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LWT - Food Science and Technology 41 (2008) 1541e1547 www.elsevier.com/locate/lwt

Effect of processing on antioxidant contents in selected dry beans (Phaseolus spp. L.) J. Boateng, M. Verghese*, L.T. Walker, S. Ogutu Department of Food and Animal Sciences, Alabama A&M University, Normal, AL 35762, USA Received 22 August 2007; received in revised form 25 October 2007; accepted 6 November 2007

Abstract Dry beans are rich sources of dietary fiber and phytochemicals such as flavonoids and phenolics. In the current study, we determined the effects of two processing methods (soaking and toasting) on total phenolics, flavonoids, proanthocyanidin and antioxidant potential by 2,2diphenyl-1-picrylhydrazyl (DPPH) activity and ferric reducing antioxidant potential (FRAP) in selected dry beans (red kidney beans (K), black-eyed peas (B), pinto beans (P) and soy beans (S)). Total phenolics (mg/g dwb) expressed as gallic acid equivalents (GAE), total flavonoids (mg/g dwb) as catechin equivalents (CE) and proanthocyanidin expressed as leucocyanidin equivalent (mg LE/g) ranged from 3.42 to 7.21, 0.61 to 0.84 and 0.51 to 3.13 in raw beans; 3.58 to 6.94, 0.19 to 0.99 and 0.43 to 3.13 in soaked beans and 4.55 to 9.52, 0.23 to1.00 and 0.20 to 3.25 in toasted beans, respectively. FRAP (mg/g) in raw, soaked and toasted dry beans ranged from 0.00097 to 0.00424 while DPPH(T30) (%) ranged from 43.9 to 62.61. Our results indicate that processing methods (soaking and roasting) influenced total phenolic, flavonoid and antioxidant contents (DPPH, FRAP) in selected dry beans. Ó 2008 Published by Elsevier Ltd on behalf of Swiss Society of Food Science and Technology. Keywords: Dry beans; Phenolics; Proanthocyanidin; FRAP; DPPH

1. Introduction Dry beans (Phaseolus spp. L.) are the most important grain legumes for human consumption. Dry beans (Messina, 1999), have been cultivated for thousands of years, and have played (and still do) an important role in the traditional diets of many regions throughout the world. While dry beans (legumes) are important in the diets of most developing countries, in contrast, they are less significant in western diets. The daily per capita consumption of all bean products in Asia alone is 110 g compared to about 9 g in the United States. Among the most popular beans consumed in the United States are pinto beans, navy beans, red kidney beans, northern and lima beans (Messina, 1999). * Corresponding author. Department of Food and Animal Sciences, Alabama A&M University, 4900 Meridian Street, P.O. Box 1628, Normal, AL 35762, USA. Tel.: þ1 256 372 4175; fax: þ1 256 372 5432. E-mail address: [email protected] (M. Verghese).

Dry beans are widely known for their fiber, mineral and protein contents; however, its nutraceutical value is yet to gain as much attention in the prevention of chronic diseases (Dinelli et al., 2006). The protective effects of dry beans in disease prevention such as cancer may not be entirely associated to dietary fiber, but to phenolic and other non-nutritive compounds (Oomah, Tiger, Olson, & Balasubramanian, 2006). Polyphenols from dry beans may possibly act as antioxidants, hindering the formation of free radicals that eventually lead to the deterioration of biological molecules. These naturally occurring phenolic compounds are predominantly present in the seed coat and possess antimutagenic and antioxidant activities (Aparicio-Fernandez, Yousef, Loarca-Pina, de Mejia, & Lila, 2005). Most studies on phenolic contents in dry beans are conducted in the seed coats, where polyphenols are concentrated. Numerous processing and cooking methods have been shown to possibly reduce antinutrients such as trypsin inhibitors and phytic acid, and increase the content of tannins,

0023-6438/$34.00 Ó 2008 Published by Elsevier Ltd on behalf of Swiss Society of Food Science and Technology. doi:10.1016/j.lwt.2007.11.025

