Food Research International 44 (2011) 436–441

Contents lists available at ScienceDirect

Food Research International

j ourna l homepage: www.e lsev ie r.com/ locate / foodres

Phenolics and antioxidant activity of lentil and pea hulls

B. Dave Oomah a,⁎, François Caspar b, Linda J. Malcolmson c, Anne-Sophie Bellido c

a National Bioproducts and Bioprocesses Program, Pacific Agri-Food Research Centre, Agriculture and Agri-Food Canada, 4200 Highway 97, P.O. Box 5000, Summerland, BC, Canada V0H 1Z0b Université de la Réunion, Ecole Supérieure d'Ingénieurs en Développement Agroalimentaire Intégré (ESIDAI), 97490 Sainte Clotilde, Ile de la Réunion, Francec Special Crops and Pulses, Canadian International Grains Institute (CIGI), 1000-303 Main Street, Winnipeg, Manitoba, Canada R3C 3G

⁎ Corresponding author. Tel.: +1 250 494 6399; fax:E-mail address: oomahd@agr.gc.ca (B.D. Oomah).

0963-9969/$ – see front matter. Crown Copyright © 20doi:10.1016/j.foodres.2010.09.027

a b s t r a c t

a r t i c l e i n f o

Article history:Received 9 July 2010Accepted 27 September 2010

Keywords:Pulse hullsLentilsYellow peaAntioxidant activitySolvent extractionPhenolicsWater extractHulls

Hulls obtained by pilot-scale dehulling process fromgreen and red lentils and yellow peaswere fractionated intohulls (N500 μm) and residues (b500 μm). These fractions together with the corresponding whole seeds wereextractedwith four solvents, aqueous (70%) acetone, (80%) ethanol, hot water (70–80 °C) andwater (22±1 °C)and evaluated for antioxidant activity in relation to phenolic contents. Aqueous acetone (70%, v/v) extracted thehighest level of total phenolics at about 87 mg of catechin equivalent per gram of sample from lentil hullsfollowed by hot water, water and aqueous ethanol (80%, v/v). Red lentil hull with maximum concentration ofphenolic compounds exhibited the strongest antioxidant activity of 260 mg (1040 μM) trolox equivalent/g hull.Hot water andwater extracted significantly higher total phenolics fromwhole pulse and their residue than othersolvents, but with weaker antioxidant activity. Aqueous ethanol was always the best extractant of phenolicantioxidant from yellow pea irrespective of its millstream. The most potent phenolic antioxidant was obtainedfromwater extract of green and red lentil hulls, followedby those extractedwith ethanol, acetone, andhotwater.Total phenolic content was highly correlated with antioxidant activity of pulses and its millstreams.

Crown Copyright © 2010 Published by Elsevier Ltd. All rights reserved.

1. Introduction

Pulse processing leaves a milling by-product which consists of amixture of hulls, cotyledon, broken pieces, and flour containing starchand protein currently sold for animal feed. The by-product can beseparated into hull and “hull-reduced” fractions that can be furtherprocessed to a high protein product (Harle & Green, 2008). Incommercial lentil processing this by-product ranging between 10 and21% is considered a significant loss (Narasimha, Ramakrishnaiah, &Pratape, 2003). Pea and lentil hulls, constituting approximately 8% ofthe seed, are considered major by-products of the pulse processingfacilities in Western Canada. Yellow pea hull fiber is already widelyused in food products and its benefits particularly in the health of theelderly has been reported (Fitzpatrick, 2007). Lentil hull has limiteduse and therefore represent a potential source of value-added healthyproducts for the expanding dietary fiber market and for furthereconomic phytochemical extraction.

Legume seed coats are rich sources of polyphenolics and naturalantioxidants that could replace the synthetic antioxidants in foods(Moise, Han, Gudynaite-Savitch, Johnson, & Miki, 2005). For example,Dueñas, Hernández, and Estrella (2006) showed that flavones,flavonols, and proanthocyanidins from methanolic extracts of theseed coat and cotyledon of lentils and dark peas contributed the mostantioxidant capacity to the seed coat, whereas the flavonoid catechin

+1 250 494 0755.

10 Published by Elsevier Ltd. All rig

was the major antioxidant contributor of the cotyledon. The lentilseed coat with higher flavonoid compounds had superior antioxidantactivity than that of peas. In both lentils and peas, the cotyledoncontained mainly non-flavonoid compounds with less antioxidantactivity than flavonoids abundant in the seed coat. Water solubleantioxidants have been derived from lentil seed coat highly enrichedin polyphenolics using various acceptable food grade solvents(Ronzio, Muanza, & Sparks, 1998). The hot water extract (carriedout at 100 °C) fractionated further with volatile organic solvent(consisting of methanol, ethanol and acetone) contained flavonols,(kaempferol, quercetin) condensed tannins (proanthocyanidins), andphenolic acids including ferulic, protocatechuic and caffeic acids andwas considered a potent antioxidant.

