Genotypic Variation in Pigeonpea for Protein, Dietary Fibre and Lectins

Indian J Agric Biochem 25(2),111-115,2012 Genotypic Variation in Pigeonpea for Protein, Dietary Fibre, Fatty acids and Lectins

R P SRIVASTAVA* and HINA VASISHTHA

Department of Biochemistry, Indian Institute of Pulses Research, Kanpur – 208 024, Uttar Pradesh, India

Six genotypes of pigeonpea viz., Bahar, UPAS 120, MAL 13, NDA 1, BSMR 853 and ICP 7035 were evaluated for their soluble protein, dietary fibre (cellulose, hemicellulose, lignin and pectin), fatty acids and lectins. Pigeonpea genotypes contained soluble protein in the range of 20.13 to 23.35%. The genotype ICP 7035 and UPAS 120 contained significantly high soluble protein in their seed. The cellulose and hemicellulose in different genotypes of pigeonpea was in the range of 3.12 to 3.70% and 1.16 to 3.31%, respectively. ICP 7035, MAL 13 and Bahar had lowest lignin as compared to other genotypes. ICP 7035 also had lowest hemicellulose and total saturated fatty acids and highest polyunsaturated (PUFA) and total unsaturated fatty acids (TUFA). Pectin in grain was present in the range of 2.89 to 4.82%. BSMR 853, NDA 1, ICP 7035 and UPAS 120 had higher pectin than other genotypes. Haemagglutinating activity (lectins) of pigeonpea grain was varying in the range of 1205 to 1415 HU/g.

Key words: Pigeonpea, soluble protein, cellulose, hemicellulose, lignin, pectin, fatty acids, lectins

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*Author for correspondence, Email: [email protected]

Pigeonpea is an important pulse crop of India. The world’s pigeonpea production is around 3.5 million tons annually. India ranks first in world pigeonpea production with annual 2 production of 3.07 million tons (1). Other important pigeonpea producers are Myanmar, Malawi, Uganda, Kenya, Tanzania, Nepal and Bangladesh. In India Maharashtra, Uttar Pradesh, Madhya Pradesh, Karnataka, Gujarat and Rajasthan are the major pigeonpea producing states. Pigeonpea is also a food crop in many tropical countries and is commercially important in countries of East Africa, the Caribbean and Latin America. This is mostly consumed in the form of dehulled and split cotyledons (dhal). Pigeonpea flour is used in the preparation of several traditional products. It is an excellent source of protein, carbohydrate, dietary fibre, minerals and vitamins. Hypocholesterolemic property of food legumes is due to the presence of high protein, high dietary fibre, presence of unsaturated fatty acids and saponins (2,3).The risk of cardiovascular disease is lower among people who consume fibre rich food (4). Because of these advantages, food legumes are becoming popular among the people living in developed countries. The dietary fibre is important as it helps in lowering the blood cholesterol. Lectins are proteinaceous toxic factors that interact with glycoprotein on the surface of red blood cells and cause them to agglutinate. Food legumes have long been known to contain protein compounds which agglutinate the red blood cells. Therefore, this investigation was carried out to work out the variability in dietary fibre components, soluble protein, fatty acids and lectins in different genotypes of pigeonpea.

