Combined Vacuum Impregnation and Electron-Beam Irradiation Treatment to Extend the Storage Life of Sliced White Button Mushrooms (Agaricus bisporus) Zeynep Sevimli Yurttas, Rosana G. Moreira, and Elena Castell-Perez

This study assessed the application of an antibrowning solution using vacuum impregnation (VI) and then electron-beam irradiation as a means to extend the shelf life of sliced white button mushrooms (Agaricus bisporus). A preliminary study helped to determine the best antibrowning solution and VI process parameters. Mushroom slices were impregnated with 2 g/100 g ascorbic acid + 1 g/100 g calcium lactate; 2 g/100 g citric acid + 1 g/100 g calcium lactate; 1 g/100 g chitosan + 1 g/100 g calcium lactate; and 1 g/100 g calcium lactate at different vacuum pressures and times and atmospheric restoration times. Selection of the antibrowning solution and VI parameters was based on texture and color of the mushroom slices. Next, the slices were irradiated at 1 kGy using a 1.35-MeV e-beam accelerator. Physicochemical, sensory, and microbial quality of mushrooms was monitored for 15 d at 4 ◦ C. The best impregnation process in this study was 2 g/100 g ascorbic acid and 1 g/100 g calcium lactate at 50 mm Hg for 5 min and an atmospheric restoration time of 5 min. The control (untreated) samples suffered structural losses throughout storage. Only the vacuum impregnatedirradiated samples had acceptable color by the end of storage. Sensory panelists consistently preferred the samples produced with VI and irradiation because exposure to ionizing radiation inhibited growth of spoilage microorganisms. Keywords: dose uniformity ratio, dosimetry, quality, spoilage

Extending the shelf life of mushrooms is important for their marketing and distribution and reliable preservation methods are still needed. Treating sliced mushrooms with vacuum impregnation and electron-beam irradiation introduces physiologically active components such as beneficial antioxidants, vitamins, and cations, while assuring safety and maintaining quality.

Practical Application:

Introduction

However, these techniques have associated drawbacks including safety, discoloration, and off-flavors production, and may be unsuitable for use on an industrial scale (Duan and others 2010). In the last decades, vacuum impregnation (VI) has been used as a means to introduce liquids into porous foods. This technique alters the product’s composition, physical, and chemical properties to improve its nutritional value (Fito and others 2001; Hironaka and others 2011). Furthermore, VI of a coating solution may be an alternative to improve the dispersion retention and to form a thicker, more effective coating. Vargas and others (2009) observed that chitosan-based edible coatings applied to fresh-cut carrots by VI enhanced all the positive effects of the coating. Sometimes, effective preservation of fresh-cut products may be achieved using a combination of several treatments (Garcia and Barrett 2002). Irradiation using gamma rays and electron beams maintains the produce quality and enhances shelf life in 2 ways: it kills spoilage organisms and retards plant ripening (Moreira and Castell-Perez 2012; Mami and others 2013). The recommended dose for enhancing the shelf life of mushrooms in different countries ranges from 1 to 3 kGy (Akram and Kwon 2010). However, irradiation at doses greater than 1 kGy can induce negative quality MS 20130805 Submitted 6/14/2013, Accepted 9/26/2013. Authors are with effects such as texture loss due to tissue softening (Koorapati and Dept. of Biological and Agricultural Engineering, Texas A&M Univ., College Sta- others 2004; Niemira and Fan 2009; Moreira and Castell-Perez tion, TX 77843-2117, U.S.A. Direct inquiries to author Castell-Perez (E-mail: 2012). Thus, the need to explore the synergistic effect of VI and [email protected]). irradiation in extending the shelf life of fresh-cut mushrooms.

White button mushrooms have an impact on treatment and prevention of breast cancer, mainly in postmenopausal women; they seem to inhibit the activity of aromatase, an enzyme involved in estrogen production (Teichmann and others 2007; Wani and others 2009). The mushroom industry presents about 5% to 25% of its fresh produce as slices and consumer demand for ready-to-use vegetables has increased the market for sliced mushrooms (Brennan and Gormley 1998). However, it is challenging to maintain their quality because of the lack of cuticle to protect them. Their larger surface area broadens the spoilage problems. Some critical changes include browning and softening (Brennan and Gormley 1998; Sommer and others 2010). Preservation of mushrooms has been attempted using packaging (Lopez-Briones and others 1992), chemicals (Sapers and others 2001), washing (Cliffe-Byrnes and O’Beirne 2008), tyrosine inhibitors (Singh and others 2010), ozone (Yuk and others 2007), and irradiation (Roy and Bahl 1984; Beaulieu and others 2002).