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catechins and polyphenols (Vidal-Valverde et al., 1993). Research on cooking and the retention of bioactive components show that while a healthy diet may contain an adequate amount of antioxidants, the way food is prepared and cooked may be just as important as what is eaten (Vallejo, Toma´sBarbera´n, & Garcı´a-Viguera, 2003). Therefore, the objective of the current study was to investigate that the effect of two processing methods (soaking and toasting) on bioactive components and antioxidant potential in dry beans. 2. Materials and methods 2.1. Processing of raw dry beans Dry beans (red kidney beans, pinto beans, black-eyed peas and soybeans) were purchased from a local food store. 2.2. Raw beans The beans were cleaned and 100 g of the four selected raw beans were ground to a fine powder (particle size of 0.5 mm) using a Wiley mill (Swedesboro, NJ). 2.2.1. Soaking of dry beans The beans were thoroughly rinsed in distilled water and 100 g were soaked in 1 l of water overnight (approximately 22 h) at room temperature (25  C). The soaked beans were removed from the soak water, rinsed three times in distilled water, flash frozen in liquid nitrogen and then freeze dried. The freeze dried beans were ground to a fine powder (particle size of 0.5 mm) using a Wiley mill (Swedesboro, NJ). 2.2.2. Toasting of beans Freeze dried beans were roasted in a microwave oven (Kenmore microwave oven, 1200 W) for 6 min. The beans were cooled and ground to a fine powder (particle size of 0.5 mm) using a Wiley mill (Swedesboro, NJ). All the ground samples were stored in amber jars at 4  C until analysis. 2.3. Preparation of dry bean extracts Dry bean (raw, soaked and toasted) extracts were prepared based on the method described by Adom and Liu (2002) with slight modifications. Briefly, 5 g of bean samples were extracted with 50 ml of chilled aqueous ethanol (ethanol: water, 80:20 v/v) for 2 h at room temperature. The samples were centrifuged at 3000g for 20 min and the supernatant was removed. Extraction was repeated twice and supernatants were pooled and evaporated at 40  C. The phenolic concentrate was made to a final volume of 10 ml with distilled water and stored at 80  C until analysis. The extraction was carried out in triplicate.

Liu, 2002; Adom, Sorrells, & Liu, 2003; Singleton, Orthofer, & Lamuela-Raventos, 1999). Briefly, 1 ml of appropriately diluted samples of bean extracts (1:20 for all beans; 1:10 for black-eyed peas. A 1:20 dilution for black-eyed peas was very low when absorbance (A750 nm) was taken.) was oxidized with 1 ml of FolineCiocalteau reagent (Sigma chemical Co, St. Louis, MO) diluted with reverse osmosis deionized water (RODI) (1:10 v/v). After a 6-min incubation period, the reaction was neutralized with 3 ml of 70 g/l sodium carbonate solution (Na2CO3) (v/v). Gallic acid was used as the standard (0.01 mg/100 ml). After 2 h incubation at room temperature, a resulting blue color was read at an absorbance of 750 nm. A reagent blank was measured against water blank. Total phenolic content in beans was expressed as mg gallic acid equivalent (GAE)/g. The samples were analyzed in four replications. 2.5. Determination of total flavonoids in dry beans Total flavonoid content in dry beans was analyzed by a colorimetric method described by Adom and Liu (2002), Adom et al. (2003), and Kim, Chun, Kim, Moon, and Lee (2003). Briefly, 0.5 ml of appropriately diluted samples of bean extracts (1:20) was added to 2 ml of RODI, the mixture was reacted with 150 ml of 50 g/l sodium nitrite (NaNO2) and the mixture was allowed to incubate for 5 min after which 150 ml of 100 g/l aluminum chloride (AlCl3) was added. AlCl3 forms a complex with the flavonoids in the extract. The samples were further incubated for 5 min and 1 ml of 1 M sodium hydroxide (NaOH) was added. Another 1.5 ml of water was immediately added to the resulting pink mixture. The absorbance was immediately read at 510 nm. The flavonoid contents in the bean extracts were compared to that of catechin standards (0.001 mg/ml). Samples were analyzed in four replications and reported as mg catechin equivalent (CE)/g. 2.6. Determination of antioxidant activities in dry beans 

2.6.1. 1,1-diphenyl-2-picrylhydrazyl (DPPH ) radicalscavenging activity in dry beans Antioxidant activity of dry beans was determined by the method of Brand-Williams, Cuvelier, and Berset (1995) and modified by Kim et al. (2003) and Lee, Kim, Lee, and Lee (2003). 1,1-diphenyl-2-picrylhydrazyl (DPPH ) radical was dissolved in aqueous ethanol (ethanol: water, 80:20 v/v) to give a final concentration of 200 mmol/l. A series of sample concentrations of 0.1 ml for each sample (10:90, 20:80, 30:70, 40:60 and 50:50) (extract: aqueous ethanol) was added to 2.9 ml of DPPH radical solution. For the control, 0.1 ml aqueous methanol (methanol: water, 80:20 v/v) was added to 2.9 ml of DPPH radical solution. The mixture was shaken and allowed to stand in the dark at room temperature for 30 min. The absorbance of the mixture was then read at 517 nm. The radical scavenging activities of the samples were expressed in terms of IC50 (concentration required for 





