Physicochemical properties and bioactive compounds of different varieties of sweetpotato flour treated with high hydrostatic pressure

Food Chemistry 299 (2019) 125129

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Physicochemical properties and bioactive compounds of different varieties of sweetpotato flour treated with high hydrostatic pressure Rongbin Cui, Fan Zhu

T

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School of Chemical Sciences, University of Auckland, Private Bag 92019, Auckland 1142, New Zealand

A R T I C LE I N FO

A B S T R A C T

Keywords: High pressure processing Ipomoea batatas Thermal property Pasting property Starch digestibility β-Carotene Phenolics Anthocyanin

Three varieties of sweetpotato flour (Orange Sunset (OS), Purple Dawn (PD), and Red) were treated by high hydrostatic pressure (HHP). Thermal analysis showed that complete starch gelatinization occurred in PD and Red subjected to 600 MPa. Starch of OS was partially gelatinized at 600 MPa. The pressure of 600 MPa caused significant decreases in peak viscosity, breakdown and setback, and an increase in pasting temperature. Compared with native samples, HHP-treated samples showed higher in vitro starch digestibility in uncooked conditions but lower digestibility in cooked conditions. HHP significantly improved the extractability of bioactive compounds from sweetpotato flour. The changes in β-carotene content, total phenolic content, in vitro antioxidant activities, individual phenolic acids, and anthocyanins profiles were investigated. This study suggests the potential of HHP as a non-thermal processing tool to modify the functional properties of sweetpotato flour.

1. Introduction Sweetpotato (Ipomoea batatas Lam.) is widely cultivated and consumed in many developing countries. The world sweetpotato production has reached over 105 million tonnes (Wang, Nie, & Zhu, 2016). Sweetpotato plays a significant role in global food security. Processing sweetpotato into flour greatly increases its storage ability and value. The major component of sweetpotato flour (SPF) is starch (> 60%). SPF has processability similar to cereal flour and has been processed into a range of products, such as soup thickener, gravy, snacks and bakery products (Wang et al., 2016). Sweetpotato roots are rich in a range of nutrients, such as carbohydrates, dietary fibers, and minerals. Sweetpotato can be a good source of slowly digestible starch (SDS) (Lehmann & Robin, 2007). Depending on genotype, some varieties contain a significant amount of bioactive compounds, such as phenolic acids, anthocyanins and carotenoids (Zhu, Cai, Yang, Ke, & Corke, 2010). However, when food is thermally processed, the composition of bioactive components and other quality attributes, such as color, are likely to change. For example, heat treatment caused a significant reduction in the anthocyanin content in purple-fleshed sweetpotatoes (Kim et al., 2012). To overcome possible negative effect of heat treatment, non-thermal processing methods such as high hydrostatic pressure (HHP) processing have been used as an alternative to traditional thermal processing (Oey,

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Lille, Van Loey, & Hendrickx, 2008; Tiwari, O'Donnell, & Cullen, 2009). Under HHP, starch granules swell and undergo a gelatinization transition. In contrast to starch gelatinization caused by heat treatment, the HHP treated starch granules usually keep their intact form due to the limited effect of pressure on hydrogen bonds in starch granules (Leite, de Jesus, Schmiele, Tribst, & Cristianini, 2017; Yang, Chaib, Gu, & Hemar, 2017). The degree of gelatinization highly depends on the type of starch, water content and pressure level. For example, A-type starch is more sensitive to pressure treatment than B-type starch, while C-type starch is in between A- and B-type starches (Yang et al., 2017). The extent of starch gelatinization significantly influences both the textural and rheological properties of starch-based food products (Liu, Hu, & Shen, 2010). HHP affects not only starch but also other food components such as proteins and fibers (Pérez-Andrés, Charoux, Cullen, & Tiwari, 2018). The effects of HHP on physiochemical and functional changes in food products such as beef, fish, ham and milk have been summarized previously (Pérez-Andrés et al., 2018). HHP can also disrupt the cell wall, organelles and membranes, which might improve the extractability of polyphenol compounds (Oey et al., 2008). Studies showed that the extractability of polyphenols from green tea leaves and the antioxidant activity in tomato puree were significantly improved by HHP processing (Patras, Brunton, Da Pieve, Butler, & Downey, 2009; Xi et al., 2009).

Corresponding author. E-mail address: [email protected] (F. Zhu).

https://doi.org/10.1016/j.foodchem.2019.125129 Received 6 March 2019; Received in revised form 3 July 2019; Accepted 3 July 2019 Available online 04 July 2019 0308-8146/ © 2019 Elsevier Ltd. All rights reserved.

Food Chemistry 299 (2019) 125129

R. Cui and F. Zhu

2.3. Analysis of morphological and physicochemical properties

The effect of HHP is variable and is dependent on not only the HHP operating conditions (e.g., pressure, hold time, temperature) but also on the food form (e.g., whole, puree or juice) (Oey et al., 2008). To the best of our knowledge, most of the previous studies have been focused on high pressure treatment of pure starch. As the type of plant material is also important, the impact of varieties within species on food quality affected by HHP is much less understood. Little information is available concerning the effect of HHP on the complex food systems such as flour, especially on the phenolic acids and anthocyanins profiles of HHPtreated flour. Therefore, the objective of this work was to study the effects of HHP up to 600 MPa on bioactive compounds and in vitro antioxidant activity of SPF from sweetpotato roots collected in New Zealand (also called kumara). The chemical composition of these SPF has been detailed in a previous report (Cui & Zhu, 2019). The functional, structural, rheological and thermal properties of different varieties of SPF under HHP processing were also examined.

2.3.1. Morphological properties Flour samples were spread on a double-sided adhesive tape attached to a metal subbase and coated with platinum under vacuum using a Hitachi E-1045 ion sputter coater (Tokyo, Japan). The morphology of SPF was examined by a Hitachi SU 70 scanning electron microscopy (SEM) (Tokyo, Japan) at an acceleration potential of 10 kV. 2.3.2. Particle size distribution The particle size distribution of SPF was analyzed using a Mastersizer 2000 particle size analyzer (Malvern Instruments, Worcestershire, UK) according to the method of (Li & Zhu, 2018). Briefly, 1% of flour suspension was prepared in an Eppendorf tube and added into the sample dispersion unit of the instrument by a dropper. The suspension was added until a proper obscuration factor was reached (5–10%). The particle size measurement was done from 0.01 to 10000 µm. A total of 100 single particles were measured in one trial. The volume-weighted mean diameter (D [4, 3]), surface-weighted mean diameter (D [3, 2]) and median diameter (d (0.5)) were recorded.

2. Materials and methods 2.1. Materials

2.3.3. Fourier transform infrared spectroscopy (FTIR) The Fourier transform infrared spectroscopy (FTIR) spectra of SPF were obtained using a Bruker Vertex 70 FTIR spectrometer (Bruker Daltonics, Bremen, Germany) equipped with a single bounce diamond accessory. Flour was covered on the crystal area of a multi-bounce plate. Spectra were obtained between 4000 and 400 cm−1 in transmission mode. Each spectrum was collected from an average of 16 scans and the results were reported as mean values.

Three varieties of sweetpotato roots (Orange Sunset (OS), Purple Dawn (PD) and Red) were collected from Kaipara Kumara, Ruawai, Northland, New Zealand in 2018. A total of 5 kg roots of each variety was washed and cut into slices. The diced roots were freeze-dried. Sweetpotato flour (SPF) was obtained by milling the dried slices using a laboratory blender and sieving through a 500 µm mesh screen. Moisture content of SPF was determined using an oven drying method (at 105 °C for overnight). The moisture contents of OS, PD, and Red were 5.63%, 6.68% and 3.54%, respectively. The total starch contents of OS, PD and Red were 52.8%, 61.8% and 66.1% respectively, according to a Megazyme Total Starch Assay Kit (Wicklow, Ireland). Pancreatin (30,000 BPU/g), invertase (300 units/mg), gallic acid, Trolox, β-carotene, chlorogenic acid, caffeic acid, and cyanidin-3-glucoside were purchased from Sigma-Aldrich (St. Louis, USA). Amyloglucosidase (200 U/mL) was purchased from Megazyme (Wicklow, Ireland). All solvents used in this study were of analytical or HPLC grade.