 R  C 2013 Institute of Food Technologists

doi: 10.1111/1750-3841.12308 Further reproduction without permission is prohibited

Vol. 79, Nr. 1, 2014 r Journal of Food Science E39

E: Food Engineering & Physical Properties

Abstract:

Fresh-cut mushroom shelf-life extension . . . groups without any treatment served as controls. The impregnated R, and control samples were placed into plastic containers (Ziploc 591 mL, with plastic lids) and stored at 4 ◦ C for analysis at days 0, 4, 9, 12, and 15 of storage. Impregnated liquid fraction (X). Sample weight before (W1 ) and after (W2 ) the impregnation treatment was measured Materials and Methods and impregnated liquid fraction (X) values were calculated. The Experimental design impregnated liquid fraction represents the total external liquid that In a preliminary study, mushroom slices were vacuum impreg- penetrates into the tissue. It was calculated by the weight differ◦ nated (VI) with several solutions. During storage at 4 C, color ence of the samples before and after the treatment (Ortiz and and texture were monitored and the best conditions (impregnation others 2003) using the following formula: solution, vacuum pressure and duration, and atmospheric restoration time) determined. The best conditions were then used to W2 − W1 X= × 100 (1) design the next set of experiments, which consisted of 1 set with W1 3 experiments: (1) irradiation, (2) VI, and (3) VI plus irradiation (see Irradiation experiment section). Ten slices were used for each treatment with 8 replications. Due Four groups of samples were used for product quality and shelf to the sheer amount of samples required, the change in mushroom life analyses: (1) control (fresh samples without any treatment), (2) composition was calculated after the impregnation process only impregnated, (3) irradiated, and (4) impregnated-irradiated slices. after determining the most effective VI process. USDA food comThe experimental design was a 4 × 4 × 2 experiment consisting position values were used as reference (USDA 2011). The change of 3 factors: impregnating solution (4 different solutions), vacuum in water content after impregnation was confirmed by measuring pressure (50, 75, 100, and 125 mm Hg), pressure duration (5 and the moisture content of the control and the selected impregnated 10 min), and atmospheric pressure restoration (5 and 10 min). The sample. experiment was conducted in duplicate. Moisture content. Moisture content of impregnated (ascorbic acid + calcium lactate) and control slices were determined by Sample preparation weight loss after drying in a vacuum oven at 70 ◦ C until conLocally grown button mushrooms (Agaricus bisporus) were stored stant weight (AOAC 1990). The samples were randomly chosen. at 10 ◦ C and 95% relative humidity in plastic bags, washed under Each sample’s weight was recorded before and after drying. Moretap water, dried with absorbent paper, and sliced (Farberware, over, the weight of canisters was recorded for higher accuracy. Hillsboro, Tex., U.S.A.). The head sides and long stems where The samples (cap and stem) were first chopped into small pieces discarded. The equipment (slicer, knife, beakers, and strainers) (approximately 10 g) and then placed in aluminum canisters prior was sanitized using a 300 μL/L chlorine solution. Slices thickness the drying process. After removal from the vacuum oven, the samples were placed in a desiccator before recording the final weight. was 6.5 ± 0.3 mm with a 3 to 5 mm cap length. Measurements were carried out at room temperature in triplicate for the control and the selected treatment. Preparation of impregnation (antibrowning) solutions Antibrowning solutions. To prepare the chitosan solution, 0.5 g/100 g acetic acid (Glacial, Mallinckrotd Baker Inc., Paris, Electron-beam irradiation Ky., U.S.A.) and 1 g/100 g calcium lactate pentahydrate (SigmaUniform dose distribution within a sample is important when Aldrich, St. Louis, Mo., U.S.A.) was dissolved in distilled wa- designing irradiation treatments for food. Uniformity is described ter at room temperature. Next, chitosan (medium molecular by a low dose uniformity ratio (DUR), the ratio between the weight, Sigma-Aldrich) was added at 1 g/100 g concentration maximum and minimum dose absorbed by the sample. Although (Hernandez-Munoz and others 2006). Ascorbic acid (L-Ascorbic the ideal DUR should be close to 1, accepted DUR values for acid, Sigma-Aldrich) and citric acid (Citric acid Anhydrous, Fisher practical applications are between 1.5 and 2 (IAEA 2002). A dose Scientific, Fair Lawn, N.J., U.S.A.) solutions were prepared by dis- mapping study was conducted using an ion farm chamber to detersolving 2 g/100 g of each acidulant and 1 g/100g calcium lactate mine dose uniformity of the irradiation setup. Irradiation dosage pentahydrate (Sigma-Aldrich) in distilled water at room temper- was measured by placing Radiochromic film dosimeters (Far West ature. Similarly, 1 g/100g of calcium lactate pentahydrate was Technology Inc., Goleta, Calif., U.S.A.) at the front, center, and dissolved in distilled water at room temperature. backside of the mushroom slices, for a total of 3 dosimeters. Preliminary study. The best impregnation solution, time, and The radiochromic films were read after stabilization using a Rapressure combinations were determined by monitoring color and diochromic reader model 92 (Far West Technology Inc.). The DUR in this study was 1.44, ensuring uniform dose distribution texture during 15 d at 4 ◦ C. Impregnation procedure. A VI system composed of a vac- within the mushroom slice. The entrance dose was approximately uum pump (Emerson Motor Div., St. Louis) and a vacuum glass 0.45 kGy, maximum dose was approximately 0.65 kGy, and backR , Brazil) was used. Sliced mushrooms were imdose was 0.6 kGy for an applied dose of 0.5 kGy. For a target of desiccator (Pyrex mersed in beakers (approximately 13 slices per beaker, a strainer 1 kGy (this study), the same relationship was applied, with enused to keep them immersed) containing the different impreg- trance, maximum and backdoses of 0.90, 1.30, and 1.2 kGy, renation solutions (ascorbic acid, citric acid, calcium lactate, and spectively (DUR = 1.44.) It should be noted that sample thickness chitosan), 1 solution at a time. During the vacuum step, different was reduced to 3.91 ± 0.16 mm because the DUR was too high pressures (50, 75, 100, and 125 mm Hg) were applied for 5 and with samples 6.5 mm thick. This sample thickness was used for 10 min and afterward, the atmospheric pressure was restored for 5 the shelf life study. or 10 min. The impregnated samples were then drained and exIrradiation experiment. A group of 4 different samples cess liquid was removed from the surface with a paper towel. The was evaluated to determine the best treatment: (1) control (no The objectives of this study were to (1) determine the best VI setup in terms of mushroom color and texture, and (2) evaluate the effect of VI and e-beam irradiation on the physicochemical, microbiological, and sensory quality of fresh-sliced mushrooms.

