Evaluating the application feasibility of thyme oil nanoemulsion coating for extending the shelf life of papaya (Carica papaya cv. Tainung No. 2) with postharvest physiology and quality parameters

Abstract

Papaya (Carica papaya L.) is a typical climacteric fruit with a brief shelf life due to the rapid degradation of quality during post-ripening, necessitating appropriate postharvest management to address this challenge. This study aimed to investigate the characteristics of thyme oil nanoemulsion (TO-NE) coating and utilize its benefits for preserving papaya. This study also investigated the physiological properties and quality changes of papaya storage at 20 ℃ and, in parallel, examined the effects of TO-NE coating to mitigate microbial infection of papaya during storage. The postharvest papaya was soaked in different concentrations (0.1, 0.25, and 0.5 mg/g) of TO-NE for coating. At the same time, the decay loss rate and effective shelf life were also evaluated. This study revealed that polygalacturonase (PG) and pectinesterase (PME) activities were inhibited during the storage of papaya treated with 0.25 mg/g TO-NE coated compared to the control group. This resulted in the preservation of the firmness of papaya fruits, in addition to a higher ascorbic acid content, delayed total soluble solids (TSS) accumulation, and total chlorophyll content (TCC) degradation, with a lagging color change for two days. The respiration rate and ethylene production were suppressed, while the 0.25 mg/g TO-NE coated group at day 14 (ethylene peak) were 63.2 mg CO₂ kg⁻¹ h⁻¹ and 7.3 µL kg⁻¹ h⁻¹, lower than control. The 0.25 mg/g TO-NE coating treatment significantly reduced the decay rate for 10 days of storage, preserving their appearance and facilitating ripening. This is a viable option for extending Tainung No.2 papaya shelf life.

Graphical Abstract

Introduction

Papaya is a tropical and subtropical fruit in the climacteric category, characterized by significant physiological changes during ripening [1]. In addition, it has high moisture content, limited shelf life, and susceptibility to airborne microorganisms, and is vulnerable to spoilage [1]. These risk factors might be compounded during storage, resulting in shortened shelf life or spoilage and severe economic losses [2, 3]. It will be imperative for the cultivation and distribution processes to incorporate postharvest technologies and solutions to mitigate the adverse effects of the ripening process through appropriate postharvest handling systems (technologies) and strategies [4]. Therefore, effective postharvest management is essential to maintain the papaya's nutritional and sensory attributes and extend its shelf life.

The coating techniques have been proven to extend the shelf life of papaya, even developed into edible films (produced with natural coatings, such as polysaccharides, proteins, or lipids (beeswax)) or non-toxic and biodegradable [1, 5,6,7,8,9,10]. These films serve to form a barrier between the fruit and the atmosphere, minimizing water loss, browning, gas exchange, ripening, and inhibiting microbial proliferation, thereby safeguarding fruits for an extended shelf life [5, 6, 8]. In addition, incorporating antimicrobial agents into edible coating films has developed into an emerging technology for protecting fruits from postharvest fungal hazards, which has been applied to bananas, strawberries, mangoes, papayas, or other fruits and vegetables with satisfactory performance [1, 7, 9, 11, 12]. Essential oils are antibacterial, natural substances extracted from plants with antibacterial and antioxidant properties and can slow down the volatilization after being added to the edible film [10]. Moreover, adding essential oils to the NE can improve its effectiveness by increasing the contact surface area against the products [13].

Thyme oil (TO; the principal components are thymol, polyapiophenol, carvacrol, etc.) is a secondary metabolite naturally present in plants [14, 15]. Its potent antimicrobial activity has made it a highly sought-after ingredient in biomedicine, pharmaceuticals, and cosmetics [1, 15]. Concretely, the phenolics in TO are the primary antimicrobial substances, which can enhance the fruit's defense-related enzymes and antioxidant capacity while affecting fungal proliferation and spore development by disrupting the cell membrane structure by blocking the activity of adenosine triphosphate (ATP) [16]. Recently, TO has attracted enormous interest as a new and effective potential candidate naturally available as an alternative to synthetic antimicrobial agents. Moreover, compared to conventional emulsions, NE is suitable for encapsulating bioactive compositions as the droplets can be controlled at a nano-size of around 10–100 nm, which provides rapid dispersion with high transparency and stability [17, 18]. In particular, TO-NE has been prepared and served as an essential oil film to prolong the shelf life of foods or fruits [1, 12,13,14, 19].

Therefore, this study was proposed to investigate the shelf life of papaya treated with TO-NE coating for storage at 20 ℃ of the papaya fruits, which analyzed the changes in physiological properties (including respiration rate, ethylene production, TCC, weight loss, and the activities of PG and PME) and quality-related parameters [containing firmness, TSS, titratable acid (TA), ascorbic acid, total plate count (TPC) on the peel, and color appearance] during the storage period (0–18 days). In addition, the effects of TO-NE coating on the mitigation of microbial infection of papaya were investigated, providing valuable information for the management of papaya during storage, shipping, and marketing.

