Abstract

Ascorbic acid (1% w/w) and calcium lactate (1% w/w) were vacuum impregnated into mushrooms to improve their physicochemical quality. Experiments were carried out using central composite design under response surface methodology with four independent variables, viz., solution temperature (ST) (35-55°C), salt concentration (SC) (4-12%), vacuum pressure (VP) (50-170 mbar), and immersion time (IT) (30-90 min) with constant solution to sample ratio (4 : 1). The ascorbic acid content of mushroom increased with vacuum pressure with a maximum content of 30.68 mg/100 g. The results also suggested that calcium lactate allowed a better maintenance of texture during storage. The optimum conditions obtained for vacuum impregnation (VI) were 41°C, 10%, 140 mbar, and 75 min of solution temperature, salt concentration, vacuum pressure, and immersion time, respectively. Under these conditions, the corresponding responses were 22.14%, 2.31%, 28.74 mg/100 g, 0.28%, 4.61, 9.85 N, and 5.37 of water loss, solute gain, ascorbic acid, titratable acidity, colour change, texture, and pH, respectively. The vacuum impregnated mushrooms had more firmness, soluble solids, and ascorbic acid content than the control group. Therefore, the result suggested that vacuum impregnation could be used as an effective and simple technique to improve the quality of mushroom.

1. Introduction

Among the edible mushrooms, white button mushroom (Agaricus bisporus) is the most appreciated mushroom worldwide because of its organoleptic, nutritional, and therapeutic properties [1]. Button mushroom have an especially high content of phosphorus, sodium, and potassium followed by Ca, Mg, Na, Fe, and Zn. It is an excellent source of good quality proteins with lysine and tryptophan that are normally deficient in cereals, also rich in several essential amino acids and vitamins (B2, niacin and folate) [2, 3]. However, mushrooms are perishable food products that deteriorate in a short period of time after harvest because of their high moisture content. At mild temperatures, osmotic dehydration (OD) with or without vacuum, is considered to be a minimal processing method that keeps the fresh-like quality of fruits and vegetables [4].

Application of vacuum in OD results in enhancing the rate of water loss (WL) with solute gain (SG) and introduces controlled quantity of an external solution into the pores of fruits and vegetables [5]. During vacuum impregnation (VI) process, due to hydrodynamic mechanisms (HDM) and deformation relaxation phenomena (DRP) internal gas or liquid voids are impregnated by the external liquid [6]. With DRP, the gas of the porous tissue subsequently flows out and is replaced by the components present in the impregnating medium [7]. After restoring the normal pressure, tissue contracts then VI solution enter into the earlier air-filled pores. The volume of liquid penetrating externally is nearly the total volume of the pores that was filled with gas initially [8]. VI results in a processed product with superior texture, nutrition, organoleptic properties, and microbial protection. Browning of fruits and vegetables with the removal of oxygen (O2) from the pores can be reduced by VI [8]. VI process depends on factors that influence the transfer mechanisms such as vacuum pressure and immersion time i.e., vacuum and relaxation, salt concentration, brine temperature, sample shape, and solution to sample ratio. The range obtained from the reported levels given in the literature review was between 50-200 mbar for vacuum pressure at temperature maintained between 30 and 50°C for vacuum time (2-20 min) followed by relaxation time (10-120 min). A weight ratio of solution to sample maintained between 4 : 1 to 10 : 1 is recommended to avoid significant dilution of the medium and subsequent decrease of the driving force during the process [2, 9, 10].

VI has emerged as a useful way for incorporation of physiologically active compounds (PAC) into the porous structure of fruits and vegetables. PAC impregnated into pores of solid matrices like calcium lactate act as firming agent which improve texture quality, ascorbic acid act as an antioxidant, and citric acid reduces microbial growth [11]. The aim of the present study was to evaluate the effect of VI parameters viz. solution temperature (ST), salt concentration (SC), vacuum pressure (VP), and immersion time (IT) on water loss (WL), solute gain (SG), ascorbic acid (AA), titratable acidity (TA), colour change (CC), texture (TE), and pH then to optimize these parameters for further developing higher quality finished product. Subsequently, the effect of VI along with composite chemicals like ascorbic acid and calcium lactate on physicochemical properties of mushroom was also studied.