2.3.4. Differential scanning calorimetry for thermal properties 2.3.4.1. Gelatinization. A Q1000 differential scanning calorimeter (DSC) (TA Instruments, New Castle, DE, USA) was used to determine the thermal properties of the SPF, following the method described by Li and Zhu (2018). Flour (3.5 mg, dry basis (db)) was weighed into an aluminum pan and mixed with water at a ratio of 1:3 (w/w). The pan was sealed and equilibrated for 30 min at room temperature. The heating process was conducted between 10 and 90 °C at 10 °C/min. An empty pan was used as reference for all measurements. For each endothermic peak, the onset (To), peak (Tp), and conclusion (Tc) temperatures and enthalpy change (ΔHgel) were recorded. The temperature range (ΔT) was calculated as the difference between Tc and To. The degree of gelatinization (DG) was calculated using the following equation:

2.2. High hydrostatic pressure (HHP) treatment of SPF SPF suspensions were prepared at a flour-to-water ratio of 1:4 in plastic bags and vacuum-sealed. This ratio was determined using a preliminary test. The packed bag was stored at 4 °C overnight for hydration. To prevent sedimentation of SPF suspensions, each bag was shaken several times before HHP treatment. The HHP treatment was carried out with a QFP 2L-700 high pressure unit (Avure Technologies Inc., Middletown, OH, USA). The packed bags were treated at 200, 400, 500 and 600 MPa for 15 min. Distilled water was used as the pressuretransmitting medium. The pressurization rate was approximately 6 MPa/s, and the depressurization time was almost instantaneous. Pressure, time and temperature were monitored by a computer program attached to the HHP unit. The experimental conditions of HHP operation at 600 MPa was given as an example (Supplementary Fig. 1). Due to compressive heating, an increase of up to a maximum of 6 °C was observed during HHP. This change was transient. The samples were equilibrated at 25 °C during holding period, so that SPF properties would not be affected. After HHP treatment, the samples (12 in total) were stored at −80 °C and freeze-dried on the following day. The HHPtreated samples were ground to powder, sieved through a 500 µm sieve and stored in Snap Lock resealable polyethylene bags (Glad, Oakland, CA, USA) until use. The native samples without pressure treatment were also freeze-dried for comparative purposes.

DG = [(ΔHns − ΔHps )/ΔHns] × 100% where ΔHns and ΔHps are the enthalpy changes in native and HHPtreated samples, respectively. 2.3.4.2. Retrogradation. The gelatinized flour was refrigerated (4 °C) for 30 days, and then heated under the same conditions as those used for gelatinization. The degree of retrogradation (DR) was calculated as the ratio of ΔHret to ΔHgel. 2.3.5. α-Amylase activity and pasting properties The α-amylase activity of SPF was determined using a Megazyme αAmylase Assay Kit (Wicklow, Ireland). The pasting properties of SPF were measured by an MCR 301 rheometer (Anton Paar, Graz, Austria) according to the method described by Li and Zhu (2017). Flour (3 g, db) was mixed evenly with 20 mL water in a 50 mL centrifuge tube before transferring into the sample canister. Five steps were involved in the pasting event: (1) equilibration at 50 °C for 5 min; (2) heating from 50 to 95 °C at a rate of 6 °C/min for 7.5 min; (3) equilibration at 95 °C for 5 min; (4) cooling from 95 to 50 °C at a rate of 6 °C/min for 7.5 min; (5) equilibration at 50 °C for 1 min. Pasting parameters included pasting temperature (Ptemp), pasting time (Ptime), peak viscosity (PV), hot paste 2

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100 g of flour db.

viscosity (HPV), final viscosity (FV), breakdown (BD = PV–HPV) and setback (SB = FV–HPV) were recorded.

2.4.3. Total phenolic content and in vitro antioxidant activity The extracts from Section 2.4.1 were used for total phenolic content (TPC) measurement according to the method described by Zhu et al. (2010). The TPC was calculated from a calibration curve using gallic acid (concentrations from 10 to 150 µg/mL) as the standard. The results were expressed as mg gallic acid equivalents (GAE)/100 g of flour db. The same extracts were analyzed for in vitro antioxidant activity using DPPH and FRAP assays based on the method described by Zhu et al. (2010). External calibration curves were made with Trolox (concentrations from 10 to 400 µM for both assays). The results were expressed as mmol Trolox equivalents (TE)/100 g of flour db.

2.3.6. In vitro starch digestibility The in vitro starch digestibility of uncooked and cooked SPF was measured according to Englyst, Kingman, and Cummings (1992) with modifications. Flour (400 mg, db) was mixed with 8 mL of water in a 50 mL centrifuge tube. For the analysis of cooked samples, the suspension was boiled for 30 min with constant stirring and then cooled down with tap water. The centrifuge tubes containing samples were equilibrated in a 37 °C water bath for 5 min. Then, sodium acetate buffer (10 mL, 0.1 M) and 5 mL enzyme mixture solution (pancreatin, amyloglucosidase, and invertase at a ratio of 27:3:2, volume basis) were added. The sample-enzyme mixture was incubated in a shaking water bath at 37 °C for 2 h. The mixture was centrifuged at 1500 × g for 3 min every 20 min. The supernatant (0.1 mL) was mixed with 80% ethanol to deactivate the enzymes. The glucose content in the sample was measured by a Megazyme GOPOD reagent (Wicklow, Ireland). The contents of rapidly digestible starch (RDS), slowly digestible starch (SDS) and resistant starch (RS) were calculated using the equations below:

2.4.4. Analysis of phenolic compounds 2.4.4.1. Quantitative analysis by HPLC. The quantitative analysis of phenolic acids was carried out using the same HPLC system and column as used in Section 2.4.2. The mobile phases were 1% acetic acid (A) and 100% acetonitrile (B). The gradients were 0–40 min, 5–30% B; 40–45 min, 30–100% B; isocratic for 5 min; 50–55 min, 100–50% B; and 55–60 min, 50–5% B. Anthocyanins were detected using the same column but with different mobile phases (1% acetic acid (A) and 100% methanol (B)). The gradients were 0–18 min, 20–70% B; 18–20 min, 70–100% B; 20–23 min, 100–50% B; and 23–25 min, 50–20% B. The injection volume was 10 µL, and the flow rate was 0.4 mL/min for the analysis of both phenolic acids and anthocyanins. The wavelengths of 320 and 520 nm were used for the analysis of phenolic acids and anthocyanins, respectively. Chlorogenic acid (concentrations from 10 to 250 µg/mL) was used as an external standard to quantify phenolic acids, and their contents were expressed as mg chlorogenic acid equivalents (CAE)/100 g of flour db, whereas the quantitation of anthocyanins was based on cyanidin-3glucoside (concentrations from 5 to 150 µg/mL), and their contents were expressed as mg cyanidin-3-glucoside equivalents (CGE)/100 g of flour db.

RDS = G20 × 0.9 SDS = (G120 − G20) × 0.9

RS = TS − RDS − SDS where G20 and G120 are the contents of glucose released after 20 min and 120 min of hydrolysis, respectively; 0.9 is the conversion factor between glucose content and starch content; and TS refers to total starch content. 2.4. Analysis of bioactive compounds 2.4.1. Extraction of bioactive compounds β-Carotene was extracted as described by Grace, Yousef, Gustanfson, Truong, Yencho and Lila (2014) with modifications. Flour (0.5 g, db) was mixed with 10 mL acetone and 10 mg butylated hydroxytoluene (BHT). BHT is an antioxidant which can minimize the degradation of carotenoids during extraction (Saini & Keum, 2018). The extraction of phenolic compounds and analysis of in vitro antioxidant activities followed a method described by Zhu et al. (2010), with some modifications. Flour (0.4 g, db) was extracted with 10 mL of 90% methanol. The extraction of anthocyanins was done using the method described by Zhu et al. (2010). Briefly, flour (0.5 g, db) was extracted with 5 mL of 80% methanol and 0.25 mL of 5 M HCl. All mixtures were shaken in a SK-O180-PRO shaker (DLAB Scientific Inc., Riverside, CA, USA) for 12 h at room temperature. After shaking, the mixtures were centrifuged at 2600 × g for 5 min. The supernatants were collected for subsequent quantitative analysis. All extraction processes were performed in a dark environment to avoid light degradation of bioactive compounds.

