Δευτέρα 26 Οκτωβρίου 2015

Effects of sprouting and postharvest storage under cool temperature conditions on starch content and antioxidant capacity of green pea, lentil and young mung bean sprouts


doi:10.1016/j.foodchem.2015.03.108
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Highlights

•
Green pea, lentil and mung bean were sprouted and storage under cool condition.
•
Phenolics, antioxidant capacity and starch quality in legume sprouts were studied.
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Reducing potential of bioavailable fraction of stored lentil sprouts was elevated.
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Storage of sprouts significantly elevated values of expected glycemic index.

Abstract

The effects of germination of selected legumes and further storage of sprouts under cool conditions on the phenolics, antioxidant activity and starch content and their potential bioaccessibility were elucidated. In green pea and mung bean sprouts a slight increase of chemically extractable phenolics (including flavonoids) during the first 4 days of sprouting was observed. Digestion in vitro released phenolics; however, flavonoids were poorly bioaccessible. Storage of green pea sprouts decreased reducing power and increased the antiradical ability. Reducing potential of potentially bioaccessible fraction of stored lentil sprouts was elevated of 40%, 31% and 23% in 3-, 4- and 5-day-old sprouts, respectively. Postharvest storage significantly increases the starch digestibility and values of expected glycemic index (eGI) – the highest eGIs were determined for 5-day-old stored sprouts; 75.17-green pea, 83.18-lentil and 89.87-mung bean. Bioactivity and nutritional quality of legumes is affected by sprouting and further storage at low temperatures.

Keywords

  • Sprouts;
  • Low-temperature storage;
  • Antioxidant capacity;
  • Bioaccessibility in vitro;
  • Starch;
  • Expected glycemic index

1. Introduction

Sprouting is a cheap, effective and simple tool useful for improving the nutritional and nutraceutical quality of cereals, pseudocereals, cruciferous and legumes (Cevallos-Casals and Cisneros-Zevallos, 2010, Guo et al., 2011, Pająkk et al., 2014 and Świeca et al., 2012). Germination is a very dynamic process, which causes significant qualitative and quantitative changes in the nutrients and bioactive compounds. These changes are associated with activation the enzymatic pathways involved in energy obtaining (e.g. amylases, proteases), new structures building (the phenylpropanoids pathway, laccase) as well as the metabolism of functional compounds such as hormones, regulators, etc. (Rosental, Nonogaki, & Fait, 2014).
The quality of sprouted food may be created on each step of its production but depends mainly on seeds quality, germination conditions and further storage. The modifications of seeds and/or germination conditions may improve the microbiological quality of sprouts e.g. combined treatments of mung bean with high pressure, temperature and antimicrobial products (Peñas, Gómez, Frías, & Vidal-Valverde, 2010). Additionally, such treatments enhance production of pro-health components eg. an increase of phenolics in lentil sprouts treated with hydrogen peroxide (Świeca & Baraniak, 2014a) or broccoli sprouts treated with yeast and willow bark extracts (Gawlik-Dziki, Świeca, Dziki, & Sugier, 2013) and diversify nutrients content and digestibility e.g. starch and protein digestibility of lentil sprouts by abiotic stress treatments (Świeca and Baraniak, 2014a, Świeca and Baraniak, 2014b, Świeca, Baraniak, et al., 2013 and Gulewicz et al., 2014). On the other hand, one of the key factors affecting sprouts quality is its age. Sprouts are usually consumed fresh; however to inhibit their growth and retain quality (microbial, nutritional and nutraceutical) they are stored at low temperatures (e.g. refrigerator). So far there are no studies about the effect of storage on the nutritional quality of sprouts and only few studies are available concerning the changes of its nutraceutical quality (Force et al., 2007, Goyal et al., 2014, Song and Thornalley, 2007 and Świeca, Surdyka, et al., 2014); however, in these studies there is no simple relationship between these determinants and sprouts age, germination conditions as well as time and storage conditions.
Biological activity of phytochemicals and nutritional potential of sprouts is strongly affected by their metabolic fate, including bioaccessibility and bioavailability (study of chemical extracts does not always mirror the real activity in vivo; however, it provides valuable information about mechanism of phytochemicals action). Accordingly, for evaluation of nutritional and nutraceutical quality of sprouts using the systems determining potential bioaccessibility (conditions simulating those observed during the gastrointestinal digestion) is very important. Digestive tract usually acts as an effective extractor realizing bioactive compounds from food matrix ( D’Archivio et al., 2007 and Gawlik-Dziki et al., 2012); however, these phytochemicals may interact with nutrients/enzymes, thus limiting nutritional potential ( Świeca, Sęczyk, and Gawlik-Dziki, 2014 and Świeca, Gawlik-Dziki, et al., 2013).
Oxidative damage, caused by the excess of reactive oxygen and nitrogen species, is proved to be the cause of many disorders such as cancer, diabetes, and inflammation. Thus, the fundamental property of phenolic food compounds is antioxidant activity, that is important for health protection (Gawlik-Dziki et al., 2012 and Rajendran et al., 2014). A very important role in dietary prevention, specially visible in case of diabetes and Alzheimer disease, plays also nutrients (their level and quality) (Rajendran et al., 2014). In this study, the effects of germination and further storage of sprouts under cool temperature conditions on the phenolic compounds (strong antioxidants) and starch contents (component creating glycemic response) were elucidated. Furthermore, the potential bioaccessibility and antioxidative activity of these compounds, the in vitro digestibility of starch and expected glycemic index (eGI) were evaluated.