2. Materials and Methods

2.1. Material

Freshly harvested mushrooms having moisture content 88-92% were procured from mushroom farm, Punjab Agricultural University Ludhiana. Mushrooms were kept at refrigeration temperature of 4°C before experiments and were equilibrated at room temperature prior to treatments ([12, 13]).

2.2. Sample and Solution Preparation

Mushrooms (35-40 mm) were sorted and thoroughly washed with cold water () (stalks were partially removed) followed by sanitation using sodium hypochlorite (100 mg/l, 25°C) for 2 min [14, 15]. Consequently, sorted mushrooms were immersed in citric acid solution at 25°C (3 g/l) for 10 min to arrest browning [16].

After pretreatment, 50 g of mushrooms were exposed to different vacuum assisted osmotic pretreatment experimental combinations using 4 factor central composite design using different concentrations of salt solution. Ascorbic acid (1%) and calcium lactate (1%) were added in hypertonic salt solution. Based on our preliminary treatments, the process parameters of VI were, i.e., solution temperature (ST) (35-55°C), salt concentration (SC) (4-12%), vacuum pressure (VP) (50-170 mbar), and immersion time 30-90 min (IT) with constant solution to sample ratio (4 : 1). Sanitized mushrooms pretreated with 3 g/l citric acid were kept as control.

2.3. VI Treatments

For each treatment, 50 g of mushroom was weighed and placed in VI osmotic solution. In all trials, the samples were immersed in hypertonic salt solutions, and vacuum pressure was applied for 15 min following atmospheric pressure restoration, keeping the mushroom dipped in the solution.

The seven dependent variables were water loss (WL), solute gain (SG), colour change (CC), ascorbic acid (AA), titratable acidity (TA), texture (TE), and pH.

2.3.1. Water Loss (WL) and Solute Gain (SG)

Mushroom samples (control and vacuum impregnated) were weighed before and after VI treatment using weighing balance. Then, the samples were dried in an oven at 105°C to a constant final weight [17]. The WL and SG of the mushroom samples were evaluated using the equations shown below [18]. where is the initial weight (g), is the weight after VI at time (g), is the initial dry matter (g), and is the dry matter after VI at time (g) of mushrooms.

2.3.2. Ascorbic Acid (AA)

Ascorbic acid content was calculated by standard indophenol method [19]. The dye used as an indicator which reduces to a colourless solution with an addition of AA. An excess of unreduced dye results in a rose-pink colour which is the end point of titration. 2,6-dichlorophenolindophenol dye solution was used for titration. Mushroom (10 g) was crushed with the help of pestle and mortar and was thereafter mixed with 10 mL of 3 percent metaphosphoric acid. The solution was filtered through Whatman’s filter paper no. 4. Briefly, 5 mL of the aliquot was titrated against the standardized dye to a pink end point which should persist for at least 15 s. The total ascorbic acid content is calculated with the formula

2.3.3. Titratable Acidity (TA)

TA of treated as well as fresh mushroom was estimated using the method of Ranganna [19]. 10 g of mushroom pulp was diluted to 50 mL volume using distilled water. 10 mL of unfiltered aliquot was titrated against 0.1 N NaOH to light pink end with a drop of phenolphthalein as an indicator. The total TA percentage was calculated with following equation:

2.3.4. Colour Change (CC)

HunterLab’s MiniScan XE plus colourimeter (Konica Minolta CR-10, Japan) was used to measure the colour of fresh and VI samples. The value measures whiteness or darkness i.e., lightness. The value shows greenness (-a) and redness (+a) while the value shows blueness (-b) and yellowness (+b). Whereas, , , and are the initial values of , , and , respectively [20].