2.4.4.2. Identification of polyphenols by HPLC-ESI-TOF-MS/MS. The identification of phenolic acids and anthocyanins was performed using a Bruker MicrOToF-QII instrument (Bruker Daltonics, Bremen, Germany). This instrument was coupled with a Dionex Ultimate 3000 HPLC system (Thermo Fisher Scientific, Sunnyvale, CA, USA) and a KD Science Syringe pump (KD Scientific Inc., Holliston, MA, USA). The same column and chromatographic conditions as described above were used. The orthogonal time-of-flight mass spectrometer (TOF-MS) was set up at negative mode for phenolic acids and positive mode for anthocyanins; the capillary temperature was 350 °C; the collision energy was 6.5–20 eV; the capillary voltage was 4.0 kV. The nebulizing gas was nitrogen with pressure of 50 psi and 10 L/min flow rate. The mass spectrometer scanned from m/z 130–1000 for phenolic acids and from m/z 260–1200 for anthocyanins. The accuracy for confirmation of tentative compound was calculated by the mass error between the theoretical and actual experimental mass peak data. The identification of the polyphenols was based on matching retention times and fragment patterns to the mass spectrum database of polyphenols from the existing literature (Grace et al., 2014; Jung, Lee, Kozukue, Levin, & Friedman, 2011; Wang et al., 2017; Xu et al., 2015; Zhu et al., 2010).

2.4.2. Quantification of β-carotene content The β-carotene content was determined using the modified method described by Grace et al. (2014). The sample extract (3 mL) was blown dry using nitrogen gas and reconstituted with methanol (3 mL). The mixture was filtered using a 0.22 µm syringe-driven filter. The quantification of β-carotene was carried out using an Agilent 1100 HPLC (high performance liquid chromatography) (Agilent Technologies, Wilmington, DE, USA) equipped with a diode array detector (DAD). The βcarotene was separated at 30 °C on a Discovery C18 reverse phase column (25 cm × 4.6 mm) (Sigma-Aldrich Chemical Co., St Louis, USA). The mobile phase was a methanol and acetonitrile mixture (9:1, v/v). The flow rate was 1.5 mL/min, and the injection volume was 20 µL. The detection wavelength was 450 nm. An external β-carotene standard (concentrations from 5 to 200 µg/mL) was used to quantify the level of β-carotene in the samples. The results were expressed as mg/

2.5. Statistical analysis All experiments were done in triplicate. Data were reported as mean ± standard deviation. Statistical analysis were done using SPSS software (Version 25.0, IBM Corp., Armonk, NY, USA). The results were subjected to one-way analysis of variance (ANOVA) using Duncan’s test (p < 0.05). 3

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3. Results and discussion

Song, & Jane, 2002; Li et al., 2015). The increases in To by HHP suggested that some weaker structural regions of starch in SPF were damaged and the more intact part was not affected by HHP (Li et al., 2015). Sugars also compete with starch for water, increasing the gelatinization temperatures (Kohyama & Nishinari, 1991). The gelatinization temperature range (ΔT) decreased with increasing pressure treatment. An increase in pressure level led to a decrease in enthalpy change of gelatinization (ΔHgel). The degree of gelatinization (DG) increased with increasing pressure. Partial gelatinization was observed in OS (70.5%) at 600 MPa, whereas PD and Red were completely gelatinized (100%) at the same pressure level. Thus, starches of PD and Red seemed to be more susceptible to HHP than that of OS. The molecular structure of the starches in the 3 sweetpotato varieties was studied previously (Zhu & Xie, 2018b). The starches in the 3 varieties had C-type polymorph. PD and Red amylopectin had a higher amount of fingerprint Achains (Afp) than OS amylopectin. These chains are not able to form double helices. They may cause structural defects in starch granules, making PD and Red starches more sensitive to HHP (Li & Zhu, 2018). ΔHgel of starch in native SPF samples (ΔHs) (PD and Red) were higher than that of the isolated starches (Zhu & Xie, 2018a). The difference suggested that another endothermic process (e.g., protein aggregation and/or denaturation) occurred together with starch gelatinization (Ahmed, Ramaswamy, Ayad, Alli, & Alvarez, 2007; Li & Zhu, 2017). Katopo et al. (2002) observed that the DSC thermogram had an additional peak right after starch was pressurized at 690 MPa. However, such additional peak was not observed in our study (Supplementary Fig. 5). The difference may be due to different drying methods (oven drying vs freeze drying) immediately after HHP treatment. The freeze drying used in this study may lead to much less retrogradation of gelatinized starch than the oven drying (Katopo et al., 2002; Kim, Kim, & Baik, 2012). The degree of retrogradation (DR) started to increase when the pressure was > 500 MPa for OS and PD, while the DR of Red was not affected by they pressure level. The enthalpy change of retrograded gels (ΔHret) for all the 3 varieties was insignificantly affected by HHP. Thus, the increase of DR was directly related to the change of ΔHgel. Lower ΔHgel resulted in higher DR. Another possible explanation could be the different amylose/amylopectin contents of starch in SPF. In general, there are two processes in starch retrogradation. One is related to the short-term amylose gelation through helix aggregation. The other one is the long-term amylopectin recrystallization (Miles, Morris, Orford, & Ring, 1985). The starches in the three varieties contained different amylopectin fractions (Zhu & Xie, 2018b). Our study focused on longterm amylopectin retrogradation (30 days). Thus, the results might be explained by the difference in the second process, which is largely dependent on the amylopectin structure of starch.

3.1. Morphology of SPF affected by HHP Starch granules in the native SPF were round, oval, and polygonal in shapes with smooth surfaces (Supplementary Fig. 2). The morphology of starch granules agreed with the results of previous studies (Zhu & Xie, 2018b). Other materials such as fibers and proteins also presented along with starch granules in the SPF. At 400 MPa, the shapes of starch in SPF remained intact after HHP. Some minor distortions at the end of starch granules were obtained (e.g., Supplementary Fig. 2B). Pressure of 600 MPa largely destroyed the starch granules. Starch and nonstarch components became infused. At this pressure, the starch in OS only partially gelatinized (Supplementary Fig. 2C), whereas significantly higher degrees of starch gelatinization and starch granule disruptions were obtained with PD and Red (Supplementary Fig. 2F and 2I). 3.2. Particle size of SPF affected by HHP The surface-weighted mean diameter (D [3, 2]), volume-weighted mean diameter (D [4, 3]) and median diameter (d (0.5)) were not significantly affected when pressure was under 500 MPa (Supplementary Table 1). However, these parameters decreased significantly when the pressure reached 600 MPa. The changes in particle size induced by HHP varied among the three varieties. For example, pressure of 600 MPa decreased the D [3, 2] of OS to 31.5 µm, which was around 20% reduction compared with native OS (40.7 µm). Under the same circumstance, 30% and 35% reductions in D [3, 2] were observed in PD and Red, respectively. Most of the SPF samples showed a distinct bimodal particle size distribution, with exceptions of OS treated at 500 MPa and Red treated at 600 MPa (Supplementary Fig. 3). The HHP led to a higher peak of volume for the first peak (particle size < 100 µm) and a lower peak of volume for the second peak (particle size > 100 µm). A significant reduction of particle size was obtained when the pressure reached 600 MPa. 3.3. Fourier transformed infrared spectroscopy (FTIR) of SPF The native and HHP-treated SPF had similar major peaks of the FTIR spectra (Supplementary Fig. 4). HHP treatment affected the intensity of the peaks due to the changes in molecular bonding of the components. A broad range in the 3515–3130 cm−1 region is the stretching of OH bonds (Kizil, Irudayaraj, & Seetharaman, 2002). The spectra of native SPF had a wide base and round peak there due to the presence of water molecules. The change of intensity in this region could be related to increasing water holding capacity of HHP-treated SPF at 600 MPa (Kizil et al., 2002). The peak observed in the region of 3000–2800 cm−1 could be related to CH bond stretching. The intensity changes in this range were attributed to the variations in the amount of amylose and amylopectin present in SPF (Kizil et al., 2002). A major change (reduced peak height) in the secondary structure of proteins in SPF treated at 600 MPa was observed in the region of 1700–1500 cm−1 (Kizil et al., 2002). The band intensities from 1200 to 900 cm−1 are sensitive to starch gelatinization (Rubens, Snauwaert, Heremans, & Stute, 1999). Within this range, the starches in PD and Red displayed higher extents of gelatinization compared to starch in OS.

