by Saleem et al.,^[63] where ascorbic acid combined with chitosan was applied on strawberries. Similar results were obtained where ascorbic acid combined with AV gel resulted in higher activities of CAT, SOD, and POX in fresh-cut lotus slices.^[59] Surface discoloration of strawberries is associated with higher activities of phenol-related metabolic enzymes, such as PPO.^[68,69] Current results revealed that strawberries subjected to different coating treatments maintained reduced activities of PPO enzyme which is in correspondence with a previous study in which gum Arabic-coated strawberries maintained the lowest PPO enzyme activity.^[70]

Conclusion

Our results revealed that all coating treatments significantly maintained strawberry quality during a 5-day ambient shelf period. Postharvest application of 5 mM ascorbic acid was more significant in reducing weight loss; while 20% AV gel coating presented significantly higher firmness than other coating treatments. Application 6% MLE exhibited the lowest respiration rate and ethylene production, respectively, whereas the lowest malondialdehyde (MDA) and TSS accumulation was recorded in 6% MLE-treated fruits. Among all treated fruits, titratable acidity, ascorbic acid contents, and carotenoid and anthocyanin contents were higher, while TPC and total antioxidants were higher in 5 mM ascorbic acid treatment. Activities of antioxidative enzymes, i.e. SOD, CAT, POX, and anthocyanins, were higher in ascorbic acid treated fruits with the lowest PPO enzyme activity recorded in 5 mM ascorbic acid treated strawberry fruits. Conclusively, the application of different edible coatings was proved to be beneficial as numerous fruit biochemical and antioxidative attributes were improved by 5 mM ascorbic acid treatment; furthermore, 6% MLE, 20% AV gel, and 1 mM oxalic acid treatments also maintained strawberry fruit cv. “Chandler” quality during a 5-day ambient storage period.

Disclosure statement

No potential conflict of interest was reported by the author(s).