2.3.5. Texture (TE)

Textural properties of fresh and impregnated mushroom samples were recorded with the help of texture analyzer (Model TA-TXT2, Stable Microsystems Ltd., UK). The samples were compressed by probe P75, compression to 50% strain. During compression, the prespeed and postspeed were fixed at 2 mm/s and 1 mm/s, respectively. The mean firmness value of 3 replicates was taken [20].

2.3.6. pH

pH of fresh as well as impregnated sample was measured by pH meter. Before the experiments, pH meter was calibrated with standard solutions of pH 4, 7, and 10. Mean of 3 replications was taken for each treatment.

2.4. Statistical Analysis

The statistical analysis was performed independently for each response variable with the help of response surface methodology (RSM) by using a commercial statistical package (Design Expert DX 13.0.5). The central composite experimental design of four variables and four levels resulting in twenty seven experiments with three central points was used. An analysis of variance (ANOVA) was conducted with a 95% confidence level, in order to determine the accuracy of the model to represent the data. ANOVA include the statistical significance of process variables on each response, the model coefficient of determination (), value, and the lack of fit.

3. Results and Discussion

3.1. Water Loss (WL)

It can be observed from Table 1 that WL varied from 11.04 to 22.71% irrespective of level of vacuum impregnation. The WL showed an increasing trend with increase in VP and SC (Figure 1(a)). The WL increased progressively with IT, ST, and VP over the entire vacuum impregnation process. Higher temperature results in faster WL due to swelling and plasticising of cell membranes as well as the better water transfer characteristics on the product surface due to lower viscosity of the osmotic medium. The WL increased with ST especially in the early stages of the immersion. The application of VI had beneficial effects on the acceleration of mass transference [8]. Similar kind of results has been reported for pineapple [21] and zucchini [22]. Statistical significance for linear, quadratic, and interaction terms was evaluated for WL by fitting the quadratic model to the experimental data (Table 2). The value calculated by least square technique was found to be 0.84, showing good fit of model to the data. The model was significant with value of 0.01 () (Table 2). The “lack of fit value” of 0.25 was insignificant, which indicated that the developed model was adequate to predict the WL. Table 2 indicated that IT and SC were the most influencing factors followed by VI; in addition, ST was least effective over water loss. The response surface graphs (Figure 1(a)) were generated for the fitted model as a function of two variables (SC and VP) while keeping IT and ST at their central value.

3.2. Solute Gain (SG)

The values of SG varied from 0.99 to 2.67% (Table 1). The SG increased rapidly with increase in SC, IT, VP, and ST (Figure 1(b)). Moreover, for a certain kind of osmotic solution, the rates of water loss and solute gain increased with increasing concentrations of the osmotic solution and vacuum pressure. Comparable results were also reported when working with celery stalks in salt solution during OD [23]. Application of vacuum during osmotic dehydration was studied to increase the rate of water loss and solute gain of various fruits and vegetables such as apples [24], papaya [5], and apple [25]. The statistical significance for linear, quadratic, and interaction terms was calculated for solute gain as shown in Table 3. The value is 0.93, showing good fit of model to the data. The model value implied that the model was significant () (Table 2). The lack of fit was insignificant, which indicated that the developed model was adequate to predicate the response. Response surface graphs (Figure 1(b)) were generated for the fitted model as a function of VP and SC while keeping IT and ST at their central value.