2.4. Determination of total phenolics in dry beans Total phenolics in dry beans (raw, soaked and toasted) were analyzed following the FolineCiocalteau method (Adom &

J. Boateng et al. / LWT - Food Science and Technology 41 (2008) 1541e1547

a 50% decrease in absorbance of DPPH, radical) relative to the control (100%) and calculated as % inhibition of DPPH .

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2.8. Statistical analysis





where, A517 control is the absorbance of the control (DPPH solution without test sample) and A517 sample is the absorbance of the test sample (DPPH , solution plus antioxidant). 

2.6.2. Ferric reducing antioxidant power (FRAP) in dry beans The FRAP assay for dry beans was based on the methods of Benzie and Strain (1999). Briefly, 100 ml of appropriately diluted samples of dry bean extracts (1:20 for all beans; 1:10 for black-eyed peas) was added to 3 ml of working FRAP reagent consisting of 300 mmol/l acetate buffer (pH 3.6), 10 mmol/l 2,4,6-tri(2-pyridyl)-s-triazine (TPTZ) in 40 mmol/l HCL and 20 mmol/l ferric chloride (FeCl3$6H2O). The FRAP reagent was made fresh. After 10 min incubation at 37  C, the absorbance was recorded at 593 nm. The absorbance changes in the test mixture were compared to those obtained from standard mixture of ferrous sulphate (FeSO4$7H20) (0.1 mmol/le 1.0 mmol/l). The increasing concentration of Fe3þ is expressed as mmol of Fe2þ/g. Samples were analyzed in four replications. 2.7. Determination of condensed tannin (proanthocyanidin) in dry beans 2.7.1. Extraction of tannins in dry beans For the extraction of tannins, 10 ml of aqueous acetone (70:30 v/v) was added to 1 g of bean samples and the mixture was subjected to ultrasonic treatment for 20 min at room temperature. The mixture was centrifuged for 10 min at 3000g at 4  C and the supernatant was collected. The mixture was subjected to another round of extraction as previously described. The supernatants were pooled and evaporated at 30  C to remove the acetone and stored at 80  C until analysis. 2.7.2. Determination of condensed tannins Condensed tannins in dry beans were analyzed following the protocol from the FAO/IAEA techniques in the Food and Agricultural Organization manual (2000). The protocol was based on the method of Porter, Hrstich, and Chan (1986). A 3.0 ml of butanoleHCl reagent (95:5 v/v) and 0.1 ml of 2% ferric reagent (20 g/l ferric ammonium sulfate dissolved in 2 N HCl) was added to (0.5 ml) dry bean extracts. The samples were thoroughly mixed and heated in boiling water for 60 min. The samples were covered with aluminum foil prior to boiling. After cooling to room temperature, the absorbance at 550 nm was recorded. To prevent the samples from exceeding the absorbance of 0.6, the extracts were diluted with aqueous acetone (70:30 v/v). Condensed tannins (% in dry matter) as leucocyanidin equivalent was calculated by the formula: ðA550 nm  78:26  dilution factorÞ=ð% dry matterÞ assuming that E1%,

1 cm, 550 nm

of leucocyanidin is 460.

Data were analyzed by (Analysis Of Variance) ANOVA using the SAS 9.0 (2005) statistical program. Values were given as means  SEM (Standard Error of the Mean) and means were separated using Tukey’s studentized range test. Level of significance was set at p  0.05. 3. Results 3.1. Total phenolics content in raw and processed dry beans and effect of processing on phenolic contents In the raw beans, total phenolics were significantly ( p < 0.05) lower in black-eyed peas (B) compared to kidney beans (K), pinto beans (P) and soybeans (S) (Fig. 1). There were no significant ( p  0.05) differences in total phenolics among K, P and S. In the processed beans (soaked), total phenolics were significantly ( p  0.05) higher in K and P compared to B and S. Total phenolic contents in toasted beans were significantly ( p  0.05) higher in K compared to P, S and B. There were no significant ( p  0.05) differences in total phenolic content between S and B. Soaking did not have any significant ( p  0.05) effect on total phenolic contents in K, but in P, total phenolics content was significantly ( p  0.05) lower after soaking (Fig. 1). Total phenolics in B, K and P were 27, 24 and 18%, respectively, after toasting compared to the raw beans. However, there was a 29% reduction in total phenolics in toasted S compared to the raw S. 3.2. Total flavonoid content in raw and processed dry beans and effect of processing on flavonoid content Total flavonoids in the raw beans were significantly ( p  0.05) higher in K and S compared to B and P. Total flavonoids in B were significantly ( p  0.05) higher compared to P. Among the raw beans, P had the lowest total flavonoid content. In the pre-processed (soaked) beans, K had significantly ( p  0.05) higher total flavonoid content and S had

