CHY547 W2024 - February 14/15, 2024 Structure of presentation 1 (10 min + 2-3 min questions) Introductory slide (1-2 slides) Objective of article (1 slide) Materials and methods (2 slides) Summary of research and conclusion (3 slides) Identify the strengths and weaknesses (2 slides) Usefulness and validity of research findings (1 slide) Pointers: Presentation structure makes sense. Show that you have the scientific foundation necessary to discuss your article Practice and be prepared to present ... avoid reading from notes! Structure of your written appraisal (~ 5 pages) (written as a duo) Introduction Objective of article Materials and methods Summary of research and conclusion Identify the strengths and weaknesses Usefulness and validity of research findings The 5 pages exclude the title page and references, should you choose to use some./n ELSEVIER ARTICLE INFO Quality of green beans (Phaseolus vulgaris L.) influenced by microwave and hot water pasteurization Keywords: Microwave pasteurization Frozen green beans Color Food Control 124 (2021) 107936 Zhi Quª, Zhongwei Tangª, Fang Liuª, Shyam S. Sablaniª, Carolyn F. Ross ¹, Sindhuja Sankaranª, Juming Tang a, Chlorophyll Aerobic mesophilic bacteria Ascorbic acid Contents lists available at ScienceDirect a Department of Biological Systems Engineering, Washington State University, Pullman, WA, 99164-6120, USA b School of Food Science, Washington State University, Pullman, WA, 99164-6120, USA 1. Introduction Food Control journal homepage: www.elsevier.com/locate/foodcont Corresponding author. E-mail address: jtang@wsu.edu (J. Tang). ABSTRACT Vegetables are critical components of a healthy diet and account for an essential part of ready-to-eat (RTE) meals. RTE vegetables can be consumed either as recipe dishes intended for the retail market or as pre- cooked ingredients for caterers or other food services (Choma et al., 2000). Thermal processing is a classical method of food preservation that has been widely used in the production of RTE vegetables. In commercial preparation of RTE meals for cold chain distribution, a thermal process that exposes food products at 90 °C for 10 min, or an equivalent lethality, at the cold spot (the spot in the food which receives the least heat) in packages is often used to achieve a 6-log reduction of non-proteolytic Clostridium botulinum. This process would provide the products up to 6 weeks' shelf life at 5 °C (ECFF, 2006). The chill tem- perature storage serves as a prudent second barrier to maintain quality and extend the shelf life of the RTE meals. The process mentioned above is sufficient to inactivate vegetative microbial flora; it may also activate dormant bacterial endospores (Krawczyk et al., 2017; Luu et al., 2015), leading to subsequent germi- nation and outgrowth of nonpathogenic vegetative cells during cold storage. Sonar, Rasco, Tang, & Sablani (2019) reported that the aerobic CONTROL CONTROL FOOD CONTROL CONTROL CONTROL CONTROL CONTROL PELL This research investigated the influence of 915 MHz microwave-assisted thermal pasteurization and conventional hot water pasteurization (F30=10 min) on chlorophylls, greenness (a*), ascorbic acid, and the growth of spoilage microorganisms in green beans in storage at 10, 7, and 2 °C for up to 100 days. Frozen cut green beans were processed in vacuum-sealed containers flushed with N₂. The beans suffered 28.3% and 33.9% losses of chloro- phyll a, 9.2% and 15.3% losses of ascorbic acid during the microwave and hot water processing, respectively. During storage, slower spoilage (21 days at 10 °C, 42 days at 7 °C, and 100 days at 2 °C), superior preservation of chlorophyll (47-50%), ascorbic acid (47-62%), and a lower increase in a* (2.8-5.2) were obtained in samples processed with microwave-assisted thermal pasteurization. Paenibacillus spp. were identified as the predominant bacteria in the spoiled green beans pasteurized with both methods. The results highlighted the potential of microwave pasteurization for producing safe, high-quality vegetable products. Check for updates https://doi.org/10.1016/j.foodcont.2021.107936 