E: Food Engineering & Physical Properties

E40 Journal of Food Science r Vol. 79, Nr. 1, 2014

Fresh-cut mushroom shelf-life extension . . . treatment), (2) impregnated (2 g/100 g ascorbic acid + 1 g/100 g calcium lactate), (3) irradiated, and (4) impregnatedirradiated slices. Samples were prepared using the same procedure described above. Prior to irradiation, a single slice was placed in a plastic bag (Mylar) (Zip SealTM , 8.64 × 10.16 cm,

48GaPET/PE/0.00035Foil/LLDPE, Transilwrap Co., Franklin Park, Ill., U.S.A) and sealed. The slices were irradiated at 1 kGy, the maximum allowed by the FDA for fresh produce. We did not irradiate at lower or higher doses because it has been proven ineffective (Sapers and others 1994; Koorapati and others 2004).

Table 1–Effect of impregnation treatment (ascorbic acid and citric acid + calcium lactate) on texture (firmness) of sliced mushrooms impregnated at different vacuum pressures and different atmospheric restoration times during storage at 4 ◦ C. Control

50 mm Hg 50 mm Hg 75 mm Hg 75 mm Hg 100 mm Hg 100 mm Hg 125 mm Hg –5 min –10 min –5 min –10 min –5 min –10 min –5 min 2 g/100 g ascorbic acid + 1 g/100 g calcium lactate solution (maximum force in newton)

a w 30.297 1 (2.092)

0

w 28.257

4

w 20.05

b

(1.010) h y 47.79 (1.237) f x 42.753 (2.409) e x 43.185 (2.315)

9 12 15

b,c,d

w 18.846

(1.663) a w 20.346 (1.904) a,b x 25.787 (1.020) e,f y 39.704 (1.924) a w 20.122 (0.751)

b,c

w 25.19

(1.562) b y 30.552 (2.270) a x 24.748 (1.337) b,c,d y 32.816 (1.331) b y 32.97 (1.008)

a,d

x 26.74

(0.756) b w,x 28.274 (1.568) b,c x 29.39 (0.871) a,b w,x 29.05 (1.836) b,c,e y 34.246 (2.413)

a

w 15.562

(0.559) c y 35.33 (1.938) c,g y 35.59 (1.548) d,e y 35.616 (1.998) c x 25.027 (1.510)

c

w 18.76

(0.686) a x 22.342 (1.271) c,f y 35.027 (2.822) a,b,c y 29.353 (0.750) a x 20.805 (1.430)

b,c

w,x 19.244

(1.473) a w,x 21.585 (2.423) a,c,e y 32.287 (2.770) a x 25.236 (2.594) a w 18.457 (1.858)

b

125 mm Hg –10 min

w,x 28.127

(0.864) a x 23.162 (1.809) a,c,d y 30.505 (2.950) c,d,e y 35.1 (3.910) a w 18.032 (0.978)

a

(0.394) 3b,c w,x 32.09 (1.920) b,d,e,f,g x 35.86 (3.226) b,c,d w,x 32.53 (1.904) a,b,c w 25.727 (2.908)

2 g/100 g citric acid + 1 g/100 g calcium lactate solution (maximum force in newton) 30.297a

0

w 1 (2.092) w 28.257

4

b

(1.010) h y 47.79 (1.237) f x 42.753 (2.409) e x 43.185 (2.315)