Several studies on phenolic and antioxidants of pulse hulls havealready been reported (Dueñas et al., 2006; Amarowicz, Troszyńska,Barylko-Pikielna, & Shahidi, 2004; Troszyńska, Estrella, López-Amóres,& Hernández, 2002; Troszyńska, Bednarska, Latosz, & Kozlowska,1997), but typically those hulls were mostly obtained manually.Yellow pea, green and red lentils were used in this study because oftheirmajor importance in pulse production, processing andmarketingin western Canada. Although the commercial production, applicationsand extensive use of fiber fractions from peas have been reasonablywell studied, lentil hulls and fibers have not been adequatelyinvestigated for future food applications.The objective of this studywas to develop an economically viable procedure for the efficientantioxidant extraction system from hulls obtained by mechanicaldehulling of pulses at the pilot-scale (pre-commercial) level. Thisprocedure differed from others in the literature in that hulls of fairly

hts reserved.

437B.D. Oomah et al. / Food Research International 44 (2011) 436–441

high purity are produced with the same latest specially developedtechnology for pulse processing currently in commercial use withplant capacities ranging from 50 to 150 t per day. Development of newbioproducts from seed hulls is one of the major priorities of the PulseResearch Network aimed at harnessing new market opportunities forthe pulse industry.

2. Materials and methods

Pulse samples consisting of green lentil (cv. CDC Plato), red lentil(cv. CDCRedberry), yellowpea (unknown origin) and their hull fractionswere kindly provided by the Canadian International Grains Institute(CIGI,Winnipeg, MB). CDC Plato and CDC Redberry are classified as largegreen (N65 mg seed weight) and small red (30–50 mg seed weight)lentils, respectively. The hulls were obtained by processing the seedsthrough the automatic “S.K. System” (S.K. Engineering & Allied Works,Bahraich, UP, India) processing plant that incorporates conditioning andcomplete milling of pulses. The pulse samples were tempered to 14%moisture prior to milling by pitting with a pitter/dehuller/splittermachine (5/8 gap size setting) (S.K. Engineering & Allied Works,Bahraich, UP, India), followed by water addition and tempering for 6 h.The samples were stirred twice during the 6 h tempering (30 min afterwater addition and then 1 h later). After tempering, the samples wereheated at 70 °C for 20 min, and then cooled at room temperatureovernight in a cooling tower. The sampleswere dehulledwith the pitter/dehuller/splittermachine (7/8 gap size setting). Thehullswere separatedwith an aspiration unit of the S.K. System.

The hull fraction was separated into hulls (N500 μm) and residues(b500 μm) using a sieving machine (type AS 200 Tap, Retsch GmbH,Haan, Germany) with 32-mesh U.S. standard, 500-μm sieve for 3 min.The “true” sieved hull represented 79.9, 77, and 87.5% (w/w) of greenlentil, red lentil, and yellow pea hull fractions, respectively. Allsamples except residues were ground to a fine powder in a smallcoffee mill.

2.1. Extraction and analysis of phenolics

Extraction of phenolics was performed according to the proceduredescribed recently (Oomah, Corbé, & Balasubramanian, 2010).Ground samples (200 mg) were extracted with aqueous ethanol80% (v/v) (8 ml), using a Reacti-Therm heating and stirring module(N 18970, Pierce, Rockford, IL) for 2 h and filtered (Acrodisc 0.45 μm,VWR International, Mississauga, ON). Similar extractions were carriedout with aqueous acetone 70% (v/v), distilled water (22±1 °C) andhot distilled water (70–80 °C). The water extracts were centrifuged(Sorvall Model RC 5B plus; DuPont Co.Wilmington, DE) at 11,000 g for30 min and filtered through the glass wool. Water was boiled in akettle for the hot water extraction and the resultant temperature 70–80 °C in the system was maintained during extraction. The recoveredsupernatant was stored at −20 °C in the dark until analysis. Allextractions were carried out in triplicates.

The phenolic content of extracts was determined according to theprocedure described previously (Oomah, Cardador-Martinez, &Loarca-Piña, 2005). Briefly, the method consisted of adding 100 μl ofsample extract with 150 μl of a solution of 2% HCl in 80% ethanol in a96-well ultra violetflat-bottomplate (Greiner Bio-One Inc., Longwood,FL). The absorbance of the solution was monitored at 280, 320, 360,and 520 nm after mixing for 2 min with a spectrophotometer(Spectramax Plus 384, Molecular Devices Corp., Sunnyvale, CA)using (+)-catechin (0–100 μg/ml), caffeic acid (0–20 μg/ml), querce-tin (0–30 μg/ml), and cyanidin-3-glucoside (0–8 μg/ml) as standardsfor total phenolics, tartaric esters, flavonols and anthocyanins,respectively. Standards were prepared in aqueous ethanol 80% (v/v).The absorbance was also read at 710 nm for turbidity correction andthe results were expressed in mg (+)-catechin, caffeic acid, quercetinor cyanidin-3-glucoside equivalents/g sample. The acetone extract

was evaporated in the Reacti-Therm module under nitrogen anddissolved in aqueous ethanol 80% (v/v) prior to phenolic assay.