Received 29 June 2020; Received in revised form 22 January 2021; Accepted 23 January 2021 Available online 29 January 2021 0956-7135/© 2021 Elsevier Ltd. All rights reserved. of HACCP and Food Safety CONTROL CONTROL mesophilic bacterial growth in pasteurized green pea puree (F=10 min) reached 6 log CFU/g after 80 days at 7 °C. Sous vide processed julienne carrots (85 °C) stored at 8 °C for four weeks had a total plate count of 6.48 log CFU/g (Nyati, 2000). Carlin et al. (2000) also observed that the count of aerobic mesophilic bacteria in pasteurized broccoli, carrot, zucchini, leek, potato, and split pea puree (80 °C, 30 min) increased up to 6–8 log CFU/g after 20-day storage at 10 °C. It has been reported that pasteurized zucchini puree showed the most rapid bacte- rial growth, while cook-chilled soybean sprouts (90 °C, 10 min) had no aerobic bacterial growth when stored at 10 °C for 24 days (Koo, Kim, Lee, Lyu, & Paik, 2008). It is evident from the literature that various vegetables after thermal pasteurization supported different bacterial growth, leading to a wide range of shelf life in cold storage at different temperatures. Thus, it is important to assess bacterial growth at different storage temperatures in studying quality losses and shelf life of RTE vegetables. The thermal processing and storage of vegetables inevitably result in changes in physical characteristics and chemical composition, such as color, bioactive compounds, and antioxidants (Baardseth, Bjerke, Mar- tinsen, & Skrede, 2010; Mazzeo et al., 2011; Peng et al., 2017). Con- sumers' demand for high-quality, fresh-like products has stimulated the Z. Qu et al. development of new technologies aiming at reducing the adverse effects of processing and storage. Microwave heating has shown advantages in reduced heating time and improved heating uniformity when compared with conventional hot water heating (Khan, Tango, Miskeen, Lee, & Oh, 2017). The microwaves penetrate into the food and generate heat throughout the whole volume of food, making thermal conductivity and heat transfer coefficients no longer the limit of heat transfer (Burfoot, Griffin, & James, 1988). Another advantage of microwave heating is the direct conversion of electromagnetic radiation into heat within foods, which could improve energy efficiency (Regier & Schubert, 2001). These advantages over hot water heating often yield an increased pro- duction rate and a reduction in undesirable thermal degradation (Ahmed & Ramaswamy, 2007; Khan et al., 2017). 93.3 Several studies have compared the quality of different RTE foods preserved by microwave with hot water heating that delivered an equivalent level of control of foodborne pathogens. Pérez-Tejeda et al. (2016) reported less color changes in tomato puree after processing using microwaves (950W, F833 = 1 min). Marszałek, Mitek, and Skąpska (2015) concluded that microwave heating better preserved the color and nutrient of strawberry puree than conventional heat pasteurization. Published shelf life studies on pasteurized vegetables have been limited to puree or smoothie (Arjmandi et al., 2017; Benlloch-Tinoco, Igual, Rodrigo, & Martínez-Navarrete, 2015; Klug et al., 2018), which do not represent a typical range of commonly consumed RTE meals. In addition, the studies on microwave pasteurization of vegetables are based on domestic microwave ovens, which do not represent the pro- cessing conditions used in industrial microwave systems for the com- mercial production of RTE meals. The objective of this study was to systematically evaluate the effects of 915 MHz microwave-assisted thermal pasteurization and conventional hot water pasteurization (F30=10 min) on the quality of green beans stored at different temper- atures (10, 7, and 2 °C). The green beans were characterized in terms of aerobic mesophilic bacteria count, pH, chlorophyll, color, and ascorbic acid. 2. Materials and methods 2.1. Sample preparation Since our intention was to simulate industrial processes for year- around operation, we selected frozen green beans for this research. Frozen cut green beans (Phaseolus