9 12 15

x 23.881

a

x 31.328

(2.177) b y 35.040 (1.897) a,b y 38.500 (2.116) a,b x,y 47.840 (10.534) a y 45.723 (2.956)

c,d

d x,y 35.573

(2.655) c y 49.355 (1.351) b,c z 43.705 (0.106) b x,z 24.820 (1.796) a,b x,y,z 33.96 (4.723)

x 29.297

(2.039) b x 32.613 (1.952) c y 46.236 (3.136) a,b x,y 50.545 (3.500) a,b x,y 28.415 (8.478)

b,c

(2.495) c y 48.436 (1.559) c y 46.32 (0.494) a,b x,y 45.77 (4.341) a,b x,y 52.46 (3.775)

x 26.127

a,b

(2.036) e y 61.34 (0.789) b,c z 43.385 (0.007) a,b x,y,z 39.86 (3.464) a,b x,y,z 26.36 (3.535)

x 31.145

b,c,d

(1.906) d y 53.315 (1.858) a,b z 37.55 (0.961) a,b x,y,z 58.775 (7.177) a,b x,y,z 18.415 (5.607)

x 28.87

a,b,c

x 30.116

(2.323) c y 46.656 (0.800) a x 34.623 (1.773) a,b z 55.805 (3.655) a z 56.01 (3.054)

b,c

(2.243) f z 67.543 (1.174) a x,y 34.83 (2.851) a,b y 39.31 (3.705) b w 19.605 (1.859)

1 Standard a,b,c

deviation. Means within a row, which are not followed by a common superscript letter, are significantly different (P < 0.05). within a column, which are not followed by a common subscript letter, are significantly different (P < 0.05).

w,x,y,z Means

Table 2––Effect of impregnation treatment (calcium lactate alone and chitosan + calcium lactate) on texture (firmness) of sliced mushrooms impregnated at different vacuum pressures and different atmospheric restoration times during storage at 4 ◦ C. Time (days)

Control a,b w 30.297 1 (2.092)

0

w 28.257

4

a,b,c

(1.010) e y 47.79 (1.237) b x 42.753 (2.409) x 43.185 (2.315)

9 12 15

50 mm Hg –5 min

y 34.61

b

50 mm Hg 75 mm Hg 75 mm Hg 100 mm Hg 100 mm Hg –10 min –5 min –10 min –5 min –10 min 1 g/100 g calcium lactate solution (maximum force in newton) y 25.125

a

y 29.106

a

(0.573) a,b,c,d x 29.74 (3.109) d z 40.503 (1.752) a w 15.35 (0.883)

(0.946) b,c,d z 31.87 (3.387) b y,z 26.97 (2.036) a x 15.94 (0.806)

(0.965) e z 40.717 (2.368) b,c y 31.71 (1.528) a x 11.675 (0.926)

∗∗∗

∗∗∗

∗∗∗

y 30.443

a,b

(1.336) d y 33.65 (1.828) a x 19.666 (1.605) a x 15.87 (1.256) ∗∗∗

y 32.313

a,b

z 27.525

125 mm Hg –5 min

a

y 24.84

a

125 mm Hg –10 min

z 36.147

b

(2.370) c,d y 33.408 (1.374) c y 33.066 (1.720) a x 19.675 (0.968)

(1.932) d y 33.57 (1.584) a x 14.943 (3.334)

(0.47) a y 26.226 (1.740) a x 15.03 (1.58)

(1.556) a,b x 27.773 (0.575) b,c y 31.478 (0.712)

∗∗∗

∗∗∗

∗∗∗

∗∗∗

∗∗∗

∗∗∗

∗∗∗

1 g/100 g chitosan + 1 g/100 g calcium lactate solution (maximum force in newton) 0 4 9 12 15

30.297c

w 1 (2.092) w 28.257

a,b

(1.010) a,c y 47.79 (1.237) d x 42.753 (2.409) a x 43.185 (2.315)

y 31.972

c,d,e

(2.012) a,b,c x 21.02 (1.230) c z 49.88 (0.494) b,c y 31.536 (3.195) ∗∗∗

y 25.52

a,b

(1.858) b,c y 28.615 (2.525) b y 29.303 (0.241) c,d z 35.996 (2.296) b x 13.07 (2.701)

e x,y 36.182

x 29.9

(2.173) d y 38.713 (2.896) b,c,d,e x 30.525 (1.421) c,d x,y 34.85 (3.255) a,b x,y 33.68 (3.422)

(2.326) c x 31.583 (2.323) a,c y 45.41 (0.895) b,c x 28.35 (1.456) a,b x 25.9 (3.733)

b,c

y 22.47

a

(0.980) c z 44.81 (1.258) a,d z 41.975 (0.601) a x 15.946 (3.506) ∗∗∗

y,z 32.426

c,d,e

(1.578) c y 30.56 (1.654) a,b,c z 43.77 (7.254) b,c y,z 31.24 (1.103) a,b z 14.23 (4.101)

y,z 30.787

c,d

(1.622) a z 37.846 (2.838) a,b,c y 26.53 (4.652) a,b x,y 23.853 (2.896) a,b x 16.785 (3.288)

y 35.17

d,e

(1.341) a,b y 30.8067 (2.731) a,b,c z 60.595 (3.486) c,d y 34.23 (5.624) a,b x 15.695 (5.395)