2.2. Antioxidant assay

Antioxidant activity was measured using the oxygen radicalabsorbance capacity (ORAC-FL) described previously (Oomah, Blanchard,& Balasubramanian, 2008), based on the procedure described by Prioret al. (2003). A SpectraMax GeminiEM microplate fluorescence reader(Molecular Devices Co., Sunnyvale, CA) was used with excitation andemission wavelength at 485 and 530 nm, respectively. Sample extractsand Trolox standards were diluted with 75 mM phosphate buffer (pH7.4) prior to transfer to a 96-well microplate (Fluotrac 200, Greiner Bio-One Inc., Longwood, FL). A peroxyl radicalwas generated byAAPH [2, 2′-azobis (2-methylpropionamide) dichloride] (Sigma Aldrich, USA)during measurement, and fluoresceine was used as the substrate.Measurements were taken every 2 min for 120 min upon addition ofAAPH. Final ORAC values were calculated using a regression equationbetween the Trolox concentration (0–4 μg/ml) and the net area underthe curve (AUC in μM Trolox equivalents (TE)/g sample) and convertedto mg/g sample.

At least three determinations were made for all assays, except forantioxidant activity that were determined in duplicate. An analysis ofvariance by the general linear models (GLM) procedure, meanscomparison by Duncan's test, and Pearson correlation were per-formed according to Statistical Analysis System (SAS Institute Inc,1990). Data were compared within each millstream for green and redlentils, whereas those of yellow pea (with values an order ofmagnitude lower than lentils) were analyzed separately.

3. Results and discussion

3.1. Acetone extract

The sifted hull residue represented 23, 20.1, and 12.5% (w/w) ofred lentil, green lentil, and yellow pea hull fractions, respectively. Theamount of red lentil residue (23%) was similar to that reportedpreviously (Wang, 2008) in dehulling Canadian red lentil varietiesindicating significant removal of cotyledons through this process.Pulse hulls had significantly (Pb0.0001) higher concentration of totalphenolic and phenolic components than their corresponding wholeseeds or residue when extracted with aqueous acetone (Table 1).Total phenolic content of lentil hulls were seven (red) to eight (green)times those in their whole seeds, whereas tartaric esters andanthocyanin contents were three to four folds the concentration inseeds. Furthermore, the total phenolic content of all red lentil acetoneextracts were significantly (Pb0.0001) higher than those of thecorresponding green lentil extracts. Differences in content of phenoliccomponents of acetone extracts between red and green lentils weresignificant (Pb0.0001) in hull, and residue. However, this differencewas not significant when comparing extracts of whole red and greenlentils, except for total phenolics. The difference in anthocyanincontent and antioxidant activity between whole seed and residueextracts of red and green lentils were also not significant.

Yellow pea hull had almost twice the concentration of totalphenolics and over six times the anthocyanin content of the seed. Theratio of the total phenolic content of the hull andwhole seed extractedwith acetone was comparable to the 2.5 to 13 reported previously(Troszyńska et al., 1997). The yellow pea hull and residue had similarand the highest anthocyanin contents, probably due to the flavoneglycosides preferentially extracted with acetone–water (Troszyńskaet al., 2002). The differences in total phenolic and anthocyanincontents were significant (Pb0.0001) for yellow pea residue, butinsignificant for red lentil residue compared to whole seeds. Flavonolcontent of hulls and residues were higher (≥1.5–2.8 times) than inseeds.

Table 1Phenolic content and antioxidant activity of pulses extracted with acetone.a

Millstreams Concentrationb TEACc (ORAC)

Totalphenolics

Tartaricesters

Flavonols Anthocyanins mg/g μmol/g

WholeGreenlentil

10.31d 0.56e 0.59d 0.026c 34.08c 136.17c

Red lentil 12.62c 0.54e 0.55d 0.026c 33.74c 134.81cHull

Greenlentil

82.95b 1.90a 1.07b 0.106b 239.18b 955.63b

Red lentil 87.16a 1.80b 1.18a 0.112b 260.18a 1040.53aResidue

Greenlentil

11.40d 0.91c 1.10ab 0.017c 27.40c 109.48c

Red lentil 12.89c 0.78d 0.93c 0.026c 29.59c 118.21cYellow peaWhole 3.45gz 0.30fz 0.16fy 0.020cy 2.59fz 10.36fzHull 6.89ex 0.81dx 0.45ex 0.136ax 5.73ey 22.89eyResidue 5.69fy 0.52ey 0.43ex 0.134ax 10.15dx 40.56dx

a Means in a column with different letters for each pulse are significantly different(pb0.05).

b Concentration of phenolic compounds are expressed as mg (+) catechin, caffeicacid, quercetin or cyanidin-3-glucoside equivalents/g sample (dry matter basis) fortotal phenolics, tartaric esters, flavonols and anthocyanins, respectively.

c Trolox equivalent antioxidant capacity.