3.5. α-Amylase activity and pasting properties of SPF affected by HHP α-Amylase is a special class of protein. HHP affects hydrogen bonds of proteins which may lead to the disruption of molecular structure. This may finally result in enzyme inactivation (Rastogi, Raghavarao, Balasubramaniam, Niranjan, & Knorr, 2007). Indeed, the HHP treatment significantly (p < 0.05) decreased the α-amylase activity in the SPF (Table 2). The pasting temperature (Ptemp) and pasting time (Ptime) of OS and PD did not change at the pressures up to 500 MPa, but increased when the pressure reached 600 MPa (Supplementary Fig. 6). The higher Ptemp of HHP-treated SPF indicated that more associating forces inside the starch granules were present after HHP treatment, which also caused higher Ptime for HHP-treated samples (Li et al., 2015). In contrast, both parameters for Red were not affected by HHP. The increase in Ptemp of the three varieties corresponded well with the changes in the gelatinization temperatures measured by DSC. HHP tended to increase the viscosity of SPF as pressure increased up to 500 MPa. Then the viscosity decreased at 600 MPa. For example, OS treated at 500 MPa had significantly higher peak viscosity (PV)

3.4. Thermal properties of SPF affected by HHP HHP up to 400 and 500 MPa had little effect on the onset (To), peak (Tp) and conclusion (Tc) temperatures of PD and the other two SPFs (OS and Red) (Table 1). Increasing HHP from 500 to 600 MPa increased To from 64.2 to 69.9 °C in OS, whereas increasing HHP from 400 to 500 MPa increased To from 56.5 to 59.8 °C in PD. An increase in To has also been reported in potato and red adzuki bean starches (Katopo, 4

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Table 1 Thermal properties of native and HHP-treated SPF. Sample

Gelatinization To (°C)

Tp (°C)

Tc (°C)

Retrogradation

ΔT (°C)

ΔHgel (J/g)

ΔHs (J/g)

DG (%)

ΔHret (J/g)

DR (%)

14.4 ± 0.0a 14.2 ± 0.5a 12.6 ± 0.0b 10.2 ± 0.1c 4.3 ± 0.3d

0.0 ± 0.0d 0.0 ± 0.0d 12.8 ± 0.2c 29.1 ± 0.8b 70.5 ± 1.7a

1.62 1.54 1.74 1.69 1.61

± ± ± ± ±

0.12ab 0.06b 0.06a 0.02ab 0.03ab

21.3 20.6 26.2 31.4 71.5

± ± ± ± ±

1.5c 1.5c 0.8bc 0.0b 0.3a

OS Native 200 MPa 400 MPa 500 MPa 600 MPa

61.5 64.1 64.1 64.2 69.9

± ± ± ± ±

0.9b 0.6b 2.5b 0.4b 0.2a

72.2 72.6 72.5 72.6 74.0

± ± ± ± ±

0.0b 0.3b 0.6b 0.1b 0.3a

78.1 77.6 77.0 77.3 78.3

± ± ± ± ±

0.4a 0.1ab 0.6b 0.3ab 0.5a

16.6 ± 0.5a 13.5 ± 0.5b 12.8 ± 2.0b 13.1 ± 0.1b 8.4 ± 0.3c

7.6 7.5 6.6 5.4 2.3

PD Native 200 MPa 400 MPa 500 MPa 600 MPa

56.3 56.5 56.5 59.8 –

± ± ± ±

0.7b 1.4b 0.7b 0.3a

68.0 69.3 67.9 68.6 –

± ± ± ±

0.4a 0.0a 0.8a 0.7a

73.8 75.2 74.3 74.3 –

± ± ± ±

0.2b 0.6a 0.3ab 0.3ab

17.7 18.8 17.8 14.6 –

11.2 ± 0.1a 11.1 ± 0.0ab 10.5 ± 0.1b 8.8 ± 0.1c –

18.2 18.0 17.0 14.2 –

± ± ± ±

0.1a 0.0ab 0.1b 0.1c

0.0 ± 0.0e 1.2 ± 0.1d 6.7 ± 0.6c 21.7 ± 0.5b 100.0 ± 0.0a

1.38 1.33 1.34 1.49 1.46

± ± ± ± ±

0.11a 0.00a 0.04a 0.08a 0.02a

12.3 12.0 12.8 16.9 –

± ± ± ±

1.0b 0.0b 0.3b 1.1a

Red Native 200 MPa 400 MPa 500 MPa 600 MPa

66.1 66.6 67.2 67.5 –

± ± ± ±

0.1a 1.1a 0.4a 0.2a

71.5 71.7 71.7 71.5 –

± ± ± ±

0.3a 0.1a 0.0a 0.1a

76.2 75.5 75.5 75.2 –

± ± ± ±

0.4a 0.6a 0.2a 0.4a

10.1 ± 0.5a 8.9 ± 1.7a 8.3 ± 0.6a 7.8 ± 0.6a –

10.5 ± 0.5a 9.8 ± 0.1ab 9.4 ± 0.0bc 8.6 ± 0.2c –

15.9 14.8 14.2 13.1 –

± ± ± ±

0.8a 0.2ab 0.0bc 0.4c

0.0 ± 0.0e 6.7 ± 1.1d 10.7 ± 0.1c 17.6 ± 2.3b 100.0 ± 0.0a

1.39 1.42 1.45 1.39 1.62

± ± ± ± ±

0.01a 0.01a 0.24a 0.07a 0.37a

13.3 14.6 15.5 16.1 –

± ± ± ±

0.8a 0.1a 2.6a 1.3a

± ± ± ±

0.9ab 2.0a 1.0ab 0.6a

± ± ± ± ±

0.0a 0.3a 0.0b 0.1c 0.1d

To, onset temperature; Tp, peak temperature; Tc, conclusion temperature; ΔT, gelatinization temperature range; ΔHgel, enthalpy change of gelatinization; ΔHs, ΔHgel of starch in flour sample based on starch content; DG, degree of gelatinization; ΔHret, enthalpy change of retrogradation; DR, degree of retrogradation; -, not detectable; values in the same column and of the same flour with the different letters differ significantly (p < 0.05).

(3.47 Pa·s) but with a much a lower BD at 600 MPa (0.73 Pa·s).