12

Total Phenolics (mg/GAE/g) dwb



DPPH radical scavenging activityð%Þ :   A517 control  A517 sample A517 control  100

a,d

10

b,d

8

ab,e a,f

6 4

a,e

a,f

c,d

b,d b,c

c,f c,e

c,e

2 0 B

P

K

S

Dry beans Fig. 1. Total phenolics (mg GAE/g) in raw and processed dry beans (K ¼ kidney beans, B ¼ black-eyed peas, P ¼ pinto beans, S ¼ soybeans, GAE ¼ gallic acid equivalent). Values are means  SEM, n ¼ 4. abcMeans on bars with similar letters do not differ ( p  0.05) using Tukey’s studentized range test. defMeans on bars with similar letters do not differ ( p  0.05) using Tukey’s studentized range test (raw, ,; soaked, G; toasted, -).

J. Boateng et al. / LWT - Food Science and Technology 41 (2008) 1541e1547

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significantly ( p  0.05) lower flavonoid contents compared to B, K and P. Total flavonoid contents in the toasted dry beans were also not significantly different among B, K and P (Fig. 2). Soaking increased total flavonoid in K and P (0.992  0.1 and 0.913  0.1) but it did not affect the flavonoid content in B (0.765  0.1) (Table 1). The total flavonoids were significantly ( p  0.05) higher after toasting in B, K and P. There was a 21, 15 and 39% increase in total flavonoids in toasted B, K and P, respectively, compared to the soaked beans, although there was a 71 and 76% reduction in toasted and soaked S compared to the raw beans. 3.3. Proanthocyanidin contents in raw and processed dry beans There were no significant differences in proanthocyanidin content between raw K and P, and there were also no significant differences between B and S (Fig. 3). Among the raw dry beans, P had the highest proanthocyanidin content compared to B, K and S. In the soaked beans, proanthocyanidin content was significantly ( p  0.05) lower in S compared to B, K and P. There were no significant differences among B, K and P. Toasting significantly ( p  0.05) increased proanthocyanidin in K (3.25  0.25) and significantly ( p  0.05) reduced the level in S (0.200  0.02) compared to the raw beans. Processing (soaking and toasting) reduced the proanthocyanidin contents in S by 54 and 79% compared to the raw beans. The highest increase in proanthocyanidin was in B, with an 82 and 79% increase after soaking and toasting compared to the raw B (Fig. 3). 3.4. DPPH and ferric reducing antioxidant potential (FRAP) in raw and processed dry beans FRAP value was significantly ( p  0.05) higher in the raw P and S compared to B and K (Table 2). Raw B had a significantly ( p  0.05) lower FRAP value compared to the other two beans. In the pre-processed beans (soaked), there were no significant ( p  0.05) differences between K and P, and there were also no significant ( p  0.05) differences between P and B. However, S had a significantly ( p  0.05) lower

Table 1 Correlation relationship between antioxidant activities and total phenolics contents in selected dry beans using Pearson correlation coefficient

Raw beans (B, K, P, S) Soaked beans (B, K, P, S) Toasted beans (B, K, P, S)

c,e

b,e

a,e

a,d

c,f

0.6 0.4

d,f

0.2

b,e

0

B

K

P

S

Dry beans Fig. 2. Total flavonoid (mg CE/g) content in raw and processed dry beans (K ¼ kidney beans, B ¼ black-eyed peas, P ¼ pinto beans, S ¼ soybeans, CE ¼ catechin equivalent). Values are means  SEM, n ¼ 4. abcMeans on bars with different letters differ ( p  0.05) using Tukey’s studentized range test. defMeans on bars with different letters differ ( p  0.05) using Tukey’s studentized range test (raw, ,; soaked, G; toasted, -).