438 B.D. Oomah et al. / Food Research International 44 (2011) 436–441

The total phenolic content of pulse hulls were higher, whereas theflavonol and anthocyanin contents were lower than those reportedpreviously for pea and lentil hulls extracted with hot aqueous acetone(80% acetone at 80 °C) (Troszyńska et al., 1997). The strongantioxidant activity of red and green lentil hulls (≥7 fold comparedto seeds) properly reflected their high concentration of total phenolicsand phenolic compounds. The residue of red and green lentilsexhibited antioxidant activity not significantly different than thoseof the whole seed. Yellow pea exhibited weak antioxidant activitywith the residue expressing almost twice and four times theantioxidant activity of the hull and whole seed extracts, respectively.The antioxidant activity of yellow peawas similar to those reported byHalvorsen et al. (2002).

Table 2Phenolic content and antioxidant activity of pulses extracted with hot water.a

Millstreams Concentrationb TEACc (ORAC)

Totalphenolics

Tartaricesters

Flavonols Anthocyanins mg/g μmol/g

WholeGreenlentil

19.31d 1.04d 1.01d 0.120d 13.49b 53.88b

Red lentil 20.83d 1.27c 1.24c 0.207b 15.67b 62.62bHull

Greenlentil

57.07a 1.92b 1.11c 0.124d 148.07a 591.60a

Red lentil 52.75b 2.00a 1.39b 0.174c 140.68a 562.06aResidue

Greenlentil

24.15c 1.37c 1.58a 0.102d 14.34b 57.28b

Red lentil 20.45d 1.03d 1.12c 0.086d 16.42b 65.58bYellow peaWhole 15.11ex 0.74ey 0.61ey 0.170cy 4.92cx 19.66cxHulls 9.10fy 0.82ey 0.32fz 0.030ez 2.93dy 11.72dyResidue 15.58ex 1.33cx 1.18cx 0.279ax 4.99cx 19.93cx

a Means in a column with different letters for each pulse are significantly different(pb0.05).

b Concentration of phenolic compounds are expressed as mg (+) catechin, caffeicacid, quercetin or cyanidin-3-glucoside equivalents/g sample (dry matter basis) fortotal phenolics, tartaric esters, flavonols and anthocyanins, respectively.

c Trolox equivalent antioxidant capacity.

3.2. Hot water extract

Lentil (red and green) hulls extractedwith hot water (Table 2) hadsignificantly higher total phenolics (≥2.5-folds) and, tartaric esters(1.8-fold) than whole seeds, whereas flavonol and anthocyanincontents were not significantly different for green lentils. The totalphenolic content of red lentil hulls was similar to the 53 mg/g drymatter of lentil seed coat reported earlier (Kubicka & Troszyńska,2003). Hot water extract of yellow pea hull had significantly(Pb0.0001) lower total phenolics, flavonol and anthocyanin contentsthan those from the whole seeds. Concentrations of phenoliccomponents from pea residue extract were significantly (Pb0.005)higher, although the total phenolic content was not different thanthose of the whole seed. The hot water extract of green lentil residuehad significantly (Pb0.0001) higher total phenolic and tartaric estercontents than the whole seed and the highest flavonol content. Thetotal phenolic content of hot water extract from the whole red lentilwas not significantly different from that of the residue (Table 2)similar to those observed with acetone extract. Conversely, hot waterextracts of green lentil hull and residue had significantly differentconcentrations of phenolic components than those of thecorresponding red lentil millstreams. The hot water extract of yellowpea hulls had the lowest total phenolic, flavonol and anthocyanincontents in contrast to the residue with the highest anthocyanin andmoderate tartaric esters and flavonol contents.

The antioxidant activities of lentil hulls were about 9 to 10 timesthose of the whole seed, disproportionate to the level of totalphenolics. This indicates that the antioxidant activity reflects otherantioxidant components of lentil hulls in addition to phenolicsextracted with hot water. The yellow pea hull extract exhibited thelowest antioxidant activity about 60% of those from the whole seedand residue extracts.

3.3. Water extract

There was no significant difference in the total phenolic contentsof water extract between green and red lentils from the whole seed,hull and residue (Table 3). Furthermore, only the hull and residueextracts showed significant differences in phenolic components whencomparing green and red lentils. Lentil hulls contained over threetimes the total phenolic content and over twelve-fold the antioxidantactivity of the whole seed. The total phenolic content of hot water andwater extracts of the lentil hull were comparable to those extractedfrom the lentil seed coat withmethanol–water (1:1 v/v) (Ronzio et al.,1998) (38–52 mg catechin equivalent/g sample dry weight). Thewater extract of yellow pea hull, similar to its hot water extract, hadthe lowest total phenolic, flavonol and anthocyanin contents.Moreover, the yellow pea hull had only half the total phenoliccontent, but exhibited antioxidant activity not different from those ofthe whole seed or residue. The total phenolic content of pulse residuewas significantly (Pb0.0001) higher than those of the whole seed,although that difference was not reflected in the antioxidant activity.The high phenolic content of the residue corresponds to the highertannin content in the powder fraction compared to the raw seedobserved in dehulling the red lentil (Wang, 2008). Therefore, watercan be considered a viable solvent for extracting antioxidantcomponents only from the lentil hull.