2. Material and methods

2.1. Chemicals

ABTS (2,2-diphenyl-1-picrylhydrazyl), α-amylase (E.C. 3.2.1.1), pancreatin, gallic acid, quercetin, amyloglucosidase (EC 3.2.1.3), ammonium thiocyanate, Trolox (6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid), invertase (EC 3.2.1.26), pepsin A (EC 3.4.2.3.1) were purchased from Sigma–Aldrich company (Poznan, Poland). All others chemicals were of analytical grade.

2.2. Material

Green pea, lentil and mung bean seeds were purchased from the PNOS S.A. in Ozarów Mazowiecki, Poland. Seeds were sterilized in 1% (v/v) sodium hypochlorite for 10 min, then drained and washed with distilled water until they reached neutral pH. They were placed in distilled water and soaked for 6 h at 25 °C. Seeds (approximately 150 per plate) were dark-germinated 6 days in a growth chamber (SANYO MLR-350H) on Petri dishes (ϕ 125 mm) lined with absorbent paper (relative humidity 70%, 25 °C). Seedlings were watered daily with 5 ml of Milli-Q water. Sprouts from subsequent days of cultivation (1–6 day-old; 1F–6F, respectively) were manually collected, rapidly frozen, lyophilized, grounded in a labor mill, sieved (60 mesh) and kept in polyethylene bags at −20 °C. For postharvest storage experiment ready-to-eat sprouts (3-, 4- and 5-day-old fresh sprouts) were manually collected and kept at polypropylene boxes at 4 °C for 7 days (3S, 4S and 5S, respectively). After 1 week storage, sprouts were collected from boxes, rapidly frozen, lyophilized, grounded in a labor mill, sieved (60 mesh) and kept in polyethylene bags at −20 °C.

2.3. Extraction procedures

2.3.1. Chemical extraction (CE)

For chemical extraction (CE) sprouts (200 mg of dry mass (d.m.)) were extracted with 5 ml 60 mM HCl in 70% acetone (v:v:v) for 1 h at room temperature (22 °C ± 2 °C), centrifuged (15 min, 3000×g, 22 °C) and the supernatants were recovered. The procedures were repeated and the supernatants combined ( Xu & Chang, 2007).

2.3.2. Digestion in vitro

For simulated mastication and gastrointestinal digestion sprouts (200 mg of d.m.) were homogenized in 3.5 mL of simulated salivary fluid (2.38 g Na2HPO4, 0.19 g KH2PO4 and 8 g NaCl, 200 U α-amylase (E.C. 3.2.1.1. in 1 L H2O, pH −6.75) and shaken for 10 min at 37 °C. Next, the samples were adjusted to pH 1.2 with HCl (5 mol/L), suspended in 1.25 mL of simulated gastric fluid (300 U/mL of pepsin A, EC 3.4.2.3.1 in 0.03 M HCl, pH 1.2) and shaken for 120 min. at 37 °C. After simulated gastric digestion, samples were adjusted to pH 6 with 0.1 mol/L NaHCO3 and suspended in simulated intestinal juice (0.05 g of pancreatin (activity equivalent 4 × USP) and 0.3 g of bile extract in 2.0 mL 0.1 mol/L NaHCO3; adjusted to pH 7 with 1 mol/L NaOH and finally 1.25 mL of 120 mmol/L NaCl and 5 mmol/L KCl was added to the sample. The prepared samples underwent in vitro intestinal digestion for 120 min ( Świeca, Baraniak, et al., 2013).

2.4. Determination of phenolics content

2.4.1. Determination of total phenolic compounds (TPC)

The amount of total phenolics was determined using Folin–Ciocalteu reagent (Singleton, Orthofer, & Lamuela-Raventos, 1974). The amount of total phenolics was calculated as a gallic acid equivalent (GAE) in mg per g of d.m.