R2

6.121 5.355 6.737

0.0021 0.00248 0.00279

0.862 0.721 0.947

FRAP value (0.00121 mg/g) compared to B, K and P. FRAP value in the toasted beans did not significantly ( p  0.05) differ between K and P. FRAP values in toasted K and P were significantly ( p  0.05) higher compared to toasted B and S. FRAP value significantly ( p  0.05) increased after processing in K and P, however, it was significantly ( p  0.05) reduced in S and B after toasting. FRAP value increased with processing in the dark colored beans. DPPH IC50 values in dry beans ranged from 49.7 to 62.6; 43.9 to 61.4 and 48.1 to 58.7 in raw; soaked and toasted beans (Table 2). There were no significant differences in DPPH IC50 values between B, K, P and S. Among the raw dry beans, the highest scavenging activity against the DPPH (IC50) value was seen in K (62.6), with S (49.79) having the lowest scavenging activity. Among the soaked beans, the greatest IC50 value was seen in P (61.4), followed by K (57.9) and B (55), again the lowest was seen in S. However, among the toasted beans, the lowest IC50 value was seen in P (48.1) and the highest was seen in K (58.7). In B, DPPH IC50 value was higher in the raw (56.8) than in the processed (soaking or toasting). In S, DPPH IC50 value was higher in toasted (54.8) compared to the raw (49.7) or soaked (43.9) beans. 4. Discussion The current study was conducted to quantify the total phenolics, flavonoids and proanthocyanidin contents and antioxidant activity as determined by FRAP and DPPH in raw and

Proanthocyanidin (mg LE/g) dwb

Total Flavonoids (mg CE/g) dwb

0.8

a,d

a,d a,d

a,d

b,c

FRAP (mg/g)* (n ¼ 4)

Abbreviations: K ¼ kidney beans, B ¼ black-eyed peas, P ¼ pinto beans, S ¼ soybeans, FRAP ¼ ferric reducing, antioxidant potential. *Average values for black-eyed peas, kidney beans, pinto beans and soybeans were used.

1.2 1

Total phenolics* (mgGAE/g) dwb (n ¼ 4)

4 3.5 3 2.5 2 1.5 1 0.5 0

a,d

a,d b,d

a,d

a,d a,e

a,e b,e b,d

b,e

b,e

B

K

P

b,e

S

Dry beans Fig. 3. Proanthocyanidins contents (mg LE/g) in raw and processed dry beans (K ¼ kidney beans, B ¼ black-eyed peas, P ¼ pinto beans, S ¼ soybeans, LE ¼ leucocyanidin equivalent). Values are means  SEM, n ¼ 4. abcMeans on bars with similar letters do not differ ( p  0.05) using Tukey’s studentized range test. defMeans on bars with similar letters do not differ ( p  0.05) using Tukey’s studentized range test (raw, ,; soaked, G; toasted, -).

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Table 2 Total antioxidant capacity (TAC) in raw and processed dry beans Dry beans

B K P S

FRAP (mg/g)

DPPH(T30) (%)

Raw

Soaked

Toasted

Raw

Soaked

Toasted

0.00190c,e 0.00233b,f 0.00268a,f 0.00274a,d

0.00241b,d 0.00332a,e 0.00297ab,e 0.00121c,e

0.00173b,f 0.00421a,d 0.00424a,d 0.00097c,f

56.15  1.27a,d 62.61  4.65a,d 55.33  5.36a,e 49.79  1.09a,e

55.03  2.57a,d 57.99  1.01a,e 61.43  7.27a,d 43.90  0.67a,f

50.51  2.80a,f 58.73  11.21a,e 48.14  1.16a,f 54.84  2.47a,d

Values are means  SEM, n ¼ 4. abc Means in a column with different letters differ ( p  0.05) using Tukey’s studentized range test. def Means in a row with different letters differ ( p  0.05) using Tukey’s studentized range test. Abbreviations: K ¼ kidney beans, B ¼ black-eyed peas, P ¼ pinto beans, S ¼ soybeans, FRAP ¼ ferric reducing antioxidant potential, DPPH ¼ 1,1-Diphenyl-2picrylhydrazyl radical.