vulgaris L.), with a size of ~2.5 cm in length and 0.8 cm in diameter, and a moisture content of 90.5%, were purchased from the local market and stored in sealed bags at -18 °C. The green beans were blanched before flash freezing, as indicated by the manufacturer. Before testing, the green beans were defrosted in 4 °C water for 10 min and then drained off for 5 min. Approximately 226 g thawed green beans were filled into each rigid tray (Silgan Plastics, Chesterfield, MO) with a PP/regrind/tie/EVOH/tie/regrind/PP struc- ture and an inner dimension of 140 × 95 × 30 mm. The trays were vacuum sealed with lidding film tal oxide coated PET/biaxial ori- ented PA/PP) with a thickness of 101 ± 1.6 µm (Printpack., Atlanta, GA), using a vacuum sealer (Multivac T-200, Multivac Inc., Kansas City, MO). The sealing conditions were set at 200 °C for 8 s, with a vacuum (6.5 kPa absolute pressure) and nitrogen flushing at 45 kPa (absolute pressure). 2.2. Thermal processing and storage Thermal processing was designed to achieve the lethality of F30 = 10 min at the cold spot in the trays. The lethality value was calculated according to Eq. (1): F= jio 0 T-Tref (1) 2 Food Control 124 (2021) 107936 where T (°C) is the temperature measured at the cold spot at time t (min), Tref is the reference temperature (90 °C), and z represents the temperature sensitivity of the heat tolerance of the target microor- ganism. We selected a z-value of 10 °C for non-proteolytic Clostridium botulinum (ECFF, 2006). The cold spot in the trays during the hot water processing was located at the geometric center of the package. For microwave-assisted thermal pasteurization, a mashed potato model food containing chemical marker precursors was employed for cold spot determination (Bornhorst, Tang, Sablani, & Barbosa-Cánovas, 2017). The heating pattern inside the microwave processed model food, as indicated by the intensity of browning, was analyzed using a computer vision method developed by Pandit, Tang, Liu, and Mikhaylenko (2007). Based on the heating pattern, the cold spot was located off the geometric center and in the middle layer of the trays. Heat penetration studies were carried out by collecting time-temperature data at the cold spot in the food trays to determine the process schedules. The temperatures were recorded every 2 s by a mobile metallic temperature sensor and a TMI data logger workstation (TMI-USA Inc., Reston, VA) (Luan, Tang, Pedrow, Liu, & Tang, 2013, 2015). This measurement method has been validated with fiber optic temperature sensors for developing microwave-assisted pasteurization processes (Tang, 2015). 2.2.1. Microwave-assisted thermal processing The microwave processing was performed in a pilot-scale 915 MHz semi-continuous Microwave-Assisted Pasteurization System (MAPS) developed at Washington State University. The system consisted of four sections (preheating, microwave heating, holding, and cooling), and each section had a separate water circulation system to control the water temperature and flow. The microwave section had four single-mode cavities connected to microwave generators. Approximately 5 kW of power was applied to each of the first two cavities, and the other two cavities equally split 8.7 kW. A metal carrier that could carry eight trays was immersed in the water and moved through the four sections mentioned above. The details of MAPS can be found in Tang, Hong, Inanoglu, and Liu (2018). 2.2.2. Conventional hot water processing Conventional hot water (HW) processing was conducted by immersing the packages in a precision digital circulating water bath (Model 260, Thermo Scientific, Marietta, OH) set at 91 °C. Two trays were processed at a time. 2.2.3. Storage conditions A total of 144 trays were processed and stored with protection from light at 10 ± 1 (a common refrigeration abuse temperature), 7 ± 1 (domestic refrigeration temperature), and 2 ± 0.5 °C (lower chill tem- perature) for 21, 42, and 100 days (Cronin & Wilkinson, 2009; James, Evans, & James, 2008). Three trays were randomly selected for each time point (Day 3, 7, 11, 14, 17, 21 at 10 °C; Day 5, 10, 15, 21, 28, 35, 42 at 7 °C; Day 7, 14, 28, 42, 60, 80, 100 at 2 °C), and each tray was measured in triplicate for quality quantification and microbiological analyses. 