2.4.2. Determination of total flavonoids content (TFC)

Total flavonoids content was determined according to the method described by Lamaison and Carnet (1990). One milliliter of extract was mixed with 1 mL of 2% AlCl3 × 6H2O solution and incubated at room temperature for 10 min. Thereafter, absorbance at 430 nm was measured. Total flavonoids content was calculated as a quercetin equivalent (QE) in mg per g of d.m.

2.5. Determination of antioxidant capacity

2.5.1. Radical scavenging activity

The experiments were carried out using an improved ABTS decolorization assay (Re et al., 1999). The affinity of test material to quench ABTS free radical was evaluated according to the following equation:
scavenging%=[(AC-AA)/AC)]×100
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where: AC – absorbance of control, AA –absorbance of sample.
Free radical scavenging ability was calculated using the Trolox standard curve prepared and expressed as mg Trolox equivalent (TE) per g of d.m.

2.5.2. Reducing power

Reducing power was determined by the method of Oyaizu (1986). Reducing power was expressed as Trolox equivalent (TE) in mg per g of d.m.

2.6. Starch content and digestibility

2.6.1. Total starch content

Total starch (TS) content was determined after dispersion of the starch granules in 2 mol/L KOH (50 mg sample, 6 ml KOH) at room temperature (30 min, constant shaking) and hydrolysis of the solubilized starch with 80 μL (1 mg/mL) amyloglucosidase (14 U mg−1; EC 3.2.1.3) at 60 °C for 45 min (Goni, Garcia-Alonso, & Saura-Calixto, 1997). Glucose content was determined by using the standard dinitrosalicylic acid (DNSA) method (Miller, 1959). Total starch was calculated as glucose × 0.9. The free reducing sugar content of the samples was determined in order to correct the total starch values. The sucrose content of the samples was also determined in order to correct the obtained total starch values. The samples dispersed in sodium acetate buffer, pH 5.0 were treated with 200 μL of (10 mg in 1 mL of 0.4 M sodium acetate buffer, pH 5.0) invertase (EC 3.2.1.26., 300 U mg−1) for 30 min at 37 °C. After centrifugation, reducing sugars were analyzed in the supernatants, using the DNS reagent. After centrifugation (3000×g, 15 min) and removal of supernatant, the pellet was dispersed with 2 mol/L KOH, hydrolyzed with amyloglucosidase and liberated glucose was quantified, as described above, for total starch (TS).

2.6.2. The resistant (RS) and potentially bioavailable (AS) starch content

The resistant (RS) and potentially bioavailable (AS) starch content was analyzed on the basis the results obtained after simulated gastrointestinal digestion (2.3.2.). After simulated digestion samples were centrifuged (3000×g, 15 min) and supernatants were removed. The pellets were washed 2 times with H2O and centrifuged. After that pellets were dispersed with 2 mol/L KOH, hydrolyzed with amyloglucosidase and liberated glucose was quantified, as described above, for total starch (TS). Resistant starch (RS) was calculated as glucose × 0.9. The potentially bioavailable starch (AS) content was calculated as the differences between TS and RS.

2.6.3. Starch digestibility

The in vitro digestibility of starch was evaluated on the basis of total starch content (TS) and resistant starch (RS) determined after digestion in vitro according to Świeca, Baraniak, et al. (2013):
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where SD – in vitro digestibility of starch, TS – total starch content, RS – resistant starch content.

2.6.4. In vitro starch digestion rate and expected glycemic index

The digestion kinetics and expected glycemic index (eGI) of the lentil sprouts were calculated in accordance with the procedure established by Goni et al. (1997). A non-linear model following the equation [C = C∝(1 − e−kt)] was applied to describe the kinetics of starch hydrolysis, where C, C1 and k were the hydrolysis degree at each time, the maximum hydrolysis extent and the kinetic constant, respectively. The hydrolysis index (HI) was calculated as the relation between the areas under the hydrolysis curve (0–240 min) of the sprout sample and the area of standard material from white bread. The expected glycemic index (eGI) was calculated using the equation proposed by Granfeldt, Björck, Drews, and Tovar (1992): eGI = 8.198 + 0.862HI.

2.7. Theoretical approach

The following factors were determined for better understanding of the relationships between biologically active compounds in the light of their bioaccessibility (Gawlik-Dziki et al., 2012):
–
The relative phenolics bioaccessibility factor (RBF):
RBF=CD/CCE
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where: CD – concentration of compounds after simulated gastrointestinal digestion, CCE – concentration of compounds after chemical extraction,
–
The relative antioxidant efficiency factor (REF):
REF=AD/ACE
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where: AD – activity of extract after simulated gastrointestinal digestion, ACE – activity of chemical extract.