(4.18 Pa·s) than native OS (1.40 Pa·s). A significant drop in PV (3.26 Pa·s) was observed at 600 MPa. The initial increase might be associated with decreased α-amylase activity. Another possible explanation could be the presence of dietary fibers in SPF. Cellulose, lignin, hemicellulose, and pectin are the main dietary fibers of SPF (Wang et al., 2016). HHP promotes structural rearrangements in cellulose fibrils, increasing the swelling and water-holding capacities (Figueiredo, Evtuguin, & Saraiva, 2010). The further decrease at 600 MPa could be due to the starch gelatinization as shown by SEM (Li et al., 2015). PD treated at 500 MPa had a higher hot paste viscosity (HPV) (2.91 Pa·s) and final viscosity (FV) (5.18 Pa·s) than native PD (HPV, 2.00 Pa·s; FV, 3.28 Pa·s). HHP also affected the breakdown (BD) and setback (SB) of SPF in the same manner. For example, OS treated at 500 MPa had a higher BD (1.36 Pa·s) than native OS (1.23 Pa·s) but with a lower BD at 600 MPa (1.08 Pa·s). PD treated at 500 MPa had a higher SB (2.28 Pa·s) than native PD (1.28 Pa·s) but with a lower SB at 600 MPa (1.70 Pa·s). Red treated at 500 MPa had a higher BD (4.06 Pa·s) than native Red

3.6. In vitro starch digestibility of SPF affected by HHP Compared with native flour (uncooked), the HHP-treated flour had a higher degree of hydrolysis for all the varieties (Fig. 1). This was due to the disruption of starch granules by HHP (Supplementary Fig. 2), making starch more vulnerable to enzymatic hydrolysis. In general, with increasing pressure level, the rapidly digestible starch (RDS) content of HHP-treated flour increased comparing to native flour, whereas the resistant starch (RS) content of HHP-treated flour slightly decreased until 500 MPa. The slowly digestible starch (SDS) content increased as the pressure level increased, reaching a maximum (27.4% for OS, 38.7% for PD and 38.0% for Red) at 500 MPa. However, there were some variations in the nutritional fraction composition affected by HHP for different varieties. For example, the RS content of OS was unaffected by pressure up to 400 MPa. The same pattern was observed

Table 2 α-Amylase activity and pasting properties of native and HHP-treated SPF. Sample

α-Amylase (CU/g)

Ptemp (°C)

Ptime (min)

PV (Pa·s)

HPV (Pa·s)

FV (Pa·s)

BD (Pa·s)

SB (Pa·s)

OS Native 200 MPa 400 MPa 500 MPa 600 MPa

0.92 0.90 0.86 0.83 0.83

± ± ± ± ±

0.01a 0.01b 0.00c 0.00d 0.00d

70.5 70.5 70.5 70.5 73.6

± ± ± ± ±

0.0b 0.0b 0.0b 0.0b 0.0a

8.50 8.50 8.50 8.50 9.00

± ± ± ± ±

0.00b 0.00b 0.00b 0.00b 0.00a

1.40 2.36 3.66 4.18 3.26

± ± ± ± ±

0.01e 0.04d 0.00b 0.02a 0.04c

0.17 0.86 2.28 2.82 2.19

± ± ± ± ±

0.07d 0.01c 0.02b 0.04a 0.04b

0.24 1.41 3.62 4.57 3.40

± ± ± ± ±

0.01e 0.02d 0.00b 0.04a 0.00c

1.23 1.49 1.39 1.36 1.08

± ± ± ± ±

0.06b 0.02a 0.02a 0.06a 0.08c

0.07 0.54 1.35 1.75 1.22

± ± ± ± ±

0.00e 0.01d 0.02b 0.00a 0.04c

PD Native 200 MPa 400 MPa 500 MPa 600 MPa

0.57 0.54 0.53 0.51 0.51

± ± ± ± ±

0.00a 0.00b 0.02c 0.00d 0.00d

72.0 72.0 72.0 72.0 75.2

± ± ± ± ±

0.0b 0.0b 0.0b 0.0b 0.0a

8.75 8.75 8.75 8.75 9.25

± ± ± ± ±

0.00b 0.00b 0.00b 0.00b 0.00a

3.64 5.17 6.47 6.39 4.30

± ± ± ± ±

0.13d 0.02b 0.16a 0.11a 0.07c

2.00 2.53 2.93 2.91 3.44

± ± ± ± ±

0.06d 0.01c 0.06b 0.03b 0.11a

3.28 4.35 5.21 5.18 5.13

± ± ± ± ±

0.01c 0.03b 0.05a 0.08a 0.07a

1.64 2.64 3.54 3.48 0.87

± ± ± ± ±

0.07c 0.03b 0.11a 0.18a 0.04d

1.28 1.83 2.28 2.28 1.70

± ± ± ± ±

0.06c 0.02b 0.01a 0.01a 0.04b

Red Native 200 MPa 400 MPa 500 MPa 600 MPa

0.12 0.08 0.07 0.06 0.06

± ± ± ± ±

0.01a 0.00b 0.00b 0.00b 0.00b

73.6 73.6 73.6 73.6 73.6

± ± ± ± ±

0.0a 0.0a 0.0a 0.0a 0.0a

9.00 9.00 9.00 9.00 9.00

± ± ± ± ±

0.00a 0.00a 0.00a 0.00a 0.00a

5.27 6.58 6.65 6.79 3.24

± ± ± ± ±

0.06c 0.14b 0.05ab 0.05a 0.03d

1.80 2.72 2.78 2.73 2.52

± ± ± ± ±

0.01c 0.05a 0.02a 0.01a 0.02b

2.95 4.86 5.27 4.91 4.34

± ± ± ± ±

0.01d 0.08b 0.05a 0.01b 0.08c

3.47 3.87 3.87 4.06 0.73

± ± ± ± ±

0.06c 0.09b 0.07b 0.04a 0.01d

1.15 2.14 2.50 2.18 1.82

± ± ± ± ±

0.01d 0.06b 0.02a 0.01b 0.06c

CU, Ceralpha unit; Ptemp, pasting temperature; Ptime, pasting time; PV, peak viscosity; HPV, hot paste viscosity; FV, final viscosity; BD, breakdown (PV – HPV); SB, setback (FV – HPV); values in the same column and of the same flour with the different letters differ significantly (p < 0.05). 5

Food Chemistry 299 (2019) 125129

R. Cui and F. Zhu

D: Cooked OS The RDS, SDS and RS contents (%)

The RDS, SDS and RS contents (%)

A: Uncooked OS 40 a

35 30

b c c

25

a Native d

20 15 10

a a a

200 MPa b

400 MPa

c bc b b

500 MPa

5

c

600 MPa

0 RDS

SDS

40 35

a a a b b

30 25

Native a a a a a

20

400 MPa

10

500 MPa a b b abab

5 RDS

35

d c

b

a

30

25 15

10

Native

e a

20 d d

c

The RDS, SDS and RS contents (%)

The RDS, SDS and RS contents (%)

a

b

200 MPa

b

c

400 MPa d

500 MPa e

5

600 MPa

0 RDS

SDS

RS

40

a a b b b

35 30

a

25

400 MPa

15

10 5

b

0 RDS

25 20 15 10

e

c d

The RDS, SDS and RS contents (%)

The RDS, SDS and RS contents (%)

a

c

a

Native

b b

b

200 MPa c

400 MPa d

5

500 MPa 600 MPa

0 RDS

SDS

SDS

a a a a

500 MPa 600 MPa

RS

F: Cooked Red

b b b

30

200 MPa

20

C: Uncooked Red a

Native

b ababab

Starch fractions

45 35

RS

E: Cooked PD 45

Starch fractions

40

SDS

Starch fractions

B: Uncooked PD 40

600 MPa

0

RS

Starch fractions

45

200 MPa

15

RS

Starch fractions

40 35

a b b b c a a a a a

30 25

Native

20

200 MPa

15

400 MPa

10

500 MPa a ab b ab ab

5

600 MPa

0 RDS

SDS

RS

Starch fractions

Fig. 1. The contents of rapidly digestible starch (RDS), slowly digestible starch (SDS) and resistant starch (RS) fractions of different SPF samples affected by HHP treatments. (A): uncooked OS; (B): uncooked PD; (C): uncooked Red; (D): cooked OS; (E): cooked PD; (F): cooked Red. Values with different letters in the same starch fraction are significantly different (p < 0.05).