Materials and methods

Materials

Papaya was purchased from a local farmer (Pingtung, Taiwan), while three batches of fruits were randomly harvested from the farm in Feb–Mar 2023 and ripened according to commercial harvesting standards. The fruits with 1–2 grooves of yellow color (10–20% color change) at around 150–180 days after anthesis. Following the harvesting process, green ripe papayas underwent commercial ripening for a duration of 1 day at room temperature (25 ± 2 ℃). Subsequently, the journey to the laboratory lasted for a duration of 30 min. Fruits that exhibit uniformity in color and size and are free from any signs of pest infestation, diseases, and abrasions are selected for subsequent trials. All the chemicals used in this study were purchased from Sigma-Aldrich^® (Merck KGaA, Darmstadt, Germany) and utilized directly without pre-treatment or purification. Difco^™ Nutrient Broth was purchased from Becton, Dickinson and Company Co. (Franklin Lakes, NJ, USA).

Preparation of nanoemulsion coating

The TO-NE coatings were prepared for use in this study according to the approaches Hou et al. [20] and Yu et al. [21] described, with slight modifications. Briefly, the aqueous phase (in the ratio of 2:1, v/v) was prepared using a homogenizer (HsiangTai Machinery Industry Co., Ltd., Taipei, Taiwan) with a speed of 2,000 rpm to homogenize the reverse osmosis (RO) water and 1,2-propanediol, while the oil phase consisted of thyme oil (10 mL) homogenized with Tween 80 (6 mL). Subsequently, the aqueous phase was slowly mixed with the oil phase at the addition rate of 10 mL/5 min with continued stirring (3000 rpm) for 50 min to form an oil-in-water NE. Then, 1 mg/g sodium alginate was added after mixing to obtain 10 mL/L TO-NE coating, followed by a dilution method (RO water as solvent) to prepare different concentrations of TO-NE coatings. In addition, the microscopic patterns of TO-NE at different concentrations were analyzed using a transmission electron microscope (TEM; HT-7500, Hitachi Ltd., Tokyo, Japan) at 100,000 × magnification.

Nanoemulsion coatings treated with papaya

All papayas were randomly grouped and soaked (2 min) in 0.1, 0.25, and 0.5 mg/g of TO-NE coating solutions, respectively, followed by picking up and air-drying before keeping them in polyvinyl chloride (PVC) boxes with sponge padding. In contrast, the control group underwent no treatment whatsoever. The micropattern of papaya peels subjected to diverse treatments was observed via a scanning electron microscope (SEM; HT S3000N, Hitachi Ltd., Tokyo, Japan) at 200 × magnification. All the groups were stored under refrigeration (20 ℃, 70–80% relative humidity), while the appearance, the rate of regulated respiration, the amount of ethylene production, and the rate of decay of papaya were measured every 2 days during the storage period, and their shelf-lives also were evaluated. In addition, papaya physiological and quality-related indicators were analyzed every 5 days (n = 3), with detailed measurements as described in the subsequent sections.

Physiological analysis

Respiration rate and ethylene production

The respiration rate and ethylene production of papaya were determined as described by Yu et al. [21], respectively, with slight modifications. Each group of papaya was placed in the 6 L breathing cylinders and sealed, where the gases were collected every 2 days with a 1 mL syringe at the port of the breathing cylinder. Subsequently, the samples were analyzed with a gas chromatograph (GC-8A, Shimadzu Co., Kyoto, Japan). Respiration rate analysis was performed using a thermal conductivity detector, a column of Porapak P (80/100, 2 m, 2 mm ID, GL Sciences Inc., Torrance, CA, USA), the temperature of the column of 90 ℃, and injection port temperature of 100 ℃, while respiration rate was expressed as mg CO₂ kg⁻¹ h⁻¹. Ethylene production was determined with the specific conditions: a flame ionization detector, the column was a Porapak Q (100/120, 2 m, 2 mm ID, GL Sciences Inc., Torrance, CA, USA), the column temperature was 80 ℃, the injection port temperature was 100 ℃, and the ethylene production was expressed as μL kg^–1 h^–1.

Polygalacturonase

The PG activity in papaya was measured as described in Tao and Pan [22], with minor modifications. The papaya pulp was homogeneously mixed with 95% ethanol in the 1:4 (v/v) ratio, which was extracted for 10 min at 4 ℃. Then, the above solution was centrifuged at 4 ℃ for 30 min (12,000 × g) using a centrifuge (CR-22, Hitachi Ltd., Tokyo, Japan). Next, the supernatant was taken, and another 10 mL of 80% ethanol was added, repeating the above steps once. The obtained supernatant was added to 5 mL sodium acetate buffer solution, followed by a standing reaction at 4 ℃ for 20 min. Afterward, the above reaction solution was taken at 0.5 mL by adding 1 mL of 50 mmol/L sodium acetate and 1.5 mL of 27.6 mM 3, 5-dinitrosalicylic acid, respectively, incubated at 37 ℃ for 1 h. Then, terminate the reaction with a boiling water bath at 100 ℃ for 5 min. After cooling, the absorbance value of the sample was determined by a spectrophotometer (U-2001, Hitachi Ltd., Tokyo, Japan) at a wavelength of 540 nm, and the PG activity was expressed as Unit (U)/kg fresh weight (FW).