3.3. Ascorbic Acid (AA)

AA content in fruits and vegetables is used as a major indicator which may reflect the changes of quality [26]. The AA impregnation significantly increased with increase in vacuum pressure. Ascorbic acid content of fresh sample was 22 mg/100 g. From Table 1, it can be observed that AA varied from 21.18 to 30.68 mg/100 g during the vacuum impregnation. The ANOVA showed that VP has the most significant effect on ascorbic acid content () (Table 2). Similar results have also been reported for potato [27], apple [28], and potato chips [4]. Results indicated that the ascorbic acid content of vacuum impregnated mushroom increased with vacuum pressure, due to deairing of void space and incorporation of impregnation solution (containing 1% AA) into the mushroom. Penetration of impregnation solution into intercellular spaces of the mushroom results in enhancing its ascorbic acid content. The value was found to be 0.89, showing good fit of the model (Table 3). VP was the most influencing factor followed by ST, while IT and SC were least effective over AA. The three-dimensional response surface plot for AA as affected by process parameters has been given in Figure 1(c).

3.4. Titratable Acidity (TA)

TA measures the total acid concentration in food [29]. The values of TA ranged from 0.18 to 0.32% (Table 1). Minimum value of titratable acidity (0.18) was observed for samples impregnated in salt solution at SC of 6%, ST of 40°C, VP 80 mbar, and IT 45 min, and maximum titratable acidity (0.32) was observed at SC of 12%, ST of 45°C, VP of 110 mbar, and IT 60 min (Table 1). The value was 0.92 demonstrating good model fit (Table 3). The TA increased with IT, SC, and VP over the entire vacuum impregnation process. Figure 1(d) shows the effect of VP and SC on TA while keeping IT and ST at their central value. The model was significant as , and “lack of fit value” of 0.83 was insignificant, which indicated that the developed model was adequate to predict the TA (Table 2). The three-dimensional response surface plot for TA as affected by process parameters has been given in Figure 1(d).

3.5. Colour Change (CC)

With respect to consumer, preference colour of developed product is an important parameter which was evaluated on the basis of , , , and colour change [20]. Minimum value of colour change (3.07) was observed for samples impregnated in salt solution at SC of 6%, ST of 40°C, VP 80 mbar, and IT 75 min, and maximum colour change (6.39) was observed at SC of 8%, ST of 55°C, VP of 110 mbar, and IT 60 min (Table 1). However, the effect of ST was more pronounced on colour change (Table 2). The low temperature used during VI prevents heat damage to tissues and greatly preserves the colour, aroma, and flavour of raw materials, especially in vegetables with high porosity. Mushroom impregnated with ascorbic acid observed better colour retention than control sample. Similar results have been reported in the literature by Shah and Nath [30] where the authors observed that VI with antibrowning agents effectively controlled browning and hence changes in colour. Comparable results were also reported by Blanda et al. [31] in nectarines; Perez-Cabrera et al. [32] in pear; Yurttas et al. [9] in sliced white button mushrooms, and Tappi et al. [33] in apple. The value calculated by least square technique was found to be 0.86, showing good fit of model to the data (Table 3). The three-dimensional response surface plot for CA as affected by process parameters has been given in Figure 1(e).

3.6. Texture

The firmness of the impregnated product ranged from 5.84 to 11.05 N (Table 1). Impregnation of calcium lactate improves the texture of mushroom than control samples. SC and VP significantly affect the firmness; their effect was shown in Figure 1(f). Similar results were reported by Tappi et al. [34] which showed that VI with calcium lactate allowed a better maintenance of texture during storage. The VI treatments increased by the firmness values, which are associated with a condition that is solute gain, were reported by Moreno et al. [35] in strawberries with VP applied for 5 min at 50 mb at the beginning of the process. Similar results were reported by Mao et al., [36] in grapes and Assis et al., [37] in apples. The ANOVA showed that VP has the most significant effect on ascorbic acid content () (Table 2). The value was found to be 0.87, implies good fit of model to the data (Table 3).