2.8. Statistical analysis

All experimental results were mean ± S.D. of three parallel experiments. Two-way analysis of variance ANOVA and Turkey’s post hoc test were used to compare groups. P values <0.05 were regarded as a significant.

3. Results and discussion

The nutraceutical potential of legumes sprouts is usually determined by phenolics compounds content that exhibit antioxidant, anti-inflammatory, and anticancer properties (Chon, 2013, Gawlik-Dziki et al., 2012, Świeca and Baraniak, 2014a and Świeca and Baraniak, 2014b). Between the studied sprouts the highest contents of total phenolics, including flavonoids, were found for lentil sprouts; however, it should be noted that their contents determined after chemical extraction (CE) were significantly lower than those found for dry seeds (Fig. 1 and Fig. 2). In the case of green pea and mung bean sprouts a slight increase in chemically extractable total phenolics during the first 4 days of sprouting (1F–4F) was observed. Similar trend was found for flavonoids; however, in case of mung bean sprouts its content remained at a constant level (Fig. 1 and Fig. 2). Digestion in vitro released phenolics from fresh sprouts (RBF values significantly exceeding 1); however, flavonoids fraction was poorly bioaccessible ( Table 1). Storage at low-temperature did not affect on the total phenolics and flavonoids in lentil sprouts (both, chemically extractable and potentially bioavailable) as well as total phenolics in mung bean sprouts (potentially bioaccessible). In the case of stored, 3-day-old green pea sprouts (3S) the level of potentially bioaccessible phenolics was higher than that determined for fresh ones (an increase of 8%), whereas for 5-day-old (5S) adverse effect was observed (a decrease of 6%). In case of potentially bioaccessible flavonoids, there was a noteworthy increase (about 2- and 3-fold in respect to fresh ones (4F and 5F)) in 4-, and 5-day-old green pea sprouts (4S and 5S). A slight, but statistically significant, reduction of flavonoids content was determined after storage of the 3-day-old green pea (3F) and 5-day-old mung bean sprouts (5F) ( Fig. 1 and Fig. 2).
Total phenolics content in fresh and stored sprouts. A – green pea; B – lentil; ...
Fig. 1. 
Total phenolics content in fresh and stored sprouts. A – green pea; B – lentil; C – mung bean. Values on selected figures designated by the different letters are significantly different (P < 0.05). 1F–6F – 1–6-day-old fresh sprouts; 3S–5S – 3–5-day-old stored sprouts.
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Total flavonoids content in fresh and stored sprouts. A – green pea; B – lentil; ...
Fig. 2. 
Total flavonoids content in fresh and stored sprouts. A – green pea; B – lentil; C – mung bean. Values on selected figures designated by the different letters are significantly different (P < 0.05). 1F–6F – 1–6-day-old fresh sprouts; 3S–5S – 3–5-day-old stored sprouts.
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Table 1. The relative phenolics bioaccessibility (RBF) of fresh and stored sprouts.