Pectin methylesterase

The determination of PME activity in papaya as described by Ren et al. [23] with slight modifications. The extract was prepared by mixing 1 g of papaya pulp with 6 mL of 8.8 mg/g NaCl and extracted at 4 °C for 1 h. Afterward, centrifugation was used for 30 min (12,000 × g at 4 ℃), and the resulting supernatant was used to determine PME activity. The aforementioned supernatant, namely 0.15 mL, was incorporated into 0.7 mL of deionized water. Subsequently, 2 mL of 0.5 mg/g citrus pectin and 0.15 mL of 0.01 mg/g bromothymol blue were sequentially added, and the mixture was thoroughly mixed. Next, the reaction was allowed to stand for a duration of 30 min at 25 ℃. The absorbance values were determined at 620 nm wavelength, and PME activity was expressed as U/kg FW.

Total chlorophyll content

The determination of TCC of papaya rind in this study was based on an approach by Huang et al. [24] and Wellburn [25] with slight modifications. The extraction was carried out by taking 0.5 g of papaya rind, adding 10 mL of 99% acetone, and standing at 4 ℃ for 24 h, while the extract was protected from light. Subsequently, the extraction solution was centrifuged for 15 min (4 ℃, 12,000 × g), and the supernatant obtained was measured for the absorbance value at wavelength 652 nm. Finally, the mass concentration of total chlorophyll (ρt; Equation (Eq. 1)) was calculated by the following equation and then converted to TCC (Eq. 2).

$$ {\text{Mass concentration of total chlorophyll }}\left( {\rho {\text{t}}} \right) = {\text{Absorbance value at 652 nm}} \times 1000/34.5 \times 100 $$

(1)

$$ {\text{Total chlorophyll content }}\left( {{\raise0.7ex\hbox{${\mu g}$} \!\mathord{\left/ {\vphantom {{\mu g} g}}\right.\kern-0pt} \!\lower0.7ex\hbox{$g$}}} \right) = \left[ {{\raise0.7ex\hbox{${{\rho t} \times {\text{Sample total volume }}\left( {{\text{mL}}} \right)}$} \!\mathord{\left/ {\vphantom {{{\rho t} \times {\text{Sample total volume }}\left( {{\text{mL}}} \right)} {{\text{Sample weight }}\left( {\text{g}} \right) \times 1000}}}\right.\kern-0pt} \!\lower0.7ex\hbox{${{\text{Sample weight }}\left( {\text{g}} \right) \times 1000}$}}} \right] \times 1000. $$

(2)

Quality-related parameters

Peel color changes

The measurement of the color changes of papaya peel was performed as described in Hou et al. [26] and Nguyen, et al. [27] with minor modifications. L, a, and b values were measured at the middle portion on one side of each papaya fruit (the front side) using a colorimeter (Nippon Denshoku Industries Co., Ltd., Tokyo, Japan). Next, another measurement is made at 180° from the above measurement point (the back side), where the values of the front and back sides are averaged, thereby determining the color of the papaya's peel. The L-value (lightness, ranges from 0 to 100; the highest value means the brightest), a value (for red–green value, positive value means red, negative value means green), and b value (for yellow–blue value, positive value means yellow, negative value means blue).

In addition, the following formula (Eq. 3) was used to calculate the hue angle (θ value), which represents the color change of the fruit (0° for reddish-purple, 90° for yellow, 180° for blue–green, and 270° for blue), and chroma (a higher C value indicates a higher color intensity of the peel) (Eq. 4), respectively, in this study

$$ {\text{Hue angle }}\left( {\theta {\text{ value}}} \right) = \left| {{\raise0.7ex\hbox{$b$} \!\mathord{\left/ {\vphantom {b a}}\right.\kern-0pt} \!\lower0.7ex\hbox{$a$}}} \right|tan^{ - 1} $$

(3)

$$ {\text{Croma}} \left( {\text{C value}} \right) = \left( {a^{2} + b^{2} } \right)^{\frac{1}{2}} . $$

(4)

Firmness

This study determined papaya firmness as described in Cheng, et al. [28] with minor modifications. The measurement was performed using a texture analyzer (EZ–Test 500N, Shimadzu Co., Kyoto, Japan) under a No. 5 probe with a diameter of 0.5 cm, a probe depth of 10 mm, and a moving rate of 2.5 cm/15 s. Each whole fruit was measured at the equatorial position on both sides and averaged and expressed in Newton (N).

Total soluble solid

TSS was determined in papaya pulp juice as described by Wu et al. [29]. The TSS of papaya juice was determined using a refractometer (N–1E, Atago Co., Ltd., Tokyo, Japan) from 1 g of papaya pulp and expressed in °Brix.

Titratable acidity

The determination of TA content in papaya pulp was based on Wu et al. [29]. The pulp of 10 g of papaya fruit was added to 100 mL of RO water, homogenized in a homogenizer, and then filtered. Then, 25 mL of the supernatant was obtained and titrated to the end of titration (pH 8.1) using a titrator (DL53, Mettler Toledo, Columbus, OH, USA) with 0.1 N NaOH. The results were expressed as citric acid, and the content of its titratable acid was obtained.