1 Standard a,b,c

deviation. Means within a row, which are not followed by a common superscript letter, are significantly different (P < 0.05). within a column, which are not followed by a common subscript letter, are significantly different (P < 0.05).

w,x,y,z Means

Vol. 79, Nr. 1, 2014 r Journal of Food Science E41

E: Food Engineering & Physical Properties

Time (days)

Fresh-cut mushroom shelf-life extension . . . Table 3–pH, color L∗ , and maximum force values for the con- Irradiation was carried out at room temperature using a 1.35 MeV trol, impregnated, irradiated, and impregnated-irradiated sam- Van de Graaff e-beam accelerator. After irradiation, samples were ples stored at 4 ◦ C for 15 d. stored at 4 ◦ C up to 15 d for shelf life analysis. Time Impregnated(days) Control Impregnated Irradiated Irradiated Quality parameters pH 0 4 9

12 15

c y 6.51 1 (0.08) y,x 6.30

a

E: Food Engineering & Physical Properties

(0.08) a x 6.18 (0.12) b x 6.18 (0.12) a x 6.20 (0.03)

a,b x 6.17

y 6.38

(0.12) a,b x 6.18 (0.08) a x 6.07 (0.07) a x 6.11 (0.06) a x 6.11 (0.07)

(0.08) a y,x 6.30 (0.11) a x 6.17 (0.10) a x 6.20 (0.09) a x 6.15 (0.03)

b,c

x 6.05

a

(0.01) a x 6.15 (9.862) a y 6.23 (0.10) a x,y 6.19 (7.796) a x 6.17 (0.02)

Color L∗ 0 4 9 12 15

b z 70.701 1 (2.501) y 67.902

c

(1.900) b x 59.994 (4.210) a,b w,x 55.743 (6.598) a w 52.631 (3.293)

4

b x 28.849 1 (5.571) a x,y 34.095

9

y 37.150

0

(4.958)

12 15

a,b

(5.707) b y 36.640 (5.622) b,c y 44.210 (8.742)

a,b c z 73.333 (1.633) (2.185) a b,c y 61.037 y 65.051 (4.298) (4.262) a c w 53.637 y 65.627 (2.599) (3.095) b a x 57.230 x 53.226 (2.454) (3.243) b a,b w,x,y 57.216 x 55.186 (4.835) (2.823) Maximum force [N] z 70.142

w 23.040

a

(3.252) a y 37.328 (6.322) a w,x,y 32.185 (8.353) a w,x 28.199 (6.168) a x,y 31.781 (6.538)

w 30.348

b

(4.178) a w,x 35.985 (7.757) b x,y 42.789 (6.155) c y,z 49.718 (8.239) c z 51.575 (6.696)

1 Standard a,b,c

w 68.503

a

(2.650) a,b x,y,z 61.821 (6.275) a,b y 56.516 (5.109) b y 58.077 (3.384) c z 63.525 (2.388) w 30.235

b

(5.604) a w 33.563 (9.862) a,b w 35.313 (10.574) a,b w 30.605 (7.796) a,b w 39.670 (7.378)

deviation. Means within a row, which are not followed by a common superscript letter, are significantly different (P < 0.05). w,x,y,z Means within a column, which are not followed by a common subscript letter, are significantly different (P < 0.05).

Soluble solids. Soluble solids concentrations in the (a) impregnated, (b) irradiated, (c) impregnated-irradiated, and (d) control samples were determined at room temperature using a handheld refractometer (Brix 35HP, Reichert Analytical Instrument, Inc., Buffalo, N.Y., U.S.A) and expressed in ◦ Brix scale. About 20 to 25 g of mushrooms were placed in stomacher bags and squeezed to yield around 10 g of juice. Measurements were carried out at room temperature and in triplicate for each treatment and control group. PH. The pH was measured using a digital pH meter (Cole Parmer, pH 500 series, #59003-20, Singapore) calibrated with standard solutions, pH 4, 7, and 10 before the experiment. About 20 to 25 g of mushrooms were placed in stomacher bags and squeezed to yield around 10 g of juice. Measurements were carried out at room temperature and in triplicate for each treatment and control group. Color. The color properties of the sliced mushrooms were determined using 20 pieces from the treated and control samples at each sampling interval at room temperature using a Lab Scan XE colorimeter (Hunter Lab, Inc, Va., U.S.A.) calibrated with a standard plate (Y = 94.00, x = 0.3578, y = 0.4567). Since our objective was to test whether the treatments maintained the whiteness of the mushroom slices, only L∗ values (lightness) were reported. The cap of the mushroom slices was placed in the aperture of the colorimeter. New samples were used for each sampling interval (days 0, 4, 9, 12, and 15) to avoid microbial cross-contamination of the slices. Texture. A shear test was applied with the Warner–Bratzler probe at 1.0 mm/s using a Texture Analyzer (TA.XT2i, Texture Technologies Corp., Scardale, N.Y.). Texture was defined as the maximum force required to shear (cut) the samples. A preliminary study showed that better results were obtained when the cap length was reduced to 3 ± 0.2 cm by cutting the sides of the mushroom slices and then combining 3 slices together. Thus, 3 slices (slice Figure 1–Mushroom slices treated under different conditions (day 0 at 4 ◦ C.)