2.3. Microbiological analyses Thawed or processed green beans were pureed using a hand blender (KHB100ER1, KitchenAid, Benton Harbor, MI). Before grinding, the blending arm and blade of the blender were sterilized using steam at 121 °C for 15 min, the motor body (holding part) was cleaned with 70% ethanol wipes. Every 30 s grinding followed by 30 s rest to allow ground green beans to cool down. The total grinding/resting time equaled 5 min. The total mesophilic bacteria were examined by diluting the green bean puree in 0.1% (w/v) of sterilized peptone water and plated on Plate Count Agar (PCA) and then incubated at 37 °C for 48 h, both aerobically and anaerobically. Yeast and mold were enumerated by dispensing Z. Qu et al. aliquots onto Potato Dextrose Agar (PDA) following incubation at 25 °C for 5 days. The PDA was acidified by adding 1 mL of tartaric acid per 10 mL of PDA. The aerobic mesophilic bacteria, yeast, and mold were analyzed before and after thermal pasteurization (Day 0), and at the selected sampling points as described in section 2.2.3. The anaerobic mesophilic count was only determined at the end of the storage (Day 21 at 10 °C, Day 42 at 7 °C, Day 100 at 2 °C). The selected dilutions from each green bean sample were plated in triplicate. All microbial counts were reported as log₁0 colony-forming units per gram of green bean (log CFU/g). The isolated colonies were identified by amplification of the 16S rRNA or the rpoB gene by PCR using universal eubacterial primers and comparing the results with sequences in the GenBank Database by the Washington Animal Disease Diagnostic Laboratory (Washington State University, Pullman, WA). 2.4. pH The pH was determined by mixing 3 g of puree with 27 mL of deionized water and measuring at room temperature (23 °C) with a pH meter (Oakton Instruments, Vernon Hills, Il). 2.5. Chlorophyll For chlorophyll determination, 3 g of green bean puree was ho- mogenized with 25 mL of acetone-water (80:20, v/v) for 5 min at 7000 rpm (Polytron PT 10/35 GT, Kinematica AG, Luzern, Switzerland). The mixture was then put in a shaker rotating at 300 rpm at room temper- ature for 30 min and centrifuged at 8870 g for 6 min (Sorvall Biofuge Primo, Thermo Scientific, Waltham, MA). The supernatant was collected and brought up to a volume of 50 mL with acetone-water for spectro- photometric analysis (V-5000, Metash Instruments, Shanghai, China) at the wavelengths of 663 and 647 nm, measured against a blank. Eqs. (2)— (4) were used for calculation (Lichtenthaler, 1987), and the chlorophyll content was expressed as mg per 100 g. Chlorophyll a = 12.25A663 -2.79A647 Chlorophyll b = 21.5A647 - 5.10A663 Total chlorophyll=7.15A663 +18.71A647 2.6. Color (2) (3) (4) The sample color was quantified in CIE L*a*b* color space using image analysis. L* corresponds to lightness/darkness, a* represents red (+) to green (-), and b* represents yellow (+) to blue (−). The green beans from the topmost layer of the trays were taken out for image analysis, which aimed to simulate the first impression of the consumers as they opened the package. A computer vision system described by Zhang, Tang, Liu, Bohnet, and Tang (2014) was used to take photos (resolution: 5184 × 3456). The RGB images were calibrated using a color reference card (QPcard 203, QPcard AB, Helsingborg, Sweden) in Adobe Photoshop CC (Adobe system, Inc., San Jose, CA). Image analysis was performed in MATLAB R2019b (MathWorks, Inc., Natick, MA) by converting RGB to L*a*b*. Thresholding was applied to select the region of interest through background removal. The color value obtained was the average color value of all pixels in the region of interest. The color difference was calculated based on Eq. (5): 2.7. Ascorbic acid 2 AE = √√ (L* − Lö)² + (a* − aj)² + (b* − bő)² where Lo, do, and be correspond to the initial values of L*, a*, and b*. (5) Quantification of ascorbic acid (AA) was determined by HPLC 3 following