Ascorbic acid

The ascorbic acid content of papaya pulp was determined according to the method described by Cheng et al. [28] and Nielsen [30]. The pulp of 5 g was homogenized with 50 mL of 3 mg/g metaphosphoric acid and then filtered. Next, 5 mL of the filtrate was added with 5 mL of metaphosphoric acid and titrated with indophenol until the solution turned pink. The above steps were repeated with 1 mg/mL of ascorbic acid standard. Finally, the ascorbic acid content of the sample was calculated using the following equation:

$$ {\text{Ascorbic acid }}\left( {\frac{{{\text{mg}}}}{{100{\text{g}}}}} \right) = \frac{{\text{S}}}{{\text{T}}} \times \frac{50}{{5{\text{ mL}}}} \times \frac{1}{{5{\text{ g}}}} \times 100, $$

(5)

where.

S indicates the volume of indophenol (mL) consumed in the titration of the sample, and T indicates the volume of indophenol (mL) used in the titration of the ascorbic acid standard.

Weight loss

Each group of samples was weighed with a digital weigher (UW4200H, Shimadzu, Co., Kyoto, Japan) following the operation’s completion in Sect. 2.3 above, which was the initial weight. Next, the weights were performed at intervals of 5 days during the storage period. Eventually, the WL was calculated by the following equation:

$$ {\text{Weight loss rate }}\left( {\text{\% }} \right) = \frac{{{\text{Weight with coating}} - {\text{Weight during storage}}}}{{\text{Weight with coating}}} \times 100. $$

(6)

Total plate count

TPC was determined according to the method described by Cheng et al. [31], with slight modifications. The papaya peels from different storage periods were collected in 1 cm² sizes and sterilized with tweezers, placed in centrifuge tubes containing 500 μL of sterile water, followed by homogenization on a sterile bench. Next, the stock solution was uniformly mixed by a one-touch vortex (FINEPCR^®, Gunpo-si, Gyeonggi-do, South Korea) vortex-induced oscillation, and serial dilution was performed for 10^–3–10^–6 dilutions. Then, 0.1 mL of the above dilutions were spread evenly on the plates of selective nutrition agar cultured plate, respectively, and were incubated (at 28 ℃ for 24–48 h), while the TPC with different dilutions was counted. In addition, plates with 30–300 colonies were considered valid calculation plates, while the stock solution’s TPC (CFU/mL) was obtained, and the TPC was expressed in Log CFU/g FW.

Decay loss rate and evaluated on shelf life

During the storage period, it was necessary to monitor the DL rate at regular intervals to ensure the papaya's preservation, while the fruit’s visual inspection was carried out every 2 days; inspection aimed to identify potential issues contributing to the loss of commercial value, such as mold, spoilage, rot, or pests and diseases. The DL rate was calculated by recording the quantity and using the following equation:

$${\text{Decay loss rate}} \%=\left(\frac{{\text{Number of damaged papaya}}}{{\text{Total papaya number}}}\right)\times 100.$$

(7)

Statistical analysis

The present study expresses all values as the mean ± standard deviation (SD) with three replications (n = 3) for all analyses except for Sect. 2.5.1. In addition, all data received were analyzed by the Statistical Analysis System (V9.0, SAS Institute, Cary, NC, USA) for analysis of variance (ANOVA) and Fisher’s least significant difference (LSD) for significant differences in the significance level of 0.05. Plotting was performed with SigmaPlot 10.0 (Systat Software GmbH, Erkrath, Germany).

Results

Thyme oil nanoemulsion microstructure and coated on papaya peel with microcosmic morphology

The particle sizes of the TO-NE droplets in this study were confirmed at 100,000 × magnification by TEM and ranged from 26.1 to 60.4 nm (Fig. 1A), exhibiting a spherical shape and distinctive particle size with uniform distribution. In addition, the results of SEM (200 × magnification) visualization of papaya with different concentrations of TO-NE in this study exhibited smoother epidermal microscopic patterns without gaps, and the epidermis formed a protective film that covered the stomata on the epidermis (Fig. 1B). Conversely, the control group showed an uneven epidermis with wrinkles and cracks, and the stomata were visible.

Fig. 1

Microstructure: A The size and shape of thyme oil nanoemulsion (TO-NE) droplets under a transmission electron microscope at 100,000 × magnification. The scale bar represents 100 nm; B the microscopic patterns (via the scanning electron microscope at 200 × magnification) of different concentrations of TO-NE coatings on the Tainung No. 2 papaya peels, Scale bar represents 200 μm

Effects of nanoemulsion coating on physiology in papaya harvesting

Changes in respiratory rate and ethylene production

This study revealed that the respiration rate of each treatment group decreased drastically during the initial storage period but showed an increasing trend as the storage time increased. In particular, the 0.5 mg/g TO-NE-coated treatment exhibited the most effective outcome in inhibiting the respiration rate (Fig. 2A). Conversely, the respiration rate of the control group remained continuously increasing until the 18th day of storage, where the respiration rate of 155.5 ± 10.9 mg CO₂ kg⁻¹ h⁻¹ appeared to be about two-to-threefold higher than that of the treatment groups.