Green pea
Lentil
Mung bean
Total phenolicsFlavonoidsTotal phenolicsFlavonoidsTotal phenolicsFlavonoids
Seeds1.23 ± 0.06b0.16 ± 0.01a1.21 ± 0.06a0.32 ± 0.02a1.35 ± 0.07c0.32 ± 0.02b
1F1.09 ± 0.05a0.21 ± 0.01b1.46 ± 0.07b0.36 ± 0.02a1.12 ± 0.06a0.36 ± 0.02c
2F1.10 ± 0.06a0.39 ± 0.02e1.41 ± 0.07b0.50 ± 0.03cd1.17 ± 0.06ab0.33 ± 0.02b
3F1.12 ± 0.06ab0.57 ± 0.03g1.48 ± 0.07bc0.55 ± 0.03d1.25 ± 0.06bc0.42 ± 0.02d
4F1.23 ± 0.06b0.47 ± 0.02f1.51 ± 0.08bc0.44 ± 0.02b1.35 ± 0.07c0.44 ± 0.02d
5F1.29 ± 0.06bc0.28 ± 0.01c1.65 ± 0.08c0.58 ± 0.03de1.46 ± 0.07c0.52 ± 0.03e
6F1.57 ± 0.08d0.31 ± 0.02d1.40 ± 0.07b0.57 ± 0.03de1.76 ± 0.09d0.57 ± 0.03e
3S1.37 ± 0.07c0.56 ± 0.03g1.44 ± 0.07bc0.48 ± 0.02bc1.41 ± 0.07c0.66 ± 0.03f
4S1.26 ± 0.06bc0.86 ± 0.04h1.57 ± 0.08bc0.51 ± 0.03cd1.43 ± 0.07c0.55 ± 0.03e
5S1.11 ± 0.06a1.00 ± 0.05i1.53 ± 0.08bc0.62 ± 0.03e1.34 ± 0.07c0.30 ± 0.02a
Values (±SD), in columns, designated by the different letters are significantly different (P < 0.05).
1F–6F – 1–6-day-old fresh sprouts; 3S–5S – 3–5-day-old stored sprouts.
Table options
Germination positively affected the reducing potential of chemically extractable and potentially bioaccessible fractions of green pea sprouts (Table 2). Storage caused a slight decrease of reducing power and an increase of antiradical abilities of green pea sprouts. It should be also noted that antioxidants were highly bioaccessible (REF value ca. 3 for reducing power and ca. 10 for antiradical activity). The observed changes of reducing potential of chemically extractable fraction of lentil and mung bean sprouts did not exceed 10% during germination. Digestion in vitro decreased the reducing abilities (lentil and mung bean) and antiradical potential (mung bean) of sprouts (REF values lower than 1). In the light of this very valuable is the fact that reducing potential of potentially bioaccessible fraction of stored lentil sprouts was increased (in respect to appropriate fresh one) of 40%, 31% and 23% for 3-, 4- and 5-day-old sprouts, respectively. On the other hand antiradical components of green pea and lentil sprouts were effectively extracted during digestion in vitro. Most importantly, free radical scavenging abilities of those sprouts were stable during storage (changes did not exceed 10%) ( Table 2).
Table 2. Antioxidant activity of fresh and stored sprouts.


Reducing power [μmol TE/g d.m.]
REFAntiradical activity [μmol TE/g d.m.]
REF


CEDECEDE
Green peaSeeds0.65 ± 0.10a8.06 ± 0.71g12.431.72 ± 0.21ab30.91 ± 1.03fg17.93
1F0.47 ± 0.09a8.14 ± 0.48g17.451.11 ± 0.41a25.56 ± 3.05ef22.99
2F0.93 ± 0.07b6.25 ± 0.61f6.691.13 ± 0.61ab25.58 ± 0.75e22.6
3F1.23 ± 0.08c8.96 ± 0.49gh7.281.14 ± 1.06ab25.60 ± 1.42e22.36
4F1.97 ± 0.33d8.36 ± 1.28fg4.251.22 ± 0.86ab30.08 ± 3.12fg24.73
5F2.53 ± 0.35de9.33 ± 0.35h3.681.97 ± 0.32b31.28 ± 2.86efg15.9
6F2.85 ± 0.36e9.33 ± 0.42gh3.274.97 ± 0.18d33.46 ± 1.57fg6.73
3S2.36 ± 0.11de6.87 ± 0.76fg2.912.55 ± 0.20bc31.60 ± 1.82fg12.4
4S2.30 ± 0.14de7.59 ± 1.44fg3.32.43 ± 0.14b28.84 ± 1.38fg11.85
5S2.28 ± 0.09d6.92 ± 1.10fg3.042.92 ± 0.23c25.21 ± 3.07ef8.62

LentilSeeds55.59 ± 3.68de49.94 ± 1.22c0.954.74 ± 1.41c61.31 ± 0.43e1.12
1F58.19 ± 1.57e22.63 ± 0.75a0.3933.95 ± 2.31b60.39 ± 0.67e1.78
2F54.00 ± 1.26d21.25 ± 4.03a0.3930.91 ± 1.38ab56.90 ± 3.04cde1.84
3F54.17 ± 7.47de23.02 ± 6.40ab0.4230.02 ± 3.62ab59.11 ± 1.51cd1.97
4F61.39 ± 5.24de26.08 ± 2.40a0.4231.66 ± 2.09ab60.23 ± 0.84ef1.9
5F51.96 ± 8.01de27.30 ± 4.07ab0.5326.58 ± 5.10ab56.48 ± 4.29cd2.12
6F53.84 ± 7.31de28.33 ± 4.61ab0.5326.62 ± 4.24ab58.49 ± 0.50d2.2
3S53.94 ± 8.06de32.02 ± 4.75b0.5928.91 ± 0.91b59.68 ± 1.89cd2.06
4S54.75 ± 3.09de34.24 ± 4.42b0.6324.90 ± 1.08a59.06 ± 0.72d2.37
5S50.99 ± 5.62de33.43 ± 6.87b0.6627.73 ± 2.68ab60.42 ± 1.10de2.18