3.4. Ethanol extract

The total phenolic contents of ethanolic extract from whole seedsand hulls of green lentil were not significantly different from those ofthe corresponding red lentil extract (Table 4); similar to thosedescribed above with water extract. The differences were also notsignificant in tartaric ester content between green and red lentils fromwithin the whole and hull extracts. However, the green lentil residue

Table 3Phenolic content and antioxidant activity of pulses extracted with water.a

Millstreams Concentrationb TEACc (ORAC)

Totalphenolics

Tartaricesters

Flavonols Anthocyanins mg/g μmol/g

WholeGreenlentil

15.33d 0.61f 0.63e 0.028e 17.36c 69.37c

Red lentil 15.88d 0.56f 0.57e 0.027e 6.70d 26.77dHull

Greenlentil

49.54a 1.57a 0.77d 0.056b 215.46a 860.83a

Red lentil 49.48a 1.43b 0.92c 0.065a 164.19b 656.01bResidue

Greenlentil

21.42b 1.20c 1.48a 0.042c 17.53c 70.04c

Red lentil 20.12b 0.91d 1.11b 0.034c 7.18d 28.68dYellow peaWhole 15.39dy 0.51fy 0.32fgy 0.046cx 5.61ex 22.43exHull 8.03ez 0.81ex 0.24gy 0.010fy 5.87ex 23.45exResidue 17.67cx 0.62fy 0.41fx 0.062ax 5.41ex 21.62ex

a Means in a column with different letters for each pulse are significantly different(pb0.05).

b Concentration of phenolic compounds are expressed as mg (+) catechin, caffeicacid, quercetin or cyanidin-3-glucoside equivalents/g sample (drymatter basis) for totalphenolics, tartaric esters, flavonols and anthocyanins, respectively.

c Trolox equivalent antioxidant capacity.

439B.D. Oomah et al. / Food Research International 44 (2011) 436–441

extract had significantly higher phenolic contents, except anthocya-nins, and antioxidant activity than those of the red lentil. Ethanolicextract of lentil (both red and green) hulls contained over four timesthe total phenolic content that was reflected in their five-foldantioxidant activity compared to those from the whole seed.Furthermore, differences in antioxidant activity between green andred lentils were not significant within the whole seed and hullmillstreams. The yellow pea hull extract had the lowest total phenolicand anthocyanin contents thereby expressing the weakest antioxi-dant activity of all ethanol extracts. The antioxidant activity of theyellow pea residue extract was significantly higher than those fromthe whole seeds and hulls; this value was similar to those of theacetone extract (Table 1).

Table 4Phenolic content and antioxidant activity of pulses extracted with ethanol.a

Millstreams Concentrationb TEACc(ORAC)

Totalphenolics

Tartaricesters

Flavonols Anthocyanins mg/g μmol/g

WholeGreenlentil

5.04d 0.49d 0.59e 0.018d 14.58d 58.26d

Red lentil 5.19d 0.41d 0.54e 0.025c 14.54d 58.10dHull

Greenlentil

24.63a 1.01b 0.67d 0.055b 82.13a 328.12a

Red lentil 27.27a 1.03b 0.79c 0.064a 85.34a 340.95aResidue

Greenlentil

9.33b 1.13a 1.45a 0.020d 23.53b 94.03b

Red lentil 7.75c 0.89c 1.12b 0.027c 17.34c 69.28cYellow peaWhole 3.08fz 0.29ey 0.15gy 0.013ey 6.95fy 27.76fyHull 2.60gy 0.30ey 0.12gz 0.008fz 5.77gz 23.06fzResidue 4.94ex 0.50dx 0.26fx 0.017dex 10.04ex 40.10ex

a Means in a column with different letters for each pulse are significantly different(pb0.05).

b Concentration of phenolic compounds are expressed as mg (+) catechin, caffeicacid, quercetin or cyanidin-3-glucoside equivalents/g sample (dry matter basis) fortotal phenolics, tartaric esters, flavonols and anthocyanins, respectively.

c Trolox equivalent antioxidant capacity.