the method described by Scherer et al. (2012) with modifi- cations. Five grams of green bean puree was homogenized with 10 mL 3% (w/v) meta-phosphoric acid for 1 min at 7000 rpm. The homogenate was incubated at room temperature (23 °C) for 2 h and centrifuged at 8870 g for 6 min. The supernatant was collected and filtered through a 0.45 μm nylon filter. Ten microliters of this filtered aliquot was injected into an Agilent 1100 HPLC system (Agilent Technology, Santa Clara, CA) to chromatographically separate the mixture into its component peaks through an RP18 5-µm 4.6 × 250 mm column (Waters Corporation, Milford, MA) equipped with a diode array detector. The mobile phase was a 0.01 mol/L monopotassium phosphate solution adjusted to a pH of 2.6 with o-phosphoric acid. The separation was accomplished using a 15-min isocratic elution procedure with a flow rate of 0.5 mL/min and a column temperature of 25 °C. The detecting wavelength was 250 nm. Pure L-Ascorbic acid was used to build a standard curve, and the AA content was expressed as mg per 100 g green beans. 2.8. Data analysis Kinetic degradation of quality attributes was analyzed based on the analytical data collected during storage. The zero-order reaction model (Eq. (6)) and the first-order fractional conversion model (Eq. (7)) were found to best fit with the experimental data in this study: C=Co - kt C-Co co Coo Food Control 124 (2021) 107936 -kt (6) (7) where Co is the initial value of the food quality attribute at t = 0 (day), C is the value at time t (day), C… is the final equilibrium value, and k is the reaction rate constant (day¯¹), which is temperature-dependent and follows the Arrhenius relationship (Peleg, Normand, & Corradini, 2012): Ea RT k = koexp( (8) 3. Results and discussion where Ea is the activation energy (kJ/mol), R is the universal gas con- stant (0.008314 kJ/mol K), T is the absolute temperature (K), and ko is the pre-exponential factor. The analysis of variance and Tukey's honestly significant difference test at a 95% confidence level (P < 0.05) were performed to identify the differences among groups using SPSS v25 (IBM Corp., Armonk, NY). Two-tailed Pearson correlation analysis was carried out at the signifi- cance level of 0.01. 3.1. Thermal processing Typical temperature profiles at the cold spots in trays are shown in Fig. 1. MAPS processing included preheating for 35 min at 51 °C, mi- crowave heating for 4.5 min in circulating water set at 91 °C, holding for 7.5 min in circulating water set at 91 °C, and cooling for 5 min at 23 °C. In HW processing, samples were heated for 47 min at 91 °C and cooled for 10 min in ice water. The lethality at the cold spots reached F 14.1 ± 0.3 min and 13.3 ± 0.7 min for MAPS and HW processing, respectively. In MAPS processing, the 35 min preheating step at 51 °C took up almost 60% of the whole processing time. This was to allow the samples to reach a uniform initial temperature before microwave heating. In an industrial process, this can be achieved by heating pre-packaged meals in a water flume at a constant temperature or through hot fill of the beans in packages before sealing. In the microwave heating section, the sample temperature increased from 51 to 89 °C at a heating rate of 8.4 °C/min. In HW processing, the temperature increase was relatively fast in the early part of the 47 min heating, as the temperature difference Z. Qu et al. Temperature (°C) 100 80 60 40 20 0 0 Aerobic mesophilic bacteria (log CFU/g) Temperature-MAPS Temperature-HW Lethality-MAPS Lethality-HW 10 8 20 30 Time (min) 40 20 I 50 I 60 40 60 Time (day) 100 Fig. 1. Temperature profiles and thermal lethality at cold spots of microwave (MAPS) and hot water (HW) processing. 80 between the heating medium and green beans was large. The heating rate gradually decreased as the green bean temperature approached the medium temperature of 91 °C, due to the reducing temperature differ- ence. The average heating rate of HW processing was 1.6 °C/min, about 1/5 of that in the microwave heating section MAPS. higher heating rate in MAPS meant less thermal exposure, which should have positive effects on the quality of food products. 