Mung beanSeeds51.93 ± 1.07h15.86 ± 1.98a0.3186.79 ± 0.75g19.57 ± 1.26c0.23
1F54.62 ± 1.18i18.29 ± 1.46ab0.3378.38 ± 3.46e22.81 ± 1.05d0.29
2F55.18 ± 1.58hi18.07 ± 1.92ab0.3379.17 ± 2.00e22.63 ± 1.31d0.29
3F53.48 ± 1.21hi19.24 ± 2.33abc0.3684.79 ± 0.34f19.66 ± 1.35c0.23
4F48.71 ± 0.69fg21.51 ± 2.03bc0.4482.43 ± 2.07ef17.85 ± 2.52c0.22
5F50.10 ± 2.26fgh25.63 ± 1.58cd0.5184.57 ± 1.42f15.78 ± 0.24b0.19
6F45.04 ± 0.39e29.90 ± 1.91d0.6685.29 ± 0.54f16.97 ± 0.80b0.2
3S46.12 ± 2.20ef20.46 ± 0.19b0.4486.26 ± 1.35fg14.90 ± 0.51a0.17
4S47.38 ± 0.65f20.82 ± 0.28b0.4484.43 ± 2.48efg15.41 ± 0.12a0.18
5S49.93 ± 0.87g23.72 ± 1.30c0.4882.21 ± 1.45e16.24 ± 0.22b0.2
Values, within the selected activity and species, designated by the different letters are significantly different (P < 0.05).
CE – chemical extracts; DE – extracts obtained after digestion in vitro; REF – the relative antioxidant efficiency factor.
1F–6F – 1–6-day-old fresh sprouts; 3S–5S – 3–5-day-old stored sprouts.
Table options
The highest starch contents were found for dry seeds (regardless of the legume type) but starch levels were decreased during germination (after 6 days of sprouting a decrease of 43%, 37% and 44% in green pea, lentil and mung bean sprouts, respectively) (Table 3). In green pea sprouts, the total starch content did not change after storage but it should be noted that the resistant starch contents were significantly lower than those determined for fresh sprouts (a decrease of 35%, 36% and 28% for 3-, 4-, and 5-day old sprouts, respectively). There were no significant changes of the resistant starch content, except for 3-day-old lentil sprouts, between fresh and stored mung bean and lentil sprouts. Sprouting caused an increase of starch digestibility that was clearly visible in case of lentil sprouts (an increase of about 70% after 4–6 day of sprouting in respect to dry seeds). Most importantly storage of sprouts significantly elevated values of expected glycemic index (eGI), wherein the highest eGI were determined for 5-day-old stored sprouts; 75.17 - green pea, 83.18 – lentil and 89.87 – mung bean (Table 3).
Table 3. Starch content and digestibility, expected glycemic index of fresh and stored sprouts.


Total starch [mg/g d.m.]Resistant starch [mg/g d.m.]Available starch [mg/g d.m.]Starch digestibility [%]Expected glycemic index
Green peaSeeds325.5 ± 32.9d125.4 ± 12.85d200.2 ± 10.02d61.49 ± 3.07a27.61 ± 0.69a
1F257.8 ± 0.3c94.9 ± 14.81c162.9 ± 6.68bc63.18 ± 2.21a32.19 ± 0.80b
2F258.5 ± 43.7abcd91.8 ± 23.74bcd166.7 ± 13.34bc64.50 ± 3.22ab36.57 ± 0.91c
3F240.1 ± 46.9abcd79.3 ± 16.93bc160.8 ± 8.20bc66.96 ± 2.68ab39.31 ± 0.98d
4F221.0 ± 0.5a75.9 ± 8.14bc145.1 ± 4.50a65.66 ± 3.28ab39.04 ± 0.98d
5F218.2 ± 20.7ab71.6 ± 20.66abc146.6 ± 2.93a67.18 ± 3.36ab43.07 ± 1.08e
6F213.3 ± 43.8abc59.9 ± 13.77ab153.3 ± 4.31ab71.89 ± 2.52bc44.85 ± 1.12f
3S236.7 ± 0.2b58.1 ± 0.05a178.5 ± 9.10c75.43 ± 3.77c45.51 ± 1.14f
4S217.3 ± 14.8a56.1 ± 14.78ab161.2 ± 6.61b74.18 ± 2.97c46.11 ± 1.15f
5S213.2 ± 40.4abc55.9 ± 10.39ab157.3 ± 9.44ab73.78 ± 3.69bc75.17 ± 1.88g