processed (soaked and toasted) whole dry beans (Phaseolus vulgaris L), kidney beans, pinto beans, black-eyed peas and soybeans. The total phenolic contents in the unprocessed or raw dry beans ranged from 3.42 to 7.21 mg GAE/g (dwb). Beans with darker seed coats, such as kidney beans and pinto beans had significantly ( p  0.05) higher total phenolics compared to those with lighter seed coats i.e., black-eyed peas and soybeans. Studies have shown that dry beans possessing darker colored seed coats have relatively higher total phenolics in comparison to those with lighter colored seed coats (Barampama & Simard, 1995). Total phenolic (29.0  0.56 mg/g) content in black soybean was found to contain higher polyphenols than yellow soybean (0.45  0.02 mg/g) (Takahashi et al., 2005). While total phenolic values in the current study are high, Heimler, Vignolini, Dini, and Romani (2005) reported phenolic content in dry bean samples ranging from 1.17 to 4.40 mg/g. Luthria and Pastor-Corrales (2006) have also reported phenolics in the range of 26.7e36 mg/100 g GAE in pinto beans and 20.9e27.4 mg GAE /100 g in kidney beans and 19.1e48.3 mg GAE/100 g in dry bean cultivars. Total phenolic contents in dry beans differ between studies, this difference according to Luthria and Pastor-Corrales (2005) could be attributed to various factors which include but not limited to genotype, agronomic practices, maturity at harvest, post-harvest storage and climatic conditions and growing and storage conditions. Recovery of polyphenols from fruits and vegetable seem to depend significantly on solvents used during extraction. Acetone and methanol extracts of samples have shown to have higher phenolic yields compared to either methanol or acetone alone. Cardador-Martinez, Castano-Tostado, and Loarca-Pina (2002) reported a yield that was more than six times that of acetone or methanol alone. Total flavonoids (raw beans) in this study ranged from 0.614 mg CE/g in pinto beans to 0.845 mg CE/g in kidney beans. Heimler et al. (2005), reported similar values for total flavonoids in dry beans. Oomah, Cardador-Martinez, and Loarca-Pina (2005) reported values of 0.24 and 0.26 mg CE/ g for flavonoid content in pinto beans and kidney beans, respectively. The high flavonoid levels in the beans maybe due to their high anthocyanin contents. Kidney beans cultivated in the United States have been reported to contain high anthocyanin levels (Choung, Choi, An, Chu, & Cho, 2003). The

seed color of beans determines the presence and concentration of flavonol glycosides, anthocyanins, and condensed tannins (proanthocyanidins). In dry beans, the most widely distributed group of flavonoids are proanthocyanidins (Aparicio-Fernandez et al., 2005). Lately, proanthocyanidins or condensed tannins have been shown to be effective antioxidants with even greater activity than simple phenolics (Hagerman, Riedl, & Rice, 1999). In this study, condensed tannin contents ranged from 0.51 to 3.130 mg LE /g in raw beans. Plahar, Annan, and Nti (1997) reported tannin concentrations of 0.3e6.9 and 7.2e116 mg CE/g flour from whole cowpea seeds and seed coats, respectively. Condensed tannin levels were 9.17e35.70 mg catechin equivalents/g flour in a study conducted by EspinosaAlonso, Lygin, Widholm, Valverde, and Paredes-Lopez (2006). It has been reported that higher condensed tannins or proanthocyanidin content are seen in colored beans than in pale (in yellow and white) colored beans (Beninger & Hosfield, 1999). However, Cardador-Martinez, Loarca-Pina, and Oomah, 2002 suggested that the highest antioxidant activity in white seeds might be due to their condensed tannin content. Since total phenolics were relatively low in the pale colored seeds it is possible to assume that the major phenolics in these seeds could be proanthocyanidins. FRAP and DPPH in raw beans are comparable to other studies (Amarowicz, Naczk, & Shahidi, 2000; Pellegrini et al., 2006). Dark colored beans (kidney beans and pinto beans) compared to pale colored beans (black-eyed peas) had higher amounts of phenolic compounds and may contribute to the high antioxidative potential (FRAP value). Although soaking and cooking dry beans have been shown to reduce antinutrients such as trypsin inhibitor and reduce phytic acid, it can also increase the content of tannins, catechins and polyphenols. Total phenolics were lower after soaking but increased after toasting. Soaking leads to softening of cell wall tissues which is usually accompanied by solubilization of bound polyphenols, hence it is likely that these compounds may have leached into the soak water during this process. However, Siddhuraju and Becker (2001) observed that total phenolics were significantly ( p < 0.05) higher after soaking and dry heating compared to cooking (boiling) or autoclaving of both raw and presoaked seeds. It is important to also note that the rate of increase in phenolic contents