E42 Journal of Food Science r Vol. 79, Nr. 1, 2014

Fresh-cut mushroom shelf-life extension . . . ysis of variance using Tukey’s multiple range tests. Statistical significance was expressed at the P < 0.05 level.

Results and Discussion Preliminary study on VI Color. By the end of the study (day 15), impregnation with citric acid, calcium lactate alone, and chitosan solutions by VI did not help maintain the lightness of mushroom slices, with L∗ values significantly (P < 0.05) lower than the control group (data not shown). This effect was true for all combinations of vacuum pressure and restoration time tested, demonstrating that these antibrowning agents are ineffective and higher concentrations of these agents may be required to achieve the desired browning inhibition effect, which may induce flavor changes in the produce (Harris and others 2007). The 2 g/100 g ascorbic acid solution containing 1 g/100 g calcium lactate applied at a 50 mm Hg for 5 min and 5 min restoration time was the most effective VI treatment in terms of color. L∗ values ranged from 60.56 on day 0 to 52.61 on day 15 compared to the control (72.91 and 48.6 on days 0 and 15, respectively). This finding is not surprising because ascorbic acid is the most effective antibrowning solution (Wang and others 2013) because of the interaction of the antibrowning agent with calcium and the mushroom’s tissue (Wang and others 2013). Perez-Cabrera and others (2011) found similar results on a study with minimally processed pears. Texture. The maximum force to shear the control samples on day 0 was around 30 N. On day 15, the force value was significantly (P < 0.05) higher since the samples were very rubbery and hard to shear. By day 15, the slices impregnated with ascorbic acid had the most similar texture characteristics to the control group on day 0, with differences (P < 0.05) due to the different conditions applied during VI (applied vacuum pressures and atmospheric restoration times) (Table 1). Figure 2–Mushroom slices treated under different conditions (day 15 at 4 ◦ C.)

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E: Food Engineering & Physical Properties

thickness approximately 6.5 ± 0.3 mm, total thickness approximately 19.5 mm) were used for the impregnation experiments. As stated earlier, we had to adjust the thickness of the slices when conducting the irradiation tests to ensure proper dose uniformity within the slices. Therefore, samples consisted of 5 (instead of 3) slices with total thickness of 19.5 mm. Ten replications were done for each shear test at room temperature. Microbiological analysis. Total aerobic plates, psychrotrophic, and yeast and mold counts were determined on days 0, 4, 9, 12, and 15 of storage. Under sterile conditions, 10 g of mushroom slices (cap and stem) from each treatment were stomached inside a sterile stomacher bag, mixed with 90 mL of 0.1 g/100 g buffered peptone water, and homogenized for 1 min; afterward, 10-fold dilutions were made in this diluent. All counts were performed using petrifilms (3M yeast and mold count plates, 3M aerobic plate count (APC), 3M microbiology, St. Paul, Minn., U.S.A.). APCs were incubated at 37 ◦ C for 48 h; psychrotrophic count plates at 4 ◦ C for 7 d; and yeast and mold count plates were incubated at 20 ◦ C for 7 d. After incubation, colonies were enumerated and results reported as log colony forming units (CFU)/g of sample. The experiments were carried out in triplicate. Sensory evaluation. Thirty-five students, faculty, and staff members at Texas A&M Univ. formed the consumer panel. Evaluation of control, impregnated, irradiated, and impregnatedirradiated slices was carried out under the same conditions on days 1, 9, and 15 of storage. The samples were placed into white plastic plates labeled with 3 random digits and presented to the panelists (Meilgaard and others 1998) who were asked to score the samples based on odor, color, firmness, and overall quality using a 9-point hedonic scale (a score of 1 represents “dislike extremely” and a score of 9 represents “like extremely”). Scores higher than 5 were considered acceptable. Statistical analysis. Data analysis was performed using SPSS software (version 20.0 for Windows 2011). Statistical differences between variables were analyzed for significance by one-way anal-

Fresh-cut mushroom shelf-life extension . . . Table 4–Aerobic, psychrotrophics, and yeast and mold plate impregnated samples had moisture contents of 92.2% and 92.9% counts for the control, impregnated, and impregnated- wet basis, respectively. irradiated samples stored at 4 ◦ C for 15 d. Aerobic plate count Time (days) 0 4 9 12 15

Control c x 5.653 1 (0.179)

E: Food Engineering & Physical Properties

c w 7.016 1 (0.134)

4

7.684c

12 15

x

(0.225) c y 9.320 (0.059) c y, z 9.433 (0.14) c z 9.774 (0.141) Yeast

0

b x 3.383 1 (0.686)