The total phenolic content and antioxidant activity of ethanol(80%, v/v) extracted whole lentil seed (Table 4) were higher thanthose reported for four Spanish lentil cultivars (Fernandez-Orozco,Zieliński, & Piskula, 2003). The yellow pea and lentil hulls had thelowest and highest total phenolic contents (2.6–27.7 mg/g catechinequivalent). The residue, representing part of the cotyledon, generallyhad over one and a half times (1.5–1.9) the phenolic content of theseeds, irrespective of pulse type. Tartaric esters and flavonols werepresent in the whole lentils and their residues in almost equalconcentrations with the highest and lowest concentration in thegreen lentil residue and the whole and hull of the yellow pea,respectively. In lentils, the hulls had twice the tartaric esterconcentration, whereas the residues had twice the flavonols thanthe whole seeds. Flavonol content of hulls was significantly lowerthan those of residue, in contrast to the higher flavonol content in thehull extracted with 80% acidified methanol (Dueñas et al., 2006). Theflavonol content of the yellow pea was about four times the amount inlentils; an observation similar to those reported earlier (Xu & Chang,2007). Anthocyanin content was highest in lentil hulls, ≥2.5 timesthan in whole lentils and similar to their proanthocyanidin range 0.66to 1.04% reported previously (Ronzio et al., 1998). Aqueous ethanolwas always the best extractant of phenolic antioxidant from yellowpea irrespective of its millstream. The antioxidant activity of ethanolicextract of the whole green lentil was similar to those of the red lentil(58 μmol/g), whereas those of water extract for the green lentil wasover twice the level in the red lentil.

3.5. Comparison of solvent extracts

The relative amounts of phenolics in pulses differed significantly(Pb0.0001) among types, millstreams and extracting solvents(Tables 1–4). Maximum differences were observed in total phenolicsof the lentil and yellow peas only when extracted with aqueousacetone comparable to those reported earlier (Xu & Chang, 2008),with two different aqueous acetone concentrations. Aqueous acetoneextracted over twice (2.2–2.5×) the antioxidant level than hot wateror aqueous ethanol from whole lentils, but less than half (0.4–0.5×)the amount from yellow peas. The lentil hull presented the best source(material) for antioxidant recovery, since the acetone extractexhibited almost three times (2.9–3×) the antioxidant activity ofthe ethanol extract. Ethanol, known as an effective solvent for therecovery of phenolic compounds, was the best antioxidant extractantfor yellow peas, similar to aqueous acetone and, 1.4–2×more efficientthan water extract. Aqueous ethanol extracted the least amount(about 3 times less than extracted with acetone) of total phenolic. Thehighest amount of total phenolic was extracted with hot water andwater from whole seed and residue and with acetone from the hull.However, these high phenolic water extracts exhibited weakantioxidant activity relative to acetone extracts. Hot water andwater actually extracted more total phenolics than aqueous acetoneor ethanol from the whole, hull and residue of yellow pea. This is incontrast to the superior polyphenol extraction of pea seed coat withacetone–water system compared with ethanol–water systemreported by Troszyńska et al. (2002).

Acetone extracts of lentil exhibited the highest antioxidant activityfollowed by water, hot water, and ethanol extracts. This contrastswith the similar antioxidant activity of phenolic compounds extractedfrom lentil seeds using three solvent systems: 80% (v/v) acetone, 80%(v/v) methanol, and 80% (v/v) ethanol observed in a beta-carotene–linoleate model system (Amarowicz, Karamac, & Chavan, 2001). Thehigh antioxidant activity of the water extract, particularly of lentilsmay be due to the presence of non-phenolic antioxidant components(Harle & Green, 2008). Hot water extracted the highest concentrationof tartaric acid, flavonols and, anthocyanins from the whole seed andresidue.

440 B.D. Oomah et al. / Food Research International 44 (2011) 436–441

Aqueous acetone was the best solvent for extracting totalphenolics and antioxidants from lentil hulls with significantly higherrecoveries from red than green lentils. Antioxidant activity wasconsistently maximally expressed in aqueous acetone extracts of thewhole, hull and residue of both green and red lentils. The totalphenolics of hot water extracts were generally higher than thoseextracted with water probably due to their release from food matrixresulting from structural breakdown or because of their thermalinstability (Ranilla, Genovese, & Lajolo, 2009). Hot water extracted themost total phenolics from the whole lentils and their residue, butdisplayed lower potent antioxidant activity than their respectiveacetone extracts. Water extracts of lentil hulls exhibited highantioxidant activity second only to their acetone extract, not reflectingtheir modest total phenolic content. The antioxidant activity of greenlentil hull extracts segregated succinctly based on the extractingsolvent, whereas those from the red lentil hull showed no significantdifference between hot water and water extracts. This may resultfrom the greater ease [clean removal of the seed coat] of dehullinggreen (with larger seed and thicker seed coat) than the small seededred lentils (therefore purer hull fraction).

Lentil hulls had higher total phenolic contents and antioxidantactivities than those of the yellow peas independent of extractingsolvent thereby confirming similar earlier reports (Dueñas et al.,2006). The total phenolics and antioxidant activities of residue,representing cotyledons were lower than those of hulls independentof extracting solvent only in lentils. The cotyledon in both lentils andpeas had lower concentration of phenolic compounds than the seedcoat due to different distributions of phenolic compounds in the twoseed parts (Dueñas et al., 2006).