1 80 60 3.2. Microbiological analyses 3.2.1. Shelf life determined by bacterial growth The initial population of aerobic mesophilic bacteria in frozen- thawed green beans before thermal processing was 4.11 ± 0.15 log CFU/g. After processing, no plated colonies were detected in the samples processed by either method (limit of detection was 100 CFU/g), which was shown as 0 at day 0 in Fig. 2. The bacterial population increased dramatically to ~4 log CFU/g in the early part of the storage (day 3 at 10 °C, day 5 at 7 °C, and day 7 at 2 °C). The increase then slowed down to reach 7-8 log CFU/g at the end of the storage (Fig. 2). The initial rapid growth of the spoilage bacteria in the green beans may be explained by the availability of more nutrients for microbial growth from the damaged green bean plant cells and the lack of competing vegetative bacteria cells after the pasteurization. No yeast or mold counts were detected in any of the samples throughout the storage period, and no 40 -- HW-7°C 20 0 -- HW-2°C MAPS-10°C - HW-10°C MAPS-7°C Lethality (min) MAPS-2°C 1 100 Fig. 2. Aerobic mesophilic bacteria count of microwave (MAPS) and hot water (HW) processed green beans during storage at 10, 7, and 2 °C. (For interpre- tation of the references to color in this figure legend, the reader is referred to the Web version of this article.) 4 Food Control 124 (2021) 107936 anaerobic growth was observed in any samples at the end of the storage. Pasteurized RTE meals could be considered as spoiled when the total aerobic microbial counts reach 7 log CFU/g or higher (FSANZ, 2001). In this study, package swelling was the first sign of spoilage, together with the presence of an off-odor that appeared when the package was opened. All swollen packages had aerobic mesophilic bacterial counts larger than 7 log CFU/g. During storage at 10 °C, the population reached 7 log CFU/g on day 17 and day 21 in HW and MAPS processed green beans, respectively (Fig. 2). Reducing the storage temperature from 10 °C to either 7 or 2 °C could delay spoilage by 2.5-4.5 times, as the lower temperatures reduced the growth of microorganisms. The MAPS pro- cessed samples took 42 and 100 days, while HW processed samples took 35 and 80 days, to reach the microbiological quality mark when stored at 7 and 2 °C, respectively. The results suggest that microwave heating had the potential to produce pasteurized green beans with longer shelf life. 3.2.2. Identification of spoilage bacteria in pasteurized green beans Four morphologically different isolates from spoiled samples after 21 days at 10 °C were identified, of which three were classified as Paeni- bacillus spp. (Paenibacillus sp. F, Paenibacillus terrae, and Paenibacillus sp. FSL H7-0357) and one as Bacillus spp. (Bacillus pumilus). Only Paeniba- cillus spp. were identified from samples spoiled at 7 and 2 °C storage. All the bacteria identified are spore formers, which survived the pasteuri- zation process and germinated into metabolically active cells during storage. Similar to our study, Guinebretiere et al. (2001) reported high fre- quencies of the presence of Paenibacillus spp. in pasteurized zucchini puree stored at 10 °C. Helmond, Nierop Groot, and van Bokhorst-van de Veen (2017) isolated P. terrae from rice-vegetable RTE meals stored at 7 °C, and Carlin et al. (2000) isolated B. pumilus from pasteurized vegetable purees stored at 10 °C. Guinebretiere, Girardin, Dargaignar- atz, Carlin, and Nguyen-The (2003) examined the contamination flows of spore-forming aerobic bacteria in a pasteurized zucchini puree pro- cessing line.Paenibacillus spp. and Bacillus spp. are known to be frequent contaminants in the soil, and the spore counts in the soil can be as high as 9 log CFU/g (Ammann, Kölle, & Brandl, 2011). The spore-forming bacteria may enter the plant through the root or attach to the surface of the vegetables by contact with soil or aerosols of soil. Another possible contamination pathway is the processing equipment, such as storage tanks and production lines (Soni, Oey, Silcock, & Bremer, 2016). Both Paenibacillus spp. and Bacillus spp. have been isolated from food pro- cessing plants as they can attach to stainless steel surfaces (Huck, Woodcock, Ralyea, & Boor, 2007; Salustiano et al., 2009). Conse- quently, they could enter the food chain easily. 