Fig. 2

Effects of different concentrations of thyme oil nanoemulsion (TO-NE) coating on (A) respiration rate, B ethylene production, C polygalacturonase activity, and D pectin methylesterase activity of Tainung No. 2 papaya during storage (0–18 days) at 20 ℃. The symbols in the figure indicate ● (Control), ○ (0.10 mg/g TO-NE), ▼ (0.25 mg/g TO-NE), and △ (0.50 mg/g TO-NE), respectively. Lowercase letters represent significant differences (p < 0.05), while NS indicates insignificant differences

In ethylene production, there were only slight increases in the treatment groups at the initial storage period, followed by a constant rate of increase (Fig. 2B). However, the control group showed a sharp increase on the first day of storage and then started to decrease, followed by the rise on the 8th storage day, which remained constant until the 16–18th day of storage. Notably, all groups followed the same trend and showed the ethylene peak on storage day 14.

This study’s findings indicate that the control group exhibited the highest levels of ethylene production. The lowest levels were observed in the 0.5 mg/g TO-NE group, followed by 0.1 and 0.25 mg/g TO-NE groups, respectively. However, the inhibitory effect of the 0.5 mg/g TO-NE group on ethylene production was notably superior by the end of the storage period on the 18th day. In addition, observations suggest that applying 0.5 mg/g TO-NE coated may be a valuable strategy to mitigate ethylene production in papaya storage.

Changes in polygalacturonase and pectin methylesterase activities

Fruit firmness is a critical determinant of fruits' shelf life and economic value, while PG and PME activities are crucial factors determining fruit firmness. Understanding the relationship between PG and PME activities and fruit firmness is vital for developing effective postharvest handling and storage strategies. This study showed that all groups' PG and PME activities increased following fruit ripening (Figs. 2C and D). Moreover, there were no significant differences in PG and PME activities during the initial 10 storage days, whereas the control group recorded slightly higher readings than other treatments. On the 15 and 18th days of storage, the PG activities of the control group were 0.02 and 0.03 U/kg FW, respectively, which were significantly higher (p < 0.05) than TO-NE-coated ones. Regarding PME activities, the control group was the highest until the end of the storage period, considerably higher than the TO-NE-coated ones. Throughout the storage period, the control group exhibited the highest level of PME activity, surpassing that of the TO-NE-coated ones by a significant margin.

Changes in total chlorophyll content

The color change of fruits can be determined more accurately by the TCC in fruits. This study revealed a decreasing trend of TCC in papaya stored at 20 ℃ (Fig. 3A). Notably, on the 5th day of storage, while there was no significant difference between the groups, both control and 0.1 mg/g TO-NE groups exhibited a decrease in TCC values. However, constant TCC was maintained in all TO-NE groups from the 10th day of storage until the 15th day, which showed a drastic decrease. This was attributed to the benefits of the TO-NE coating treatment, which effectively prevented chlorophyll degradation in papaya, with 0.5 mg/g TO-NE providing the best results compared to the control group.

Fig. 3

Effects of different concentrations of thyme oil nanoemulsion (TO-NE) coating on (A) total chlorophyll content, B L-value, C Chroma (C value), and D Hue angle (θ value) of Tainung No. 2 papaya during storage (0–18 days) at 20 ℃. The symbols in the figure indicate ● (Control), ○ (0.10 mg/g TO-NE), ▼ (0.25 mg/g TO-NE), and △(0.50 mg/g TO-NE), respectively. Lowercase letters represent significant differences (p < 0.05), while NS indicates insignificant differences

Effects of nanoemulsion coating on postharvest quality of papaya

Changes in peel lightness, chroma, and hue angle

This study showed that the lightness of papaya peels increased as the storage time increased. The control group exhibited the highest L-value (63.60 ± 0.80) on the 10th day of storage, which was significantly higher than all TO-NE coated groups, yet there was no significant difference between the groups at the end of storage (15–18th days) (Fig. 3B). Similarly, chroma increased with storage time (Fig. 3C), which was significantly higher in the control group than the TO-NE-coated group during the entire storage period, with a maximum value of 66.5 ± 2.3 reached on the storage 10th day. Regarding the hue angle, which can indicate peel color, the Papaya hue angle decreased with storage time (Fig. 3D). The hue angle of the control group decreased relatively fast compared to the 0.25 and 0.5 mg/g TO-NE coated groups (p < 0.05) from storage days 5–15th. Despite the absence of any significant difference, it is noteworthy that the control group registered the lowest hue angle of 74.6 ± 1.0 on the 18th day of storage.

Changes in firmness, total soluble solids, titratable acid, and ascorbic acid contents

The present study showed a decreasing trend in hardness with increasing days of storage in all groups (Fig. 4A), with a more drastic decrease in the control group (p < 0.05). Subsequently, the firmness of all groups tended to flatten out by days 15–18 of storage. The firmness on the 18th day of storage ranged from low to high as follows: control (5.5 ± 0.4 N), 0.1 mg/g TO-NE (5.7 ± 0.4 N), 0.25 mg/g TO-NE (6.3 ± 0.4 N), and 0.5 mg/g TO-NE (7.4 ± 0.2 N), while 0.5 mg/g TO-NE was the highest, which were significantly different compared to other groups (p < 0.05).