The comparison of the phenolic content of pulse with antioxidantactivity revealed strong correlation (Table 5). The total phenoliccontent correlated positively with tartaric esters and flavonolcontents (r=0.85 and 0.47, pb0.0001, respectively), indicating thatextraction yielding high total phenolic content will result in extractsenriched in tartaric esters and flavonols. The total phenolic andtartaric esters were the best indicators of antioxidant activity, andtotal phenolic content can therefore be used as the best predictor ofantioxidant activity based on its significant correlation. The correla-tion coefficient between total phenolic and flavonols was reduced,whereas that with anthocyanins improved when considering lentilsamples only.

The most potent phenolic antioxidant was obtained from the waterextract of green and red lentil hulls (17.3 and 13.3 μM trolox/mgphenolics), followed by those extractedwith ethanol (12.8 and 13.4 μMtrolox/mg phenolics), acetone (11.5 and 11.9 μM trolox/mg phenolics),and hot water (10.4 and 10.7 μM trolox/mg phenolics). The lentil hullsare antioxidant-enriched fibers that can be very useful as newingredients to allow new product development platforms (Cudrado etal., 2002; Thavarajah et al., 2008) in reduced-calorie foods with all the

Table 5Correlation coefficients for phenolics and antioxidant activity of pulses.

Tartaric esters Flavonols Anthocyanins TEAC

All samples (n=108)Total phenolics 0.853a 0.468a 0.335b 0.938a

Tartaric esters 0.751a 0.532a 0.760a

Flavonols 0.416a 0.324Anthocyanins 0.184

Lentil samples (n=72)Total phenolics 0.864a 0.263c 0.537a 0.938a

Tartaric esters 0.620a 0.703a 0.771a

Flavonols 0.475a 0.078Anthocyanins 0.365b

TEAC = Trolox equivalent antioxidant capacity.a Pb0.0001.b Pb0.005.c Pb0.05.

health benefits of dietary fiber to control body weight and combatobesity for the fortified/functional products market. Although severalextraction and isolation procedures have been proposed, new markettrends dictate using solventwith the statusGRAS (Generally Recognizedas Safe) (Takeuchi et al., 2009). The selectivity and unlimited restrictionof water makes it an excellent cost-effective solvent for extractingpotent phenolic antioxidants from lentil hulls.

4. Conclusions

This investigation in addition to numerous other studies confirmsthat most of the bioactive phenolics and antioxidants in lentils areconcentrated in the hulls. Water was the most and hot water, the leastsensitive solvent in differentiating antioxidant activity of green andred lentil millstreams.The benefit provided by the antioxidantperformance of lentil hull extracted by water can respond to therequirements of numerous food manufacturers seeking to eliminatesolvent based fiber ingredients in their food. Lentil hull impartssuperior antioxidant benefits compared to whole seed and mayretain/carry that benefit when incorporated into processed foods. Theantioxidant advantage of lentil hull provides significant differentia-tion and may be better than the pea hull or fibers that reduce insulinresistance and fasting insulin levels in clinical trials (Hughes, 2010).This study highlights both secondary processing components involv-ing dehulling, and tertiary level of pulse processing where the by-product, the hulls, have been investigated to improve their functionalantioxidant value. We have used a wide range of solvents to producephenolic antioxidants, and developed a platform that allows itspractical application of phytochemical antioxidant extraction as a firststep in adding value to waste products like hulls that are now used foranimal feeding.

Acknowledgements

We thank Université de la Réunion, Ecole Supérieure d'Ingénieursen Développement Agroalimentaire Intégré (ESIDAI) for providing aninternship grant to François Caspar and Judy Harrison for technicalsupport.

References

Amarowicz, R., Karamac, M., & Chavan, U. (2001). Influence of the extraction procedureon the antioxidative activity of lentil seed extracts in a beta-carotene–linoleatemodel system. Grasas y Aceites Sevilla, 52(2), 89−93.

Amarowicz, R., Troszyńska, A., Barylko-Pikielna, N., & Shahidi, F. (2004). Polyphenolicsextracts from legume seeds:correlations between total antioxidant activity, totalphenolics content, tannins content and astringency. Journal of Food Lipids, 11,278−286.

Cudrado, C., Grant, G., Rubio, L. A., Muzquiz, M., Bardocz, S., & Pusztai, A. (2002).Nutritional utilization by the rat of diets based on lentil (Lens culinaris) seedmeal orits fractions. Journal of Agricultural and Food Chemistry, 50, 4371−4376.

Dueñas, M., Hernández, T., & Estrella, I. (2006). Assessment of in vitro antioxidantcapacity of the seed coat and the cotyledon of legumes in relation to their phenoliccontents. Food Chemistry, 98, 95−103.

Fernandez-Orozco, R., Zieliński, H., & Piskula, M. K. (2003). Contribution of low-molecular-weight antioxidants to the antioxidant capacity of raw and processedlentil seeds. Nahrung/Food, 47, 291−299.

Fitzpatrick, K. (2007). Novel fibers in Canada: Regulatory developments. Cereal FoodsWorld, 52(1), 22−23.