3.3. pH The initial pH of the green beans was 6.28. Thermal processing reduced the pH to 6.23 and 6.16 when MAPS and HW were employed, respectively. This may be due to the release of organic acids from the food matrix. The pH decreased in all samples during storage (Table 1). This was also demonstrated in other pasteurized vegetable puree during storage at 4-10 °C (Carlin et al., 2000; Sonar et al., 2019b). The pH drop during storage was caused by the growth of microbial flora, as observed in section 3.2. In the current study, the significant pH drop started somewhere around the middle of the shelf life. MAPS processed samples always had similar or higher pH as compared to HW processed ones. Referring to the microbial analysis in section 3.2, a pH value lower than 5.5 may suggest the end of the shelf life of pasteurized green beans due to spoilage. 3.4. Chlorophyll degradation The total chlorophyll content of unheated green beans was 10.46 mg/100 g, out of which 68.0% was chlorophyll a. Both heating methods Z. Qu et al. Table 1 pH of microwave (MAPS) and hot water (HW) processed green beans during storage at 10, 7, and 2 °C. 10 °C 7 °C Day 0 3 7 11 14 17 21 MAPS 6.23 ± 0.01ªA 6.00+ 0.01bCA 6.03 ± 0.01 BA 5.99 ± 0.01 CA 5.95 + 0.01 A 5.81 ± 0.02⁹A 5.47 ± 0.02fA Typical image L* a* b* ΔΕ pH caused significant (P < 0.05) degradation of chlorophyll a and b (Table 2). Compared with HW, MAPS induced less loss in chlorophyll a. Chlorophylls continued to degrade during storage following the first- order fractional conversion model described in Eq. (7) (Fig. 3). The ki- netic parameters of degradation are summarized in Table 3. As expected, the higher the storage temperature, the faster the degradation of chlo- rophylls over time, as indicated by higher degradation rates (k). Reducing the storage temperature from 10 to 2 °C could bring about a threefold reduction in k for both chlorophyll a and b. To better interpret the influence of storage temperature on chlorophyll degradation, chlo- rophyll content in samples after 17 days at 10 °C, 21 days at 7 °C, and 28 days at 2 °C were compared. MAPS processed green beans had total chlorophyll retentions of 49.6%, 53.2%, and 55.4%, while preservation of 45.4%, 49.3%, and 52.2% was obtained in the HW processed samples. Microwave technology provided green beans with greater chlorophyll retention over time, due to the lower k in MAPS processed samples than HW. The chlorophyll content of MAPS processed samples at the end of storage at 7 °C was significantly higher as compared with HW processed ones, and the values of those stored at 10 and 2 °C were still comparable (Fig. 3). HW 6.16 + 0.01 B 5.99 ± 0.02bA 6.04 ± 0.01 DA Schwartz and Von Elbe (1983) suggested that chlorophyll degrada- tion in vegetables is the result of chlorophyll degrade to pheophytin and further to pyropheophytin. First-order degradation kinetics of chloro- phyll were reported for fresh asparagus (Tenorio, Villanueva, & Sagar- doy, 2004), pasteurized green pee puree (Sonar, Rasco, et al., 2019), and frozen green beans (Martins & Silva, 2002) during storage. But Benl- loch-Tinoco et al. (2015b) reported second-order chlorophyll degrada- tion kinetics in microwave and conventionally heated kiwifruit puree; Table 2 Quality attributes of green beans before and after thermal processing. Before processing After processing MAPS 50.51 ± 1.41ª -29.52 ±0.08ª 44.06 ± 1.24ª 5.72 ± 0.01 CB 5.52 ± 0.01 dB 5.27 ± 0.03eB 5.15 + 0.02eB 6.28 ± 0.02² 7.11 0.10² 3.35 ± 0.18a 6.12 ± 0.10² Day 0 5 10 15 21 28 35 42 Values with different superscript letters have a significant difference (P < 0.05). Small superscript letters indicate the results in the same column, and capital su- perscript letters compare rows within the same temperature. 