Materials and Methods

Seed material: Six varying genotypes of pigeonpea namely Bahar, UPAS 120, MAL 13, NDA 1, BSMR 853 and ICP 7035 were selected for this study. The seeds/grains of these genotypes were collected from the crop grown at the Indian Institute of Pulses Research, Kanpur, UP, India during 2006-07. The grain samples were dried at 70 ºC and powdered to a uniform particle size in a seed grinder Perten model 3303 and analysed in triplicate. The results were calculated on dry weight basis. Determination of protein, dietary fibre and lectins: Soluble protein in grains of different genotypes was estimated by Lowry’s method (5) and reported as percent soluble protein on dry weight basis. The cellulose was determined by the method of Updegroff (6) and results expressed as percent on dry weight basis. The hemicellulose was determined according to method of Goering and Van Soest (7) and the results calculated as difference of NDF (neutral detergent fiber) and ADF (acid detergent fiber), and reported as percent on dry weight basis. The lignin was 3 determined by the method of Van Soest (8) and the ADL (acid detergent lignin) reported as lignin percent on dry weight basis. The pectin was determined by the method of Ranganna (9) and reported as percent on dry weight basis. Lectins were quantified by haemagglutinating test as per method of Grant et al. (10) and reported as haemagglutinating units (HU) per gram grain on dry weight basis. Determination of fatty acids: The fatty acids in pigeonpea grains were determined as per the method described by Srivastava and Srivastava (11) with minor changes. The powdered seed samples were extracted with 12% sulphuric acid in methanol by shaking for 20 hrs in a rotary shaker and the filtrate was extracted in chloroform, the moisture was removed using anhydrous sodium sulphate. The methyl esters of fatty acids so prepared were quantified against standard fatty acid methyl esters purchased from Sigma-Aldrich (USA) on a Nucon 5765 gas chromatograph. A stainless steel column of 15% DEGS (2 m x 2 mm) with FID detector was used for separation. Temperature programming was 130 C (2 min) to 150 C (2 min) at 4 C/min increase, and then 150 C to 195 C at 3 C/min was followed. Injector and detector temperatures were set at 230 ºC and 240 ºC. 2 µL of methyl esters prepared from pigeonpea along with standard methyl esters of fatty acids were injected in gas chromatograph. Winacds software was used for computation. The flow of carrier gas (nitrogen) was adjusted to 30 mL/min and the fuel gas hydrogen and air were in the ratio of 30:300 mL/min. Three chromatograms were run for each sample and the average values used for statistical analysis. The palmitic, stearic, oleic, linoleic and linolenic acids were quantified as % of total fatty acids in seed. Total saturated, polyunsaturated (PUFA) and total unsaturated (TUFA) fatty acids of pigeonpea were also calculated. Statistical analysis: The Statistical Package for the Social Sciences (SPSS, version 13) was used for all data analysis. The data were analysed by one- way analysis of variance (ANOVA). Significance of the differences was defined as P<0.05.

Results and Discussion

Soluble protein: There was a wide variability in soluble protein content of seed in pigeonpea. Soluble protein in seed was found in the range of 20.13 to 23.35% (Table 1). Average soluble protein of pigeonpea genotypes was 21.78%. UPAS 120 and ICP 7035 contained significantly higher soluble protein in the grain. Other genotypes such as Bahar, MAL 13, NDA 1 and BSMR 853 contained moderate soluble protein (Fig 1). Reddy et al. (12) have reported protein content in pigeonpea in the range of 18.4 to 28.8%. Srivastava and Bajpai (13) reported crude protein in the 4 range of 15.37 to 21.87%. Ghadge et al. (14) have reported 24.43% crude protein in raw seeds of pigeonpea. High soluble protein in grain is an important quality attribute and is considered beneficial from nutrition point of view. High protein helps in lowering cholesterol in blood vascular system (15). Dietary fibre: The dietary fibre of seed consists of cellulose, hemicelluloses, lignin and pectin. The average cellulose in seed of pigeonpea genotypes was 3.41% and present in the range of 3.12 to 3.70%. Highest content of cellulose was observed in Bahar (3.70%) and lowest in ICP 7035 (3.12%). The average hemicellulose in pigeonpea genotypes was 2.40% and the range of hemicellulose in different genotypes of pigeonpea was 1.16 to 3.31%. Highest hemicellulose was found in NDA 1 (3.31%) and lowest in ICP 7035 (1.16%). The average lignin in seed of pigeonpea genotypes was 3.12% and present in the range of 2.07 to 4.56%. High lignin was found in seed of NDA 1 and UPAS 120, whereas MAL 13 contained low lignin in their seed. Apata (16) reported lignin in pigeonpea seed in the range of 1.36 to 2.04%. Insoluble fibre mainly consists of cellulose, hemicellulose and lignin. It was present in the range of 7.19 to 11.20%, and highest insoluble fibre was observed in NDA 1 (11.20%), followed by UPAS 120 (9.76%), BSMR 853 (8.78%), Bahar (8.49%), MAL 13 (8.19%) and ICP 7035 (7.19%) (Fig 1). The average pectin in pigeonpea seed was 4.19% and present in the range of 2.89 to 4.85%. High pectin was found in seed of NDA 1, BSMR 853 and UPAS 120, whereas MAL 13 contained low pectin in its seed. Pectin helps in binding lipids and cholesterol (17,18). Pectin is not used in human digestion, but passes through small intestine more or less intact. In the large intestine and colon, microorganism degrades pectin and converts them into short chain fatty acids that have positive influence on health. NDA 1 and UPAS 120 also contained high total dietary fibre, whereas ICP 7035 and MAL 13 had the least total dietary fibre in their grains (Fig 1). Importance of dietary fibre is well known in lowering the cholesterol (4,19). 5