Halvorsen, B. L., Holte, K., Myhrstad, M. C. W., Barikmo, I., Hvattum, E., Remberg, S. F.,et al. (2002). A systematic screening of total antioxidants in dietary plants. Journalof Nutrition, 132, 461−471.

Harle, D., & Green, R. (2008). Lentil extract. United States Patent, 0280024, A1.Hughes, M. (2010). Nutritional trials prompt launch of new pulse flour products.

University News April 30, 2010. www.universitynews.org/f2ShowScript.aspx?i=23312.

Kubicka, E., & Troszyńska, A. (2003). Antioxidative potential of lentil seed coat towardslipoxygenase activity and beta-carotene oxidation. Polish Journal of Food andNutrition Sciences, 12(SI 1), 147−150.

Moise, J. A., Han, S., Gudynaite-Savitch, L., Johnson, D. A., & Miki, B. L. A. (2005). Seedcoats: structure, development, composition, biotechnology. In Vitro CellularDevelopmental Biology — Plant, 41, 620−644.

Narasimha, H. V., Ramakrishnaiah, N., & Pratape, V. M. (2003). Milling of pulses. In A.Chakraverty, A. S. Mujumdar, G. S. V. Raghavan, & H. S. Ramaswamy (Eds.),

441B.D. Oomah et al. / Food Research International 44 (2011) 436–441

Handbook of Postharvest Technology: Cereals, Fruits, Vegetables, Tea, and Spices(pp. 427−454). New York: Marcel Dekker, Inc.

Oomah, B. D., Corbé, A., & Balasubramanian, P. (2010). Antioxidant and anti-inflammatory activities of bean (Phaseolus vulgaris L.) hulls. Journal of Agriculturaland Food Chemistry, 58, 8225−8230.

Oomah, B. D., Blanchard, C., & Balasubramanian, P. (2008). Phytic acid, phytase,minerals, and antioxidant activity in Canadian dry bean (Phaseolus vulgaris L.)cultivars. Journal of Agricultural and Food Chemistry, 56, 11312−11319.

Oomah, B. D., Cardador-Martinez, A., & Loarca-Piña, G. (2005). Phenolics andantioxidative activities in common beans (Phaseolus vulgaris L). Journal of theScience of Food and Agriculture, 85, 935−942.

Prior, R. L., Hoang, H., Gu, L., Wu, X., Bacchiocca, M., Howard, L., et al. (2003). Assays forhydrophilic and lipophilic antioxidant capacity (oxygen radical absorbancecapacity (ORACFL)) of plasma and others biological and food samples. Journal ofAgricultural and Food Chemistry, 51, 3273−3279.

Ranilla, L. G., Genovese, M. I., & Lajolo, F. M. (2009). Effect of different cookingconditions on phenolic compounds and antioxidant capacity of selected Brazilianbean (Phaseolus vulgaris L) cultivars. Journal of Agricultural and Food Chemistry, 57,5734−5742.

Ronzio, R. A., Muanza, D. N., & Sparks, W. S. (1998). Antioxidant derived from lentil andits preparation and uses. US Patent, 5(762), 936.

SAS Institute Inc (1990). SAS/STAT user's guide, version 6 (4th ed.). Cary, NC: SASInstitute.

Takeuchi, T. M., Pereira, C. G., Braga, M. E. M., Maróstica, M. R., Jr., Leal, P. F., & Meireles,M. A. A. (2009). Low-pressure solvent extraction (solid–liquid extraction,microwave assisted, and ultrasound assisted) from condimentary plants. In M. A. A.Meireles (Ed.), Extracting Bioactive Compounds for Food Products (pp. 137−218). BocaRaton: FL: CRC Press.

Thavarajah, D., Ruszkowski, J., & Vandenberg, A. (2008). High potential for seleniumbiofortification of lentils. Journal of Agricultural and Food Chemistry, 56,10747−10753.

Troszyńska, A., Estrella, I., López-Amóres, L. M., & Hernández, T. (2002). Antioxidantactivity of pea (Pisum sativum L.) seed coat acetone extract. LebensmittelWissenchaft und Technologie, 35, 158−164.

Troszyńska, A., Bednarska, A., Latosz, A., & Kozlowska, H. (1997). Polyphenoliccompounds in the seed coat of legume seeds. Polish Journal of Food and NutritionSciences, 6/47(8), 37−44.

Wang, N. (2008). Effect of variety and crude protein content on dehulling quality andon the resulting chemical composition of red lentil (Lens culinaris). Journal of theScience of Food and Agriculture, 88, 885−890.

Xu, B., & Chang, S. K. C. (2008). Effect of soaking, boiling, and steaming on total phenoliccontent and antioxidant activities of cool season food legumes. Food Chemistry, 110,1−13.

Xu, B. J., & Chang, S. K. C. (2007). Comparative studies on phenolic profiles andantioxidant activities of legumes as affected by extraction solvents. Journal of FoodScience, 72, S159−S166.