3.7. pH

The pH value of fresh mushroom was approximately 6.5. pH of impregnated mushroom ranged from 5.32 to 6.4 (Table 1). Due to the acidic nature of AA and citric acid, the pH of all the developed samples were considerably lower than the control sample. Similar results have been reported in the literature by [38], where the authors observed that VI with lactic acid solution decreased the pH value to a greater extent than processing carried out at atmospheric pressure. Similar results have also been reported for zucchini [39], mushroom [12], carrot and eggplant [13], and broccoli [40]. The ST and VP significantly affected the pH () (Table 2). pH variability of VI samples with VP and SC is shown in Figure 1(g). The value was found to be 0.78, implies good fit of model to the data (Table 2).

3.8. Optimization

The second order polynomial equation was fitted to the experimental data of each dependent variable. Maximum importance was given to the texture and colour. Table 4 shows the desired goals for each independent variable and response. The software generating optimum values of independent variables were ST of 41°C, SC of 10%, VP of 140 mbar, and IT of 75 min. Optimum conditions of responses corresponding to the values of independent process variables were WL of 22.14%, SG of 2.31%, AA of 28.74 mg/100 g, TA of 0.28%, CC of 4.61, TE of 9.85 N, and pH of 5.37 with overall desirability of 0.85 (Table 4).

4. Conclusion

The response surface methodology was found effective for optimization of vacuum impregnation of button mushrooms. Optimal values of desired responses were WL of 22.14%, SG of 2.31%, AA of 28.74 mg/100 g, TA of 0.28%, CC of 4.61, and TE of 9.85 N with pH of 5.37 corresponding to solution temperature of 41°C, salt concentration of 10%, vacuum pressure of 140 mbar, and immersion time of 75 min with 15 min vacuum. The study manifested that applying vacuum during osmotic treatment had a considerable effect on the rate of WL and SG of mushroom. In addition, VI mixed solution (1% ascorbic acid and 1% calcium lactate) maintained ascorbic acid content, firmness, improved colour, and reduced pH of button mushroom. Therefore, the vacuum impregnation is simple and efficient way to incorporate PAC into the porous structure of fruits and vegetables.

Abbreviations

ST:Solution temperature
SC:Salt concentration
VP:Vacuum pressure
IT:Immersion time
VI:Vacuum impregnation
OD:Osmotic dehydration
WL:Water loss
SG:Solute gain
HDM:Hydrodynamic mechanisms
DRP:Deformation relaxation phenomena
PAC:Physiologically active compounds
AA:Ascorbic acid
TA:Titratable acidity
TE:Texture
RSM:Response surface methodology
ANOVA:Analysis of variance.

Data Availability

The datasets generated and/or analyzed during the current study are available from the corresponding author on reasonable request.

Additional Points

Novelty Impact Statement. White button mushrooms are highly perishable and lose texture and colour after a few days postharvesting. The suitability of vacuum impregnation process for incorporation of calcium lactate and ascorbic acid to improve its functionality was evaluated. The results obtained in the present study establish that the developed technology can be helpful in meeting the specific nutritional demands of consumers.

Disclosure

The contributing authors declare that: (i) the work described has not been published before, (ii) it is not under consideration for publication elsewhere, (iii) its submission to JFPP publication has been approved by all authors as well as the responsible authorities—tacitly or explicitly—at the institute where the work has been carried out, (iv) if accepted, it will not be published elsewhere in the same form, in English or in any other language, including electronically without the written consent of the copyright holder, and (v) JFPP will not be held legally responsible should there be any claims for compensation or dispute on authorship.

Conflicts of Interest

The contributing authors declare that there is no conflict of interest between them.

Authors’ Contributions

(1) Wagh Muktabai Dinesh was involved in the methodology, validation, formal analysis, data curation, and in writing the original draft. (2) Mohammed Shafiq Alam was responsible for the conceptualization, design of experiments, validation, resources, supervision, and funding acquisition. (3) Raouf Aslam was assigned for the formal analysis and software and for writing the original draft, reviewing, and editing.

Acknowledgments

The authors are grateful to ICAR-All India Coordinated Research Project on Post-Harvest Engineering and Technology for the financial assistance.