The digestibility of starch in SPF is important for nutritional and food applications. The in vitro starch digestibility of HHP-treated SPF in the uncooked condition increased with increasing of pressure level (Fig. 1). This could be explained by high pressure induced starch gelatinization which made the HHP-treated samples more susceptible to enzyme hydrolysis (Yang et al., 2017). This is reflected by the higher RDS content and lower SDS content in the HHP-treated samples than native samples.

in the SDS content of Red. When the pressure level reached 600 MPa, the content of RDS markedly increased to 32.6%, 35.4%, and 35.4% for OS, PD and Red, respectively. Meanwhile, the SDS and RS contents were significantly decreased. SDS and RS (Type 2) are the major starch fractions of uncooked SPF (Fig. 1A–C). The significant decrease of SDS and RS at 600 MPa could be attributed to the structural modification and rupture of starch granules (SDS and RS) induced by HHP. For cooked samples, as the pressure level increased, the RS content increased and reached a maximum (2.6% for OS, 3.7% for PD and 3.7% for Red) at 600 MPa, while the RDS content decreased gradually (Fig. 1D–F). The higher RS content was mainly attributed to the cooked samples treated at HHP. These cooked and HHP-treated samples had increased retrogradation (Table 1), leading to increased RS (Type 3) fraction. Moreover, the native and HHP-treated and cooked SPF had a relatively low content of SDS which was little affected by increasing pressure except for the SDS content of PD at 200 MPa.

3.7. β-Carotene content, total phenolic content, and in vitro antioxidant activity of SPF affected by HHP β-Carotene contents of OS and PD were not significantly affected by the pressure between 200 and 500 MPa (Table 3). However, pressure treatment at 600 MPa significantly increased the extractability of βcarotene from OS and PD by 50 and 52%, respectively, compared to the 6

7

– – – – – – – – – – – – – – – 5.7 ± 0.0b 3.5 ± 0.0e 4.1 ± 0.0d 4.8 ± 0.1c 12.4 ± 0.1a tr tr tr tr tr – – – – – 6.4 ± 0.0e 20.2 ± 0.0d 21.0 ± 0.1c 22.5 ± 0.2b 25.4 ± 0.1a – – – – tr 0.09a 0.00e 0.03d 0.08c 0.06b ± ± ± ± ± 9.83 4.85 5.40 6.13 9.44 – tr tr tr tr

3-CQA, 3-caffeoylquinic acid; 5-CQA, 5-caffeoylquinic acid; 4-CQA, 4-caffeoylquinic acid; CA, caffeic acid; FQA, feruloylquinic acid; 4,5-diCQA, 4,5-dicaffeoylquinic acid; 3,5-diCQA, 3,5-dicaffeoylquinic acid; 3,4-diCQA, 3,4-dicaffeoylquinic acid; CFQA, caffeoyl-feruloylquinic acid; diCFQA, dicaffeoyl-feruloylquinic acid; the content of HAD are expressed as mg chlorogenic acid equivalents (CAE)/100 g of flour db; tr, trace amount; -, not detectable; values in the same column and of the same flour with different letters differ significantly (p < 0.05).

± ± ± ± ± 22.9 30.3 32.4 35.7 51.9

1.9e 0.6d 3.4b 0.7c 1.9a ± ± ± ± ± 270.5 322.5 349.8 341.9 404.5 0.1c 0.0b 0.3a 0.0a 0.4a ± ± ± ± ± 17.3 17.9 21.2 20.8 20.9 0.04d 0.02e 0.04b 0.01c 0.03a ± ± ± ± ± 5.06 4.23 5.32 5.17 6.25 12.7 ± 0.3b 9.7 ± 0.0d 11.6 ± 0.2c 11.6 ± 0.1c 13.6 ± 0.3a 82.7 ± 0.7d 76.6 ± 0.1e 90.3 ± 0.5b 88.9 ± 0.2c 131.1 ± 0.7a 0.23c 0.30c 0.21b 0.08b 0.21a ± ± ± ± ± 5.68 5.78 6.56 6.38 7.94 0.1d 0.0e 0.2b 0.0c 0.1a ± ± ± ± ± 14.6 14.1 16.0 15.7 17.4 24.7 ± 0.5e 101.4 ± 0.1a 94.1 ± 0.5b 88.6 ± 0.0c 86.6 ± 0.3d 0.06c 0.02b 0.17a 0.02b 0.03a ± ± ± ± ± 1.79 2.55 2.82 2.52 2.99 100.8 ± 0.4b 79.7 ± 0.1d 91.3 ± 1.1c 92.2 ± 0.0c 105.7 ± 0.4a

± ± ± ± ± 20.9 22.9 21.7 22.7 27.5 8.6 ± 0.0b 8.8 ± 0.0b 8.6 ± 0.1b 8.9 ± 0.1b 11.7 ± 0.2a 0.03c 0.03bc 0.06bc 0.01b 0.30a ± ± ± ± ± 2.84 3.07 3.13 3.29 4.82

Red Native 200 MPa 400 MPa 500 MPa 600 MPa

A total of 10 different phenolic acids (mainly hydroxycinnamic acid derivatives (HAD)) were identified (Supplementary Fig. 7 and Supplementary Table 2; the profile of OS treated at 600 MPa is given as an example). Peaks 2 and 4 were identified as 5-caffeoylquinic acid (5-

5.3 ± 0.0c 10.5 ± 0.1b 10.7 ± 0.3b 10.1 ± 0.4b 12.0 ± 0.2a

3.8. Phenolic acid profile of SPF affected by HHP

PD Native 200 MPa 400 MPa 500 MPa 600 MPa

Table 4 Contents of hydroxycinnamic acid derivatives (HAD) in native and HHP-treated SPF.

FQA

± ± ± ± ±

0.0b 0.1d 0.3d 0.2c 0.0a

native samples. The total phenolic content (TPC) increased with increasing pressure level (Table 3). All treated samples showed a significant increase in TPC at 600 MPa. For example, the TPC of OS treated at 600 MPa (1956 mg GAE/100 g of flour db) was much higher than that of native OS (1603 mg GAE/100 g of flour db). Among the different varieties, the highest increase in TPC at 600 MPa was observed in Red (42%), followed by PD (23%) and OS (22%). Increasing pressure level also increased the in vitro antioxidant activity of SPF. For example, HHP significantly increased the antioxidant activity measured using DPPH assay by 4.2% for OS, 13.1% for PD, and 30.9% for Red at 600 MPa compared to the native samples. Likewise, the increase in antioxidant activity when pressure reached 600 MPa was observed in that measured by the FRAP assay, though a higher percentage of increase was observed from this assay compared with that from the DPPH assay. Significant increments in TPC and in vitro antioxidant activities were observed (Table 3). HHP improves extractability of bioactive compounds due to the disruption of plant cell wall. Solvents have higher accessibility to these bioactive compounds and more compounds are released from the damaged cell (Oey et al., 2008). Even the cellular structure is completely disrupted, the presence of other food matrix components and related interactions may affect the bioavailability of nutrients. For example, the presence of dietary fibers could decrease carotenoid absorption (Palafox-Carlos, Ayala-Zavala, & GonzálezAguilar, 2011). Since carotenoids are hydrophobic, their absorption also depends on the solubilization by bile salts and related digestive enzymes (Palafox-Carlos et al., 2011). Therefore, the ingestion of dietary lipids is recommended to improve the bioavailability of carotenoids in the food systems.

± ± ± ± ±

0.0d 0.0b 0.2c 0.2b 0.0a

1035.7 1010.3 1003.7 1034.1 1356.9

Total HAD 4,5-diCQA

db, dry basis; TPC, total phenolic content; DPPH, free radical scavenging activity against 2,2-diphenyl-1-picrylhyl radicals; FRAP, ferric reducing ability of plasma; GAE, gallic acid equivalents; TE, Trolox equivalents; –, not detectable; values in the same column and of the same flour with the different letters differ significantly (p < 0.05).