4

3.620b

9 12 15

Irradiated

4.891b (0.098) c b y 7.164 x, y 4.980 (0.593) (0.458) c b y, z 7.817 x 6.560 (0.404) (0.22) b b x, y, z 7.431 z 8.632 (0.169) (0.728) c b z 7.984 z 8.693 (0.206) (0.041) Psychrotrophic plate count

0

9

Impregnated

Next, we explored whether the above VI treatment combined with irradiation at 1 kGy would help preserve the physicochemiImpregnated- cal, sensory, and microbiological quality of the mushroom slices.

x

(0.573) b x 4.412 (0.206) b x, y 5.625 (0.206) b y 4.458 (0.342)

y

N/D N/D N/D N/D N/D

5.437b (0.389) b x 5.331 (0.159) b y 7.915 (0.106) b y 8.203 (0.616) b y 8.815 (0.379) and molds plate count x

2.761b (0.111) b x 3.043 (0.245) b x, y 3.960 (0.499) b x, y 4.621 (0.927) b y 4.985 (0.091)

N/A N/D N/A N/A N/A

x

N/A N/D N/D

2.536a (0.629) a y 3.370 (0.438)

x, y

1 Standard a,b,c

deviation. Means within a row, which are not followed by a common superscript letter, are significantly different (P < 0.05). w,x,y,z Means within a column, which are not followed by a common subscript letter, are significantly different (P < 0.05). Detection limit = 2.39 CFU/g. N/A = No values observed, N/D = Under detection limit.

Impregnation with citric acid (Table 1) yielded samples with different texture characteristics. Similarly, calcium lactate alone was not effective when applied by VI due to loss of quality in terms of structural losses (Table 2). The structural deformation was caused by the pressure changes during the VI process as observed by other authors (Perez-Cabrera and others 2011). Increasing vacuum pressure and vacuum time resulted in more structure deformation due to the applied pressures. Impregnation with chitosan (Table 2) produced sticky and rubbery samples by day 9 with force values lower (P < 0.05) than the controls by day 15. In brief, ascorbic acid + calcium lactate impregnation at 50 mm Hg for 5 min and 5 min restoration time effectively maintained the firmness of the sliced mushrooms stored at 4 ◦ C for 15 d. This VI treatment was also effective in increasing the amount of physiologically active components in the mushroom composition with calcium increasing by 10 times and ascorbic acid by 150 times, compared to the USDA database values of 0.003 g/100 g and 0.0021 g/100 g, respectively (USDA 2011). After 5 min under 50 mm Hg and 5 min restoration time, W1 = 22.96 ± 2.29 g, W2 = 27.19 ± 3.17, and X = 18.42 ± 3.04%. Control and ascorbic acid–calcium lactate– E44 Journal of Food Science r Vol. 79, Nr. 1, 2014

Combination Treatment: VI and Irradiation at 1 kGy Effect on quality attributes PH. The pH of fresh mushrooms was approximately 6.5 (Table 3). The pH values of all the impregnated samples were significantly (P < 0.05) lower than the nonimpregnated (control and irradiated alone) samples, because of the acidic nature of ascorbic acid. On day 15, all samples had a pH of 6.2, still acceptable for consumption (US FDA/CFSAN 2012). Soluble solids (o Brix). All samples had ◦ Brix values between 5.0 and 6.0 (data not shown) but no significant (P > 0.05) differences were found among the samples. Color. By day 15, lightness values between the control and the treated samples were different (P < 0.05) (Table 3). On day 9, the impregnated samples had higher L∗ values than the control and irradiated samples, probably due to the precipitation of calcium in the cell walls, which may have increased the sample’s opacity. Furthermore, dark brown spots and dark staining occurred on the surface of the controls. Figure 1 and 2 show the appearance of the slices on days 0 and 15 of storage, respectively. Figure 2 shows the beneficial effect of the combined vacuum impregnation and irradiation treatment in the color of the samples. Although ascorbic acid is effective in preventing enzymatic browning once it oxidized completely, darkening of samples occurred in impregnated samples due to melanin formation. Irradiation seems to reduce browning by slowing down enzyme oxidation (Niemira and Fan 2009). Texture. Irradiation induced softening of the sliced mushrooms (Table 3). This is a problem when exposing fruits and vegetables to ionizing radiation that causes depolymerization of cellulose, hemicelluloses, starch, and pectin, which results in tissue softening (Niemira and Fan 2009; Moreira and Castell-Perez 2012). VI also induced softening due to loss of structure. However, application of the antibrowning solution (2 g/100 g ascorbic acid) containing 1 g/100 g calcium lactate maintained the firmness of the sliced mushrooms (approximately 35 N). Impregnatedirradiated mushroom slices had more fresh-like texture than the irradiated slices, demonstrating the beneficial effect of VI with a calcium-containing solution when samples are going to be irradiated. The controls and nonimpregnated samples suffered considerable structural loss (very soggy); hence, the blade could not shear the samples.