Fig. 4

Effects of different concentrations of thyme oil nanoemulsion (TO-NE) coating on (A) firmness, B total soluble solid, C titratable acidity, and D ascorbic acid of Tainung No. 2 papaya during storage (0–18 days) at 20 ℃. The symbols in the figure indicate ● (Control), ○ (0.10 mg/g TO-NE), ▼ (0.25 mg/g TO-NE), and △ (0.50 mg/g TO-NE), respectively. Lowercase letters represent significant differences (p < 0.05), while NS indicates insignificant differences

TSS content is an indicator of the sweetness of fruits. This study showed an increasing trend in all groups during storage (Fig. 4B), particularly in the control group, where TSS content accumulated relatively rapidly and was significantly higher than in other groups at the storage period of 0–10 days (p < 0.05). However, there were no significant differences in the TSS content for each TO-NE coated group between the storage periods of 15–18 days.

In terms of TA, except for the control group, the TA contents of the groups coated with TO-NE exhibited a slightly increasing trend with the increase in storage time. Yet, each group showed no significant difference (Fig. 4C).

During the storage period, the ascorbic acid contents of the TO-NE-coated groups exhibited a slight increase from days 0 to 15, followed by a decrease on Day 18, which were significantly different (p < 0.05) for each group (Fig. 4D). Conversely, the ascorbic acid content in the control group decreased on day 10, followed by a slight increase, and eventually reduced again on day 18. This implies that the TO-NE-coated membrane contributed to the delay in the decline of ascorbic acid content on storage day 10.

Changes in weight loss rate

Following the harvest, crops undergo a period of water deprivation while respiration and transpiration processes continue to operate. Over time, the crops experience WL as a direct effect of the aforementioned water deficit, directly impacting their economic returns. This study showed that the WL rate of papaya for all groups was positively correlated as storage time increased. All the TO-NE coated groups exhibited slightly higher WL rates than the control group at the initial storage stage (0–5 days). In contrast, the 0.5 mg/g TO-NE coated group demonstrated the highest WL rate, while a significant difference was observed for all groups (p < 0.05). However, there was no significant difference in WL rates within each group during the post-storage period (10–18 days) (Fig. 5A).

Fig. 5

Effects of different concentrations of thyme oil nanoemulsion (TO-NE) coating on (A) weight loss, B total plate count, C decay loss, and D shelf life of Tainung No. 2 papaya during storage (0–18 days) at 20 ℃. The symbols in the figure indicate ● (Control), ○ (0.10 mg/g TO-NE), ▼ (0.25 mg/g TO-NE), and △ (0.50 mg/g TO-NE), respectively. Lowercase letters represent significant differences (p < 0.05), while NS indicates insignificant differences

Effects of nanoemulsion coatings on the antibacterial properties of papaya

This study showed that the TPC of all groups increased as storage time increased (Fig. 5B), where the highest TPC was observed in the control group. Notably, the 0.5 mg/g TO-NE group exhibited a significant antibacterial effect on the first storage day. Collectively, at later storage periods (days 10–18), all TO-NE groups (TPC between 6.7 ± 0.1 and 6.8 ± 0.0 Log CFU/g) exhibited a significant inhibitory effect (p < 0.05) compared to the control group (7.5 ± 0.0 Log CFU/g). However, the antimicrobial effect of different concentrations of TO-NE without exhibiting a dose dependence remained the lowest TPC for 0.5 mg/g TO-NE overlay.

Effects of nanoemulsion coatings on decay loss rate and shelf life of papaya

This study revealed a positive correlation between the decay loss rate of papaya and the storage period across all groups (Fig. 5C). Specifically, the control group achieved a decay rate of 35% on the 10th storage day, whereas the decay rate exceeded 50% on the 12th storage day, which was significantly higher than the 0.25 and 0.5 mg/g TO-NE groups (p < 0.05). However, the 0.1 mg/g TO-NE group exhibited no statistically significant difference despite having lower values than the control group. Therefore, the 0.25 and 0.5 mg/g TO-NE coated groups delayed papaya decay during storage. Decay was initiated after the 6th day of storage, and the decay loss rate reached 35–42% on the 12th day of storage, which was 2 days later than the control group. Unfortunately, all groups experienced a decay loss rate of over 80% at the end of the storage period (days 16–18th). The average shelf life of a control group of papaya was 11.3 days, while the shelf life of 0.1, 0.25, and 0.5 mg/g TO-NE coated papaya was 11.9, 13.2, and 13.6 days, respectively (Fig. 5 D). The 0.5 mg/g TO-NE coated was predicted to obtain maximum shelf life (p < 0.05).

Effects of nanoemulsion coatings on the appearance color of papaya

This study showed that during storage, the control group appeared to have a color change on storage day 4, followed by a complete yellowing on storage day 8 (Fig. 6). In contrast, the 0.1 and 0.25 mg/g TO-NE groups experienced color change on storage day 6, and the complete yellowing occurred on storage day 12. Notably, the 0.5 mg/g TO-NE group showed incomplete color change on the 18th day of storage. Moreover, the control group displayed black spots, mold, and decay on storage day 14, while the 0.1 and 0.25 mg/g TO-NE coated showed slight decay at the papaya’s tip, consistent with findings of decay loss rate in Sect. 3.5 above. The group that employed a 0.25 mg/g TO-NE coating demonstrated notable success in prolonging the shelf life of papayas. The findings of this study indicate that the TO-NE coating proved to be efficacious in reducing the respiration rate of papaya fruits.