1.8e 0.6d 0.2c 0.4b 0.9a

0.01c 0.01bc 0.01bc 0.01b 0.02a

± ± ± ± ±

± ± ± ± ±

412.0 415.1 421.5 432.0 653.6

0.51 0.53 0.53 0.55 0.68

9.6 ± 0.1b 9.9 ± 0.0b 9.5 ± 0.3b 10.1 ± 0.2b 13.1 ± 0.4a

0.01c 0.01c 0.01c 0.05b 0.02a

24.5 22.9 22.6 23.5 27.0

± ± ± ± ±

5c 8b 5b 11b 2a

0.0a 0.0b 0.5c 0.4d 0.7e

0.42 0.42 0.42 0.49 0.55

± ± ± ± ±

± ± ± ± ±

288 349 361 376 408

61.1 56.2 53.8 52.1 51.4

0.06c 0.09b 0.06a 0.07a 0.02a

0.1b 0.6ab 0.0b 0.7ab 0.8a

± ± ± ± ±

± ± ± ± ±

2.77 3.14 3.45 3.47 3.49

13.1 12.8 11.4 12.0 13.4

0.08b 0.02b 0.09a 0.01a 0.01a

1.1b 0.2d 2.0e 1.4c 3.6a

± ± ± ± ±

± ± ± ± ±

2.52 2.57 2.76 2.87 2.85

451.7 428.3 422.1 438.8 519.0

906 ± 3c 1030 ± 10b 1088 ± 15a 1114 ± 9a 1117 ± 3a

0.1b 0.1c 0.3d 0.2c 0.3a

0.70b 0.34b 0.08b 0.13b 0.24a

± ± ± ± ±

± ± ± ± ±

31.3 30.0 29.3 30.1 35.6

7.34 7.43 7.42 7.59 8.99

OS Native 200 MPa 400 MPa 500 MPa 600 MPa

0.01b 0.01b 0.02b 0.01b 0.01a

diCFQA

– – – – –

1b 2b 1b 1b 3a

± ± ± ± ±

CFQA

Red Native 200 MPa 400 MPa 500 MPa 600 MPa

± ± ± ± ±

3.08 3.07 3.06 3.07 3.21

3,4-diCQA

40 39 41 40 61

3c 12b 9b 6b 7a

± ± ± ± ±

3,5-diCQA

PD Native 200 MPa 400 MPa 500 MPa 600 MPa

8c 5b 2b 6b 3a

CA

1603 1640 1651 1629 1956

± ± ± ± ±

FRAP (µmol TE/ 100 g of flour db)

4-CQA

560 753 750 756 842

DPPH (µmol TE/ 100 g of flour db)

5-CQA

OS Native 200 MPa 400 MPa 500 MPa 600 MPa

In vitro antioxidant activity

3-CQA

TPC (mg GAE/ 100 g of flour db)

Sample

β-Carotene (mg/100 g of flour db)

Sample

3.1b 1.4c 3.6c 1.4b 7.3a

Table 3 β-Carotene content, total phenolic content and in vitro antioxidant activity of native and HHP-treated SPF.

0.2e 0.1d 0.1c 0.5b 0.3a

Food Chemistry 299 (2019) 125129

R. Cui and F. Zhu

Food Chemistry 299 (2019) 125129

R. Cui and F. Zhu

Table 5 Contents of anthocyanins (AC) in native and HHP-treated SPF. cy 3-phydroxybenzoylsoph5-glc

pn 3-phydroxybenzoylsoph5-glc

cy 3-(6″feruloylsoph)5-glc

pn 3-(6″feruloylsoph)5-glc

cy 3-(6″caffeoyl-6″′feruloylsoph)-5glc

pn 3-(6″-caffeoyl-6″′-phydroxybenzoylsoph)-5glc

pn 3-(6″caffeoyl-6″′feruloylsoph)-5glc

Total AC

OS Native 200 MPa 400 MPa 500 MPa 600 MPa

2.03 2.02 1.87 1.91 2.16

± ± ± ± ±

0.01b 0.01b 0.01c 0.01c 0.04a

1.89 1.86 1.86 1.89 1.99

± ± ± ± ±

0.16a 0.02a 0.02a 0.10a 0.03a

2.82 2.91 2.87 2.70 3.15

± ± ± ± ±

0.05b 0.03b 0.06b 0.04c 0.04a

5.82 5.90 5.56 5.70 6.02

± ± ± ± ±

0.41a 0.08a 0.09a 0.08a 0.04a

13.8 14.3 13.3 13.6 14.5

± ± ± ± ±

0.3b 0.1a 0.1c 0.0bc 0.1a

23.3 23.9 22.4 22.7 23.9

± ± ± ± ±

0.4b 0.1a 0.1c 0.1c 0.0a

2.69 2.73 2.66 2.77 2.79

± ± ± ± ±

0.33a 0.05a 0.05a 0.11a 0.04a

52.3 53.6 50.5 51.3 54.5

PD Native 200 MPa 400 MPa 500 MPa 600 MPa

1.19 1.29 1.32 1.21 1.56

± ± ± ± ±

0.01c 0.03bc 0.02b 0.07c 0.04a

0.76 0.83 0.83 0.78 0.80

± ± ± ± ±

0.01a 0.01a 0.00a 0.12a 0.14a

10.3 10.4 10.9 10.5 11.3

± ± ± ± ±

0.0c 0.2c 0.0ab 0.4ab 0.6a

0.74 0.64 0.73 0.70 1.00

± ± ± ± ±

0.00a 0.00a 0.17a 0.28a 0.01a

8.58 8.53 8.81 8.86 9.29

± ± ± ± ±

0.14c 0.06c 0.04b 0.04b 0.03a

84.9 83.7 83.6 82.9 80.4

± ± ± ± ±

0.0a 0.0b 0.2b 0.3b 0.0c

7.51 7.43 7.63 8.07 7.77

± ± ± ± ±

0.11c 0.27c 0.11ab 0.29a 0.01ab

114.0 112.7 113.9 113.1 112.1

Red Native 200 MPa 400 MPa 500 MPa 600 MPa

1.10 0.97 0.87 0.96 1.77

± ± ± ± ±

0.00b 0.02c 0.00d 0.01c 0.02a

– – – – –

1.45 1.22 1.29 1.31 2.27

± ± ± ± ±

0.00b 0.01e 0.00d 0.01c 0.01a

– – – – 1.32 ± 0.01

Sample

– – – – –

– – – – –

– – – – –

2.55 2.19 2.15 2.27 5.36

± ± ± ± ±

1.4bc 0.4ab 0.3c 0.0c 0.5a

± ± ± ± ±

± ± ± ± ±

0.1a 0.6a 0.5a 1.6a 1.1a

0.01b 0.02d 0.00d 0.02c 0.02a

cy, cyanidin; soph, sophoroside; glc, glucoside; pn, peonidin; the content of AC are expressed as mg cyanidin-3-glucoside equivalents (CGE)/100 g of flour db; –, not detected; values in the same column and of the same flour with different letters differ significantly (p < 0.05).

with other elements such as vitamins, simple sugars, and carotenoids (Truong, McFeeters, Thompson, Dean, & Shofran, 2007). The presence of phenolic acids in SPF may inhibit starch hydrolysis (Zhu, 2015). Pearson correlation analysis showed that the SDS content negatively correlated with phenolic acids (Supplementary Table 3). HHP at 600 MPa led to a higher phenolic acids content in SPF, resulted in the degradation of the starch fraction (SDS) (Fig. 1A–C). The inhibitory effect of phenolic acids on starch hydrolysis followed the order of p-benzoquinone > chlorogenic acid > gallic acid > caffeic acid > ferulic acid > quinic acid (Rohn, Rawel, & Kroll, 2002). Indeed, our results showed that chlorogenic acid (r = −0.793, p < 0.01) had a higher impact on starch nutritional fractions than that of caffeic acid. Other factors affecting the inhibition of phenolic acids on starch hydrolysis included the type of starch, the specific enzymes used in the model system, and the concentrations of the enzymes, phenolic acids and starch (Zhu, 2015).