Effect on Microbial Quality Three sample groups were evaluated: (1) untreated control, (2) impregnated, and (3) impregnated-irradiated.

Aerobics Impregnation with ascorbic acid–calcium lactate solution was effective in reducing the aerobics counts (P < 0.05) (Table 4). As expected, the combined VI and irradiation treatment was effective (P < 0.05) in reducing microbial growth (Kooropati and others 2004). The trend of microbial growth for the control and impregnated samples was similar, with an increase in counts with storage time. Impregnation treatment alone reduced counts only by 1-log compared to the combined treatment (under the detection limit).

Psychrotrophics The organisms usually responsible for spoilage of mushrooms are Gram-negative, psychrotrophic bacteria, particularly belonging to the Pseudomonae family. These microorganisms are more susceptible to irradiation than other types of spoilage bacteria (Koorapati and others 2004). The combined VI and irradiation treatment was very effective (P < 0.05) in reducing psychrotrophic microbial growth, under the detection limit by day 9 (Table 4). Ascorbic acid impregnation reduced psychrotrophs by 1.5 logs, though this treatment alone is not sufficient to stop their growth.

Yeast and molds There was a significant (P < 0.05) difference in counts among the control, impregnated, and impregnated-irradiated samples on day 0 (Table 4). After day 12, there was a drastic increase in growth, showing an approximately 2-log counts increase by day 15. This finding suggests that the 1.0 kGy dose may be insufficient to inactivate yeast and molds in sliced mushrooms. It is known that yeasts are more resistant to irradiation than molds (da Silva Aquino 2012).

Effect on sensory attributes Sensory results indicated that the impregnated, irradiated, and impregnated-irradiated samples were acceptable (P < 0.05) (Figure 3). On day 1, there was no difference (P > 0.05) among the color acceptability of the samples (Figure 3A). The control samples were unacceptable to the panelists by day 15 of storage (scores approximately 4.0), while the impregnated, irradiated, and impregnated-irradiated samples were acceptable (scores approximately 6.0). This finding is supported by the objective color measurements (Table 3). The odor scores of the control were significantly (P < 0.05) different after day 1 (Figure 3B). Since irradiation inhibited microbial growth, no odor changes were observed in the irradiated and impregnated-irradiated samples with scores > 6.0. Mushroom slices lost their original mushroom smell with time, but treated samples did not exhibit unfavorable odor changes. Throughout the evaluation period, the controls were less acceptable in terms of their texture attributes (Figure 3C). Although the objective texture measurements showed clear changes in firmness of the treated samples (Table 2), the panelists found all the samples acceptable throughout storage (scores > 6.0). The controls exhibited significant (P < 0.05) quality loss and became unacceptable by day 15 of storage (Figure 3D). The

Figure 3–Sensory (A) color scores; (B) odor scores; (C) texture scores; and (D) overall quality scores for control, impregnated, irradiated, and impregnatedirradiated samples stored at 4 ◦ C during 15 d. A score of 1 = dislike extremely, 5 = neither like nor dislike, and 9 = like extremely. A value above 5 is considered acceptable.

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Fresh-cut mushroom shelf-life extension . . .

Fresh-cut mushroom shelf-life extension . . . impregnated, irradiated, and impregnated-irradiated samples had consistently higher scores than the control group with the impregnated-irradiated samples receiving slightly (though not significant) higher scores ( >6.0). Product appearance is the most appealing attribute to the consumers. Since irradiation prevented microbial growth and reduced microbial-induced browning, irradiated and impregnatedirradiated samples were consistently rated higher.

Conclusions

E: Food Engineering & Physical Properties

Although calcium is a texture enhancer for fresh-cut fruits and vegetables, it was not effective in sliced mushrooms when applied using VI and spoilage by fungi resulted in loss of quality. Irradiation treatment alone caused softening of the sliced mushrooms; however, impregnation with 1 g/100 g calcium lactate–ascorbic acid solution helped to maintain mushroom firmness. Microbiological analysis demonstrated that e-beam irradiation had an impact on reducing aerobic and psychrotrophic populations, whereas higher doses were required for inhibition of yeasts and molds. Sensory tests indicated consumers’ acceptance of the impregnated, irradiated, and impregnated-irradiated mushrooms. Product appearance was the most important quality attribute. Irradiated and impregnated-irradiated samples had the highest overall sensory scores because of reduced microbial-induced browning. Results from this study demonstrated that e-beam irradiation and VI of a solution containing 2 g/100 g of ascorbic acid and 1 g/100 g of calcium lactate can extend the shelf life of sliced mushrooms. Future work includes evaluation of the effect of impregnationirradiation on poyphenoloxidase activation to understand the mechanism of browning inhibition; irradiation at doses higher than 1 kGy to reduce growth of yeasts and the effect on quality; evaluation of combination of ascorbic acid and citric acid solutions; and optimization of impregnation solution and irradiation dose combinations.

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