LentilSeeds307.8 ± 8.5e179.7 ± 1.63d128.1 ± 6.53b41.62 ± 2.08a36.00 ± 0.90a
1F283.4 ± 8.4d152.3 ± 9.24c131.1 ± 5.37b46.26 ± 1.62b44.75 ± 1.12b
2F271.0 ± 21.3d137.9 ± 13.82bc133.2 ± 10.65bc52.82 ± 2.64c48.57 ± 1.21c
3F268.5 ± 36.2cde126.3 ± 7.02b142.2 ± 7.25c49.23 ± 1.97bc49.07 ± 1.23c
4F201.1 ± 22.2b57.0 ± 4.12a144.0 ± 4.47c71.64 ± 3.58e55.90 ± 1.40d
5F169.9 ± 2.0a55.2 ± 13.68a114.7 ± 2.29a67.52 ± 3.38de63.09 ± 1.58e
6F180.5 ± 25.6ab50.1 ± 11.25a130.5 ± 3.67b72.26 ± 2.53e69.04 ± 1.73f
3S196.9 ± 23.0b56.6 ± 10.61a140.4 ± 7.16cb71.27 ± 3.56e57.74 ± 1.44d
4S213.3 ± 30.1bc68.9 ± 7.23a144.4 ± 5.92c67.68 ± 2.71ed58.54 ± 1.46d
5S193.7 ± 14.1b71.3 ± 16.34a122.4 ± 7.35ab63.21 ± 3.16d83.18 ± 2.08g