J. Boateng et al. / LWT - Food Science and Technology 41 (2008) 1541e1547

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y = 0.0003x + 0.0017 R2 = 0.9187 FRAP

10 8 6 4 2 0

0.0035 0.0025

y = 1.0186x + 3.575 R2 = 0.5275 Total phenolics

B

K

0.0015

FRAP (ug/g)

Total phenolic (mg GAE/g)

primarily depends on the type of legume and the preparation procedure used. Hence, toasting rather than boiling or cooking yielded higher phenolic and flavonoid contents in dry beans. Additionally, the disruption of the cell wall through heating or the breakdown of insoluble phenolic compounds could have led to better extractability of these compounds. Dewanto, Wu, and Liu (2002) explained that thermal processing may release more bound phenolic acids from the breakdown of cellular constituents. We found a strong correlation between total phenolics and FRAP activity in toasted dry beans (R2 ¼ 947, p < 0.05) (Fig. 4) and a positive but lower correlation in soaked dry beans (R2 ¼ 0.721, p < 0.05) (Table 1). The high correlation values of this relationship indicate that phenolic compounds in the dry beans were responsible for their antioxidant capacity. Dewanto et al. (2002) also reported an increase in antioxidant activity following thermal treatments of sweet corn. The authors attributed the improved antioxidant activity to the additive and synergistic effects of other phytochemicals. The correlations between total phenolics and DPPH activity in the soaked (R2 ¼ 0.616, p < 0.05) and toasted (R2 ¼ 0.309, p < 0.05) dry beans was, however, less strong (data not shown). This could mean that other phytochemicals other than phenolics may potentially play a role in the antioxidant activity in dry beans. In their study Duenas, Fernandez, Hernandez, Estrella, and Munoz (2005) found no significant ( p < 0.05) differences in DPPH inhibition by raw and fermented beans although there was a slight increase after heating. In previous studies, Barroga, Laurena, and Mendoza (1985) showed that boiling and roasting of dry beans resulted in 73

0.0005 P

S

10 8 6 4 2 0

y = -0.0004x + 0.0035 R2 = 0.3043 FRAP

K

0.0035 0.0025

y = 0.1966x + 4.864 R2 = 0.0245 Total phenolics

B

0.0045

0.0015 0.0005 P

FRAP (ug/mg)

Total Phenolics (mg/GAE/g)

Dry beans

S

10

y = -0.0002x + 0.0034 R2 = 0.0296 FRAP

8

0.005 0.004 0.003

6 y = -0.067x + 6.905 2 R = 0.0012 Total Phenolics

4 2

0.002 0.001

FRAP (ug/g)

Total phenolics (mg GAE/g)

Dry beans

0

0 B

K

P

S

Dry beans Fig. 4. Correlation between total phenolics (y-axis) and antioxidant activities (z-axis) in raw, soaked and toasted dry beans (Total Phenolics, ; FRAP, ; Linear FRAP ). A; Linear Total Phenolics,

and 17% reductions in polyphenols, respectively. The authors (Barroga et al., 1985) positively correlated the lowering of polyphenols in dry beans with a decrease in protein-precipitable phenols. According to Siddhuraju (2006) and Duenas et al. (2005), the stability of antioxidant products such as phenolics and flavonoids during heating maybe due to the formation of mailliard products such as hydroxymethylfurfuraldehyde (HMF) which produces high antioxidant activity. An earlier study by Nicoli, Anese, Parpinel, Franceschi, and Lerici (1997) showed that medium dark roasted coffee brews had the highest antioxidant properties due to the formation of maillard reaction products. Plahar et al. (1997) also noted that roasted cowpeas (black-eyed peas) contained similar nutritional and functional qualities as protein supplements. The authors concluded that roasted cowpeas could thus be used in cereal-based weaning foods. After a series of thermal treatments of sweet corn and masa, Dewanto et al. (2002) and de la Parra, Saldivar, and Liu (2007) reported significant increases in the contents of total free phenolics. Dry beans are an integral part of diets in a significant portion of the world population, but the potential benefits of consuming beans from a nutraceutical standpoint have largely been overlooked. Since dry beans contain compounds other than fiber that may have significant antioxidant potential, it will be useful to investigate their potential in alleviating diseases and maximizing their use in food industry.

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