CQA) and caffeic acid (CA), respectively. The two peaks eluting before (peak 1) and after (peak 3) 5-CQA were tentatively identified as 3caffeoylquinic acid (3-CQA) and 4-caffeoylquinic acid (4-CQA), respectively (Padda & Picha, 2008). Three parent ions observed at m/z 515 (peak 6–8) were characterized as 4,5-dicaffeoylquinic acid (4,5diCQA), 3,5-dicaffeoylquinic acid (3,5-diCQA) and 3,4-dicaffeoylquinic acid (3,4-diCQA), respectively, according to their retention times found in the literature under similar separation conditions (Grace et al., 2014; Jung et al., 2011). These compounds (peak 1–8) have been reported in sweetpotato samples from China, Korea, the US and Europe (Jung et al., 2011; Padda & Picha, 2008). Peak 9 was identified as caffeoyl-feruloylquinic acid (CFQA), and this compound was reported in the peels of sweetpotato cultivated in Tanzania and China (Zheng & Cliford, 2008). Peak 10 was identified as di-caffeoylferuloylquinic acid (diCFQA) due to the addition of a caffeoyl group (m/z 162) in CFQA. The major HADs in OS and PD were 5-CQA and 3,5-diCQA, whereas CA was the most abundant HAD in Red (Table 4). The contents of most of the HADs significantly increased as a result of increased HHP. For example, after pressurization at 600 MPa, the concentrations of 4,5diCQA, CFQA and diCFQA increased by > 30% in OS compared to the native sample. The concentration of CA at 200 MPa was 4-fold higher in PD than the native sample. Pressurization at 600 MPa induced 31%, 50% and 126% increases in total HADs for OS, PD and Red, respectively. However, HHP also led to a reduction in the contents of some of the HADs in SPF. For example, the CA content of OS treated at 600 MPa (51.4 mg CAE/100 g of flour db) was lower than that in native OS (61.1 mg CAE/100 g of flour db). A decrease in 5-CQA content in PD was observed at 200 MPa (79.7 mg CAE/100 g of flour db) compared with that in native PD (100.8 mg CAE/100 g of flour db). Polyphenoloxidase (PPO) and peroxidase (POD) are the intracellular enzymes in SPF responsible for the decrease of the contents of some phenolic compounds (Oey et al., 2008). The complete inactivation of PPO and POD usually requires HHP (> 600 MPa) in conjunction with a mild thermal treatment (40–50 °C) (Rastogi et al., 2007). The decrease in the content of these phenolic acids may also be affected by their interactions with proteins as protein ionization is favored by HHP (Rastogi et al., 2007). It was noted that the TPC determined by the Folin-Ciocalteu method was much higher than the total HADs quantified by the HPLC method. This may be due to the fact that the FolinCiocalteu reagent can react not only with phenolic compounds but also

3.9. Anthocyanin profile of SPF affected by HHP Seven major peaks related to anthocyanins were observed in the HPLC chromatograph (Supplementary Fig. 8 and Supplementary Table 4; the profile of OS treated at 600 MPa is given as an example). The abbreviations for these anthocyanins were used here (Supplementary Table 4). The anthocyanins detected were cyanidin and peonidin glycosides acylated with phenolic acids such as p-hydroxybenzoic acid, caffeic acid and ferulic acid. For example, the most abundant anthocyanin in OS and PD was pn 3-(6″-caffeoyl-6″′-p-hydroxybenzoylsoph)-5-glc. The precursor ions were detected by the 1st MS after electrospray ionization (ESI) (m/z 1069) and further dissociated by collision energy for the 2nd MS detection (m/z 907, 463, 301). The total anthocyanin content was the highest in PD, followed by OS. The anthocyanin content was significantly less in Red than OS and PD (Table 5). A trace amount of anthocyanins in the peels of whitefleshed sweetpotato root has been reported (Zhu et al., 2010). In general, the individual anthocyanin and total anthocyanin contents had no significant changes after HHP treatment regardless of the pressure level, indicating that anthocyanins were well retained after HHP treatment. Other authors have also demonstrated that anthocyanins were stable during HHP treatment at moderate temperature (Oey et al., 2008). 8

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Declarations of Competing Interest and Funding

Instead, storage time and temperature are the major factors for anthocyanin degradation (Patras, Brunton, O'Donnell, & Tiwari, 2010). Recent research showed that cyanidin based pigments decreased in vitro starch hydrolysis by blocking digestive enzymes and forming crosslinking structure with starch during cooking, limiting the amylolytic attack (Camelo-Méndez, Agama-Acevedo, Sanchez-Rivera, & BelloPérez, 2016; Zhu, 2015). However, HHP had no significant effect on anthocyanin contents in our study. The changes in starch hydrolysis are unlikely related to the presence of anthocyanins. Other factors such as disruption of starch granules and increased content of phenolic acids as described before may take a major responsibility.

The authors declare no conflict of interest. The research did not receive any specific grant. Appendix A. Supplementary data Supplementary data to this article can be found online at https:// doi.org/10.1016/j.foodchem.2019.125129. References

3.10. Significance of HHP processing for food applications: A general discussion

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In recent decades, applications of HHP in the food industry have aimed at reducing the food spoilage, improving shelf life of food products and creating novel functionalities and sensory properties (Rastogi et al., 2007). The effects of HHP on the functionalities of SPF are well demonstrated in this study (Section 2). Pre-gelatinized SPF can be used in food formulations such as in ready-to-eat food products and as thickening agents. The rheological behaviors of HHP-treated SPF (e.g., reduced BD) suggested that this material may be incorporated into the gluten-free bread making to retard bread staling. The FV and SB indicate the retrogradation tendency. The higher FV and SB of SPF treated at 200–500 MPa suggested their tendency towards retrogradation. Other quality attributes, such as hardness, bread specific volume and cohesiveness, remain to be studied. HHP-treated SPF (up to 500 MPa) can be used as an SDS source. The advantages of eating SDS-rich food include controlling glucose metabolism, reducing the risk of diabetes and improving mental performance (Lehmann & Robin, 2007). Nutritionally, HHP enhances the intake of dietary nutrients in humans by the disrupting cell wall of plants. The increased extractability of phenolic acids and stability of anthocyanins in SPF may be beneficial to the gut microbiome and for value-added processing such as fractionation. However, the bioavailability of these bioactive compounds remains to be studied. Finally, HHP is a proven technology to reduce microbiological contamination (Rastogi et al., 2007). Therefore, food formulated with HHP-treated SPF may be expected to have an extended shelf life. 4. Conclusions Full gelatinization of starches in PD and Red was achieved at 600 MPa, while higher pressure was required to completely gelatinize starch in OS. HHP under 500 MPa increased the viscosity of SPF during pasting. At 600 MPa, HHP significantly decreased the viscosity. Particle size of SPF was decreased due to the damage of starch granules caused by HHP. The in vitro digestibility of starch in HHP-treated samples behaved differently in cooked and uncooked conditions. The TPC and in vitro antioxidant activity in the HHP-treated samples increased with increasing pressure. The contents of bioactive compounds were significantly improved at 600 MPa due to the cell membrane disruption caused by HHP. The predominant phenolic acids in OS and PD were 5CQA and 3,5-diCQA, whereas CA was the most abundant phenolic acid in Red. Anthocyanins were well retained in HHP-treated samples. Overall, the selection of sweetpotato varieties, together with the conditions of HHP treatment are important for desired functional and nutritional properties with potential for food product formulations. Acknowledgments The authors wish to thank Mr Anthony Blundell for providing the sweetpotato roots; Mr Peter Buchanan for setting up the high pressure unit; Dr Yuan Tao for helping with the scanning electron microscopy analysis; and Mr Radesh Singh for training in FTIR. 9

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