Mung beanSeeds310.1 ± 8.9e151.9 ± 14.23d158.1 ± 8.06e51.00 ± 2.55a32.68 ± 0.82a
1F244.3 ± 1.5d104.7 ± 9.46bc139.5 ± 5.72d57.13 ± 2.00bc37.33 ± 0.93b
2F236.2 ± 1.1c102.2 ± 9.21bc144.0 ± 11.52de60.98 ± 3.05c42.48 ± 1.06c
3F230.3 ± 1.1bc110.4 ± 3.27bc119.9 ± 6.12bc55.90 ± 2.24ab46.03 ± 1.15d
4F221.9 ± 5.1b83.0 ± 1.43a138.9 ± 4.30d62.60 ± 3.13c53.81 ± 1.35e
5F210.9 ± 15.7ab81.5 ± 2.46a129.4 ± 2.59c61.37 ± 3.07c61.75 ± 1.54f
6F192.4 ± 4.1a79.9 ± 0.89a112.5 ± 3.16ab58.47 ± 2.05c76.73 ± 1.92g
3S229.1 ± 14.8bcd91.5 ± 5.47b137.5 ± 7.01d60.04 ± 3.00c52.77 ± 1.32e
4S222.4 ± 2.8b78.4 ± 14.06ab144.0 ± 5.90d49.92 ± 2.00a60.54 ± 1.51f
5S174.2 ± 26.0a70.0 ± 7.14a104.2 ± 6.25a59.82 ± 2.99c89.87 ± 2.25h
Values, within the selected characteristic, designated by the different letters are significantly different (P < 0.05).
1F–6F – 1–6-day-old fresh sprouts; 3S–5S – 3–5-day-old stored sprouts.
Table options
In foods of plant origin antioxidant potential is closely linked with its low-molecular antioxidants level, especially phenolics. Generally, the determined amounts of phenolics and antioxidant capacities levels, as well as a kinetic of changes during sprouting are comparable with available literature data (Cevallos-Casals and Cisneros-Zevallos, 2010, Pająkk et al., 2014 and Świeca, Sęczyk, and Gawlik-Dziki, 2014). The differences may be caused by the different start material (varieties), variable sprouting conditions and extraction systems (Xu and Chang, 2007 and Świeca et al., 2012). Digestion in vitro released phenolics from sprouts which confirm results obtained for total phenolics. Similar observations were found previously for sprouts e.g. broccoli ( Gawlik-Dziki et al., 2012), lentil ( Świeca, Baraniak, et al., 2013) or food products enriched with polyphenols e.g. bread enriched with quinoa leaves ( Świeca, Seczyk, Gawlik-Dziki, & Dziki, 2014) and coffee enriched with willow bark ( Durak, Gawlik-Dziki, & Sugier, 2014). Surprisingly, flavonoids were poorly bioaccessible ( Table 1). According to literature data flavonoids are stable during digestion (pH and temperature as well as enzymes released phenolics from glycosides and/or cell wall elements). Lowered bioaccessibility may be caused by formation of complexes with components of digestive tract and/or sprouts protein and starch ( Scalbert & Williamson, 2000).
According to RBF value a reductive potential of lentil sprouts is mainly created by flavonoids. Although, total phenolics of lentil sprouts were well bioaccessible a reducing powers of extracts obtained after digestion were significantly lower than those determined for chemical extracts. This observation highly corresponded with changes in flavonoids fraction. Phenolics play a key role as antioxidants in legumes; however, according to these studies it may be speculated that creation of antioxidant activity may also contribute to other compounds. In some cases, especially in green pea sprouts, an increase in antioxidant activity was disproportionate to increase in polyphenols content, both during germination and after in vitro digestion. Thus, it may be speculated that in this case the role in the creation of antioxidant potential is also played by bioactive peptides and oligosaccharides. The presence of antioxidant peptides in legume proteins hydrolysates obtained after treatment with digestive enzymes was already confirmed by Karaś, Jakubczyk, and Baraniak (2010). On the other hand these disparities may be caused by the interaction of sprouts phenolics among themselves and with other sprouts components. There are only few studies concerning changes in the antioxidant level of sprouts during storage at low temperatures; however, they do not provide any information about the effects of postharvest storage on the antioxidant activity and level and bioaccessibility of bioactive constituents.
In the study performed by Świeca et al. (2014), potentially bioaccessible fraction of lentil sprouts, treated during germination at low and high temperature, recorded the same reducing and chelating power as fresh samples. Goyal et al. (2014) proved that during storage of mung bean at room and low temperature ascorbic acid, total phenols and antioxidant activity of sprouts firstly increased and then decreased significantly. Similar observation was found in this study where reducing potential of mung bean sprouts was lowered after storage; however, antiradical activity of sprouts has not been changed. During 1 week storage of broccoli sprouts at 4 °C and 8 °C ascorbic acid contents were decreased, while total phenol contents were generally stable (Waje et al., 2009). The effect of cold storage of the 7-day-old-sprouts of broccoli, kohl rabi, white radish and rocket was studied by Force, O’Hare, Wong, and Irving (2007). They proved that there is no significant loss of glucosinolates (potential anticancer compounds) under domestic refrigeration conditions.
Sprouting significantly changes the nutritional quality of legumes seeds. Nutrients and micro- and macroelements are more accessible; usually vitamin content is also increased. Legumes are known to be an excellent source of nutrients (valuable source of starch and protein) and importantly their consumption does not cause an abrupt increase in postprandial blood glucose level, which in turn induces immediate oxidative stress (Hoover & Zhou, 2003). Somehow pro-health benefits of legume sprout-rich diet is linked with a high content of phenolics, an important role plays also starch content and its quality. On germination, a significant decrease in the starch contents was observed, which could be due to the use of starch as an energy source in the sprouting process. These data agree with the findings of Ghavidel and Prakash (2007) for the germinated green gram, cowpea, lentil, and chickpea. Relatively high amounts of resistant starch, in comparison to other studies (Eyaru et al., 2009 and Hoover and Zhou, 2003), are probably caused by the method used for its determination – in vitro conditions (gastrointestinal digestion). Generally, no information is available on changes in starch content and its digestibility during cool storage. Some researchers postulate that the changes are linked with modification of the starch structure (reduction of amylose content, which is very resistant due to its higher crystallinity), and content and the structure and/or activity of factors influencing the rate of its mobilization (e.g., amylase inhibitors, tannins, phytic acid) ( Cevallos-Casals and Cisneros-Zevallos, 2010, Ghavidel and Prakash, 2007 and Hoover and Zhou, 2003). It is thus suggested that during storage (similarly to sprouting), starch structure is loosened, which probably creates a large space within the matrix and increases the susceptibility to enzymatic attack ( Benítez et al., 2013). This statement is also supported by the previous studies of Fernandez and Berry, 1989 and Frias et al., 1998, who observed that germination sharply increased the susceptibility of chickpea and lentil starch to digestion by α-amylase, indicating the influence of dextrinization in producing material more susceptible to enzymatic attack. On the other hand, although at low temperature metabolic rate is reduced, germination caused dynamic changes in the amylases level and activity. These factors consequently improved the digestibility of starch and reduced the resistant starch content which may partially explain a significant increase of starch digestibility and expected glycemic index values (in respect to fresh sprouts) determined after 1 week storage at 4 °C.

4. Conclusion

Legumes are food products highly desired by the modern communities because of their low glycemic index and high amounts of potentially resistant starch and polyphenolics. Sprouting and further storage can effectively modify the nutraceutical and nutritional values. Both, time of germination and storage, diversified the phenolics antioxidant levels, antioxidant activity of sprouts and affect their bioaccessibility in vitro. Postharvest storage significantly increases the starch digestibility and expected glycemic index value that was linked with reduction of resistant starch content. In the light of these results, it may be concluded that the bioactivity and nutritional quality of sprouted legumes are affected by storage at low temperatures, however, there is no a simple pattern which could predict potential changes.

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