t 15 n e c r e

0 Bahar UPAS 120 MAL 13 NDA 1 BSMR 853 ICP 7035 Varieties Sol Protein Total fibre Insoluble fibre

Fig 1: Soluble protein and fibre contents of pigeonpea genotypes

Fatty acids: The fatty acid composition of different genotypes of pigeonpea is summarised in table 2. Palmitic and stearic acids were the main saturated fatty acids of pigeonpea grain, whereas oleic, linoleic and linolenic were main unsaturated fatty acids. The most predominant saturated fatty acid of grain was palmitic acid. The average palmitic acid in whole seed of pigeonpea was 28.85% of total fatty acids. The range of palmitic acid in grain of different genotypes was 22.46 to 33.62%. Highest content of palmitic acid was found in UPAS 120 and Bahar, whereas ICP 7035 (22.46%) contained lowest palmitic acid. The stearic acid was another saturated fatty acid of pigeonpea and was much lower than palmitic acid in seed. The average content of stearic acid in whole seed of pigeonpea was 3.18% of total fatty acids and present in the range of 2.54 to 3.88% of total fatty acids in different genotypes. Higher stearic acid was found in BSMR 853, Bahar and MAL 13, whereas NDA 1 contained significantly low stearic acid. ICP 7035 had the least total saturated fatty acids, whereas Bahar and UPAS 120 contained highest saturated fatty acids in their grain (Fig 2). 6

60 s d i c a

50 y t t a f

40 l a t o t

30 f o

0 Bahar UPAS 120 MAL 13 NDA 1 BSMR 853 ICP 7035 Varieties TUFA TSFA

Fig 2: Total unsaturated and saturated fatty acids in pigeonpea genotypes

The oleic acid was the main mono unsaturated fatty acid (MUFA) of pigeonpea. The average oleic acid in pigeonpea grain was 8.90% and present in the range of 7.14 to 13.72%. Highest content of oleic acid in pigeonpea was found in BSMR 853 and lowest in UPAS 120 genotypes. The linoleic acid was most predominant poly unsaturated fatty acid (PUFA) of pigeonpea. The average linoleic acid in pigeonpea grain was 41.72% and present in the range of 35.75 to 45.44%. Highest linoleic acid in grain of pigeonpea was found in MAL 13 and lowest in BSMR 853. Oshodi et al. (20) have reported 54.8% linoleic acid in pigeonpea seed. The linolenic acid was another poly unsaturated fatty acid (PUFA) of pigeonpea. The average linolenic acid in pigeonpea genotypes was 17.33%. The range of linolenic acid in different genotypes of pigeonpea was 13.68 to 20.34%. Higher linolenic acid was present in BSMR 853, NDA 1 and ICP 7035, whereas MAL 13 and UPAS 120 contained low linolenic acid. High polyunsaturated fatty acids (PUFA) were observed in ICP 7035 and NDA 1 genotypes of pigeonpea. These two genotypes in addition to BSMR 853 also contain high total unsaturated fatty acids (Fig 2). The fatty acid profile of pigeonpea grain was more or less similar to chickpea, lentil and fieldpeas (21,22,23). It has been reported that high unsaturated fatty acids help in lowering cholesterol in blood vascular system. Lectins: The variation in haemagglutinating activity (HA) in grain of different genotypes of pigeonpea was in the range of 1205 to 1415 HU/g. The average HA activity of pigeonpea grain was 1273.3. High HA activity was observed in ICP 7035 and UPAS 120, whereas Bahar, MAL 7