50.55 ± 0.52² -13.94 +0.71b 41.88 0.12ª 15.82 ± 0.44ª 6.23 ± 0.01b 5.10+ 0.12b MAPS 6.23 ± 0.01ªA 6.19 + 0.01 A 6.21 ± 0.02ªA 6.18 ± 0.02ªA 6.17 ± 0.02abA 6.08 ± 0.01 BA 5.92 ± 0.03CA 5.60 ± 0.08dA 1.86 ± 0.13b Chlorophyll a (mg/ 100 g) Chlorophyll b (mg/ 100 g) Ascorbic acid (mg/ 0 g) Means in rows followed by different letters differed significantly (P < 0.05). 5.56 ± 0.08¹ HW 49.63 ± 1.48² -9.39 ± 0.29€ 43.45 ± 0.37a 20.27± 0.52b 6.16 0.01⁹ 4.70 ± 0.13º 1.70 ± 0.16b 5.18± 0.03 HW 6.16 + 0.01 B 6.13 +0.00B 6.14 ± 0.01 B 6.13 ± 0.02aB 6.01 + 0.01 B 5.57 ± 0.02CB 5.39 +0.03dB 5.17 ± 0.03eB 5 2 °C Day 0 7 14 28 42 60 80 100 Food Control 124 (2021) 107936 MAPS 6.23 ± 0.01 A 6.16± 0.02¹A 6.14 ± 0.01 BA 6.12 + 0.01 DA 6.06 ± 0.02CA 6.02 +0.04cdA 5.99 ± 0.02dA 5.69 ± 0.01 A HW 6.16 + 0.01 B 6.06 ± 0.02bB 6.08 ± 0.01 B 6.11 +0.03abA 5.93 0.01 CB 5.26 +0.03dB 5.13 + 0.03eB the degradation rate decreased from 0.031 to 0.007 day-¹ when the storage temperature changed from 22 to 4 °C. Besides storage temper- ature, the kinetic of chlorophyll degradation is also affected by pH and microbial growth. Gunawan and Barringer (2000) showed that micro- bial growth accelerates the color change in broccoli by producing acids. The decrease in pH favors converting chlorophyll to pheophytin by facilitating the replacement of Mg²+ by H+ (Gunawan & Barringer, 2000). Koca, Karadeniz, and Burdurlu (2007) reported that the degra- dation rate of chlorophyll a in green peas at pH 5.5 was twice that at pH 6.5. The higher pH in MAPS processed samples may be another reason that more chlorophyll was preserved. Klug et al. (2018) reported that microwave (30s, 11 kW) caused less loss in chlorophyll in faba bean sauce compared to conventional pasteurization (85 °C, 5 min). Benlloch-Tinoco et al. (2015b) also confirmed the superiority of microwave heating in preserving chloro- phylls and carotenoids in kiwifruit puree stored at 4-22 °C. The observed difference could be explained by the higher heating rate and less exposure time to high temperature in microwave heating. 3.5. Color degradation Both MAPS and HW processing induced considerable changes in color, as indicated by an increase in the a* values (Table 2). No signif- icant changes in L* and b* occurred during processing or storage, so a* was used as the greenness indicator. A larger increase in a* was observed when the green beans were conventionally heated. The more significant change in a* of HW processed samples led to a significantly higher AE. A AE value of 3 is perceptible by most people (Paciulli, Palermo, Chiavaro, & Pellegrini, 2017), and the color changes were visually noticeable as shifting from bright green to olive green, as shown in typical images in Table 2. The increase of a* during storage followed the first-order fractional conversion model (Eq. (7)). Storage temperature showed a considerable effect on the rate of reduction in greenness. A reduction in the storage temperature from 10 to 2 °C could reduce the color degra- dation rate from 0.149 and 0.173 to 0.0359 and 0.0363 day in MAPS and HW processed green beans, respectively (Table 3). As shown in Fig. 4, microwave processed green beans had significantly (P < 0.05) lower a* at each measured time point regardless of storage tempera- tures. The a* of microwave heated green beans at the end of storage at all temperatures was between 2.8 and 5.2, while it was between 5.1 and 6.8 for conventionally processed samples. This suggests that MAPS better preserved the green color of green beans during storage. -1 An increase in a* during thermal processing of green vegetables was observed previously (Aamir, Ovissipour, Rasco, Tang, & Sablani, 2014; Steet & Tong, 1996). Besides the increase of a*, Kotani, Yamauchi, Ueda, Imahori, and Chachin (1999) reported that L* and b* were unchanged in boiled broccoli florets during storage at 10 °C. Several studies compared the effects of microwave and conventional heating on color. Faba bean sauce preserved by microwave had a smaller increase in a* after pro- cessing (Klug et al., 2018). Microwave heated kiwifruit puree had a color change of 6.54 after 123 days storage at 4 °C, while the color change was