Fig. 6

Effect of different concentrations of thyme oil nanoemulsion (TO-NE) coating on the appearance color of Tainung No. 2 papaya during storage (0–18 days) at 20 ℃

Discussion

This study showed that during the initial storage period, the lightness and chroma of the control group were significantly higher than all TO-NE groups. In contrast, the papaya rapidly changed from green to yellow (Fig. 6). However, the color angle of papaya coated with 0.25 and 0.5 mg/g TO-NE was significantly higher than that of the control group. This indicates that the TO-NE coating could effectively defer the ripening process and induce a gradual color change in the peel. The results of this study agreed with the slower color change of mango peel treated with chitosan coating results reported by Cosme Silva et al. [32]. In the course of fruit post-ripening, chlorophyll degradation occurs concurrently with the accumulation of carotenoids, which contributes to the color change of the fruit and is one of the characteristics of post-ripening [33]. This was also the critical point for defining the level of ripeness [1]. This study revealed that a concentration of 0.25 and 0.5 mg/g TO-NE coated papaya could mitigate chlorophyll degradation during storage, thereby facilitating slower color changes in the rind. However, a concentration of 0.5 mg/g TO-NE coated papaya may result in color change disorders in certain papaya fruits. According to a study, the slower color change observed in navel oranges may be attributed to the antioxidant properties of the essential oil concentration, which prevent the rapid oxidation of color-related substances [34]. Namely, a high concentration of essential oils might have the side effect of causing color change disorders [34]. In addition, high essential oil concentrations have been reported to have caused damage to the fruit, including severe peel burn and browning, thus affecting the fruit's appearance [34, 35]. This observed phenomenon may be attributed to high concentrations of essential oils, which cause structural alterations in the cellular components of phospholipids, fatty acids, and polysaccharides, leading to cellular toxicity and disrupting their normal functions [36, 37]. However, the decrease in fruit firmness observed during storage has been widely reported to be related to changes in the cell wall composition, particularly due to hydrolytic enzyme reactions in the fruit, including PG, PME, and cellulose enzymes [29, 38]. Moreover, PG is responsible for softening the fruit by breaking down the pectin, while PME promotes the loss of firmness by de-esterifying the galacturonic acid in the cell wall [29, 39], and the enzyme activities increase as the ripening level increases [40, 41]. This study agreed with the results reported by Yu et al. [42] on maintaining mango firmness with cinnamon essential oil and chitosan-coated films, where the primary key was regulating PME and PG activities in the fruits. During post-ripening, various physiological changes occurred, including hydrolyzing polysaccharides (starch or pectin) into monosaccharides and consuming organic acids during respiration [9, 28, 33]. This study showed that TO-NE-coated papaya had lower TSS content and higher ascorbic acid content, because the starch hydrolyzed slower into sugar when it was stored. This was similar to the results obtained with citrus essential oil NE-coated tomato as reported by Das et al. [43]. Maintaining the proper levels of ascorbic acid when storing agricultural commodities is crucial while facilitating the mitigation of oxidative reactions that can spoil the quality of the commodities and shorten their shelf life [44]. In addition, Radi et al. [45] reported that applying orange peel essential oil NE coating to citrus can maintain a higher ascorbic acid content. This was due to the coating’s exceptional fruit protection, which can provide a more effective barrier to oxygen contact, thereby reducing the oxidation of ascorbic acid. Furthermore, an additional explanation for the elevated ascorbic acid content could be attributed to the percentage increase in acidity and a decrease in the conversion rate of ascorbic acid to dehydroascorbic acid [46]. This study revealed that there were insignificant differences in the TA contents of the treatments. Nevertheless, it was observed that the papaya subjected to the TO-NE-coating treatments showed a slight tendency to increase TA contents. Notably, according to the report presented by Pérez-Soto et al. [47], organic acids serve as substrates in the respiration process that occurs during the ripening of fruits, thereby altering both pH and TA, then receiving the necessary energy [48]. Moreover, a study of carboxymethyl cellulose-coated films noted that bananas had higher TA content, negatively correlated with ethylene production and respiration rate [11]. However, it can be inferred that the primary reason for this phenomenon lies in inhibiting the post-ripening process and the consequent decrease in the metabolic consumption of organic acids [49]. Specifically, the fruit ripening process has been delayed by reducing the conversion of specific organic acids into sugars [50]. Therefore, the decrease in TA content of the control papaya in this study can be attributed to the more active physiological metabolism of the fruits and increased ethylene production and respiration rate, ultimately leading to the decrease in TA. The relationship between respiration rate, fruit ethylene production, and their senescence process is well established [38, 48, 51]. Ethylene production is a critical factor in the post-ripening of mature fruits. The rate of respiration, which reflects the energy consumption of fruits, is positively correlated with ethylene production [28]. Consequently, it has a significant impact on the shelf life of fruits. A higher respiration rate indicates faster energy consumption, which, in turn, results in more excellent ethylene production, leading to gradual fruit aging [51]. As a result, monitoring respiration rate and ethylene production is essential when considering the shelf life of fruits [52,53,54]. Li et