13, BSMR 853 and NDA 1 have shown relatively low HA activity. High lectins in pulses are harmful; therefore they are required to be degenerated before consuming as food. Processing techniques such as boiling, cooking or pressure cooking help in complete destruction of lectins, therefore pulses should be consumed after necessary processing. Savelkoul et al. (24) have reported degradation of lectins during different processing of legumes.

Acknowledgements

Thanks are due to Dr I.P.Singh for supplying seeds of pigeonpea genotypes.

References

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Table 1: Protein, dietary fibre and lectins in different genotypes of pigeonpea ______

Genotypes Soluble protein Cellulose Hemicellulose Lignin Pectin Lectins (HU/g) (%) (%) (%) (%) (%) ______

Bahar 21.88±0.52 3.70±0.23 2.40±0.07 2.39±0.01 3.87±0.15 1205±1.00 UPAS 120 23.35±0.50 3.37±0.13 2.56±0.07 3.83±0.03 4.67±0.05 1310±1.00 MAL 13 20.13±0.57 3.38±0.05 2.74±0.05 2.07±0.02 2.89±0.15 1226±4.58 NDA 1 20.47±0.82 3.33±0.07 3.31±0.48 4.56±0.06 4.85±0.17 1235±5.56 BSMR 853 21.50±0.13 3.54±0.07 2.23±0.12 3.01±0.01 4.82±0.08 1249±2.00 ICP 7035 23.34±0.17 3.12±0.12 1.16±0.10 2.91±0.01 4.05±0.09 1415±4.35

Mean 21.78 3.41 2.40 3.12 4.19 1273.3 SEm 0.18 0.10 0.17 0.26 0.10 2.92 CD (P<0.05) 0.38 0.23 0.38 0.56 0.22 6.36 ______

Each value is mean of three replications. Mean values ± SD.

Table 2: Fatty acids in different genotypes of pigeonpea (In percent of total fatty acids)

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Genotypes Palmitic Stearic Oleic Linoleic Linolenic Sat. fatty PUFA Unsat. 10

acid acid acid acid acid acids fatty acids ______

Bahar 33.36±0.30 3.75±0.02 7.81±0.80 38.75±0.80 16.33±0.01 37.11 55.08 62.89 UPAS 120 33.62±0.60 2.63±0.01 6.83±0.62 42.66±1.96 14.26±0.06 36.25 56.92 63.75 MAL 13 29.67±0.60 3.65±0.08 7.56±0.20 45.44±0.40 13.68±0.02 33.32 59.12 66.68 NDA 1 27.72±0.70 2.54±0.02 7.14±0.10 42.65±0.20 19.95±0.05 30.26 62.60 69.74 BSMR 853 26.31±0.30 3.88±0.07 13.72±0.56 35.75±0.50 20.34±0.04 30.19 56.09 69.81 ICP 7035 22.46±0.40 2.65±0.05 10.37±0.30 45.08±1.00 19.44±0.44 25.11 64.52 74.89

Mean 28.85 3.18 8.90 41.72 17.33 32.04 59.06 67.96 SEm 0.41 0.04 0.40 0.81 0.15 - - - CD 0.90 0.09 0.88 1.76 0.33 - - -

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Each value is mean of three replications. Mean values ± SD.