al. [55] reported a significant increase in 1-aminocyclopropane-1-carboxylic acid (ACC) concentration in bananas as a result of a nanoemulsion coating, implying that the coating altered the internal gas composition of the fruit, which helped to limit the hydrolysis of ACC to ethylene, thereby reducing the accumulation of ethylene in the fruit. These results are consistent with the research reported by Salvia-Trujillo et al. [56], in which the lemongrass essential oil NE coating was found to form a thin film on apples. This film reduced the respiration rate by limiting the exposure of fruits to the surrounding atmosphere, thereby hindering the gas exchange process [48]. Moreover, TO-NE coating in this study showed lower ethylene production, which implied that TO-NE coating was effective in delaying fruit ripening and senescence. This was consistent with the findings of Nasrin et al. [57], who reported that lemon treatment with coconut oil and beeswax coating provided a reduction in ethylene production and extended the shelf life. This study reveals that TO-NE-coating treatment resulted in a substantial weight loss rate of papaya. The underlying hypothesis is that this weight loss can be attributed to the films’ enhanced water vapor permeability (WVP) rate, a consequence of TO essential oils, namely, the phenolic compounds present [58]. It is pertinent to note that the films are conventionally employed as a barrier between the food surface and the environment, thereby mitigating water evaporation from food surfaces and extending the shelf life of the food [59]. Notably, coatings might display varying degrees of penetration due to irregular structures and thicknesses forming during the membrane consolidation process [50]. Typically, incorporating essential oils into the coating matrix decreases its WVP, primarily attributed to essential oil droplets, making it more difficult for water vapor to diffuse through the film [60]. In addition, Shahrampour and Razavi [61] have reported that the WVP value of the film increases with increasing concentration of the added antimicrobial agent. Despite this, the antimicrobial agent facilitates moisture transmission through the film by extending the intermolecular interactions while helping to loosen the structure and reduce its tightness, thus increasing the WVP value of the film. Moreover, Pranoto et al. [62] reported an elevation in WVP values from 0.02309 to 0.03363 g m/m² day kPa following the addition of 150 mg/g potassium sorbate to chitosan films. In contrast, the addition of 153 U/g streptozotocin lactate to chitosan resulted in a significant elevation in WVP values from 0.02309 to 0.02762 g m/m². Therefore, the size and distribution uniformity of the oil droplets distributed in the film are the main factors affecting the WVP of the film [63]. There are two possible reasons for the reduction of film substrate density. First, there could be larger sized droplets and instability in the NE. Second, droplet flocculation in the film during the drying process may form larger oil holes or cracks, which can also decrease film density [63]. It is worth mentioning that the essential oils of NE have been proven to provide antimicrobial activity, while the thymus daenensis essential oil NE coated application has the property of inhibiting tenfold E. coli development compared to the free oil ones [64]. In this study, the utilization of TO-NE coating can effectively mitigate the TPC of papaya within the storage period, which was consistent with the published findings, such as 400 μL L⁻¹ TO coating inhibited the anthracnose development in mangoes [16] and ginger oil positively protected papaya from disease infections [65]. Therefore, this study indicated that TO-NE-coated papayas were effective in delaying the onset of spoilage with a lower decay rate. It was attributed to the TO-NE coating, effectively preventing microbial attachment, infection, and spoilage. The results of this study were similar to those of Manzoor et al. [66], who reported that NE coating (antioxidants and antimicrobial) had been used to reduce the rate of kiwi fruit decay. Thus, the antimicrobial mechanism of essential oils delays fruit decay by depolarizing the mitochondrial membrane [50]. Specifically, it increases cell permeability and ion transport imbalance, leading to apoptosis through microbial death [50]. Moreover, satisfactory results were obtained in this study regarding the shelf life prolongation of papaya treated with TO-NE coating. These results were also in line with the results reported by Chu et al. [67], who used NE coating with cinnamon essential oil for the shelf-life extension (2–3 days) of strawberries. Above all, the TO-NE-coating treatment in this study extended the shelf life of papaya. This approach may improve decay rates by considering possible combinations with other essential oils [10, 68], bioactive extracts of plant or microbial origins [69,70,71], or physical methods.

Conclusions

This study revealed that using a 0.25 mg/g TO-NE coated was remarkably successful in preserving the postharvest quality and prolonging the shelf life of the Tainung No. 2 papaya. More specifically, the TO-NE-coating treatment for papaya in this study was applied principally to delay ripening and maintain firmness by decreasing the papaya's physiological activities (such as respiration rate, ethylene production, and PG and PME activities). In addition, the papaya kept ascorbic acid content during the shelf life while retaining a visually appealing appearance. The delay in the accumulation of TSS and chlorophyll degradation was linked with the delayed color change. Moreover, the TPC on the papaya's surface was minimized, leading to a lower decay rate. Unfortunately, the 0.5 mg/g TO-NE-coated treated results were color change disordered, which requires more in-depth investigation of more suitable auxiliaries or different formulation ratios for improving the defects of WVP, which will be a valuable direction for the following research stage. Hence, the results of this study present an opportunity for notable advancements in the preservation, shipping, and marketing of papaya.