Comparative evaluation of protein quality of raw and differentially processed seeds of an under-utilized food legume, Abrus precatorius L.

M Pugalenthi*, V Vadivel and P Janaki

Department of Biotechnology, Karpagam Arts and Science College,
Coimbatore - 641 021, Tamil Nadu, India
pugalmuthiah@rediffmail.com

Abstract

In the present study, the nutritional and antinutritional characteristics, biological value and protein quality of raw and differentially processed seed samples of a South Indian under-utilized food legume, Abrus precatorius L. were investigated.

The mature seeds were found to contain 21.99% of protein, 8.9% of lipid, 6.4% of fiber, 5.3% of ash and 57.3% of carbohydrates. Although, various antinutritional compounds were present in the raw seeds, the autoclaving treatment effectively reduced their maximum levels without affecting the nutritional value of Abrus precatorius seeds. When considering the biological value, the rats fed on autoclaved seeds exhibit better growth performance such as higher feed intake (FI) (239g) and body weight gain (BWG) (67.4g). Moreover, the protein quality parameters such as protein efficiency ratio (PER), true digestibility (TD), biological value (BV), net protein utilization (NPU) and utilizable proteins (UP) of Abrus precatorius seeds were also significantly improved by autoclaving treatment when compared to other processing methods such as soaking, cooking and roasting.

Key words: Abrus precatorius, antinutritional compounds, biological value, processing methods, protein quality

Introduction

The recent research trends indicate that there is an increase interest towards the search for alternative/additional protein source to meet the increasing requirements for protein source for expanding livestock industries, especially in the developing countries (Janardhanan et al 2003). The food scientists and nutritionists have attempted to develop certain novel, alternative protein sources mainly due to two main reasons:  (i) the low production of oil seeds and grain legumes;  and (ii) the stiff competition between man and the livestock industries in utilizing the existing legume seeds as protein source.

Even though soy bean and other common legume grains plays a key role as protein source for both human beings and animals, their production is not sufficient to meet the protein requirements of increasing population and expanding livestock industries (Vijayakumari et al 2007). The heavy demand for these common legumes has given rise to a disproportionate increase in their prices, and consequently, the cost of the food and feedstuffs. Hence, the recent research efforts have directed to identify and evaluate the under-utilized legume seeds as alternative/additional protein source for the future world (Janardhanan et al 2003; Pugalenthi et al 2005).

In this context, the seeds of Abrus precatorius L., an under-utilized food legume widely distributed through out the tropics and Andaman Islands, having good nutritional properties receives more attention as an alternative protein source. The seeds were reported to contain appreciable amount of protein (16-23%) with major proportion of albumin proteins, has a balanced amino acid composition and high content of lipid (2.0-9.4%) and desirable mineral composition (Rajaram and Janardhanan 1992; Mohan and Janardhanan 1995; Banerji and Dixit 1998; Vadivel and Janardhanan 1999). The cooked seeds have been consumed by Onges of Andaman and Katkharis of Pune District, Maharastra, India (Janardhanan et al 2003).

The exploitation and development of such potential non-conventional legume seeds as protein source for human beings/animals may offer a good scope to meet the increasing protein requirements at large, particularly in the developing countries. However, before recommending such indigenous foodstuffs, their nutritional properties and biological value should be thoroughly investigated. Although reports are available on the biochemical composition and nutritional value of Abrus precatorius, the information regarding the biological value of seed proteins appears to be meager. Hence, in the present study an attempt has been made to analyze the biological value and protein quality of raw and differentially processed seeds of Abrus precatorius collected from South India.

Materials and methods

Seed sample

The seed materials of Abrus precatorius L. were collected from Anaimalai Hills, Pollachi, Coimbatore District, Tamil Nadu, South India. Soon after collection, the immature and damaged seeds were removed and the mature seeds were dried in the sun light for 24 h and stored in plastic containers in refrigerator (5^oC), until further use.

Processing methods

Five separate batches of whole seeds of Abrus precatorius were taken and the first batch was soaked in distilled water for 6 h at room temperature (30 ± 2^oC) in the bean to water ratio of 1:10 (w/v). The second batch of seeds was cooked at 90-95^oC for 1 h in the bean to water ratio of 1:10 (w/v). The third batch of seeds was taken in the bean to water ratio of 1:10 (w/v) in a metal container and autoclaved at 15 lb pressure (121^oC) for 30 min. The fourth batch seeds was roasted for 30 min at 100-110^oC in an iron pot along with clean fine sand to prevent the burning of the seed coat and to ensure the uniform distribution of heat. After each treatment, the treated seeds were rinsed with distilled water, separately, and then dried at 55^oC for 6 h in a hot air oven. The fifth batch of raw seeds was stored as such without any treatment. 

Analytical methods

All the processed as well as raw seeds of separate batches were powdered in a Willey Mill to 60-mesh size and the powdered samples were used for further analysis. The proximate composition such as moisture, crude protein, crude lipid, crude fiber and ash content of raw as well as processed seed flour was determined by following AOAC (1990) method. Nitrogen free extractives (NFE) and calorific value were calculated by following the method of Siddhuraju et al (1992).

The antinutritional compounds such as total free phenolics and tannin content of raw and processed seed samples were extracted and estimated by following the method of Sadasivam and Manickam (1992) and Burns (1971), respectively. The L-Dopa (L-3,4-Dihydroxyphenylalanine) content was quantified according to the method of Brain (1976), whereas, the phytic acid content was determined by following the method of Wheeler and Ferrel (1971) and the oligosaccharides content was estimated by following Pugalenthi et al (2006) method. The haemagglutinating activity was analyzed according to the method of Makker et al (1997) and trypsin inhibitor activity was determined by casein digestion method (Mulimani and Vadiraj 1993) while α- amylase inhibitor activity was measured according to the method  of Mulimani and Rudrappa (1994).

Biological evaluation of protein quality

Fifty numbers of 23 days old male albino rats with an initial body weight of 40 ± 5 g were equally divided into five groups with 10 animals in each group and housed individually in cages. The animals were maintained at 22^oC with 12 h light and 12 h dark at Karpagam Animal House (Approved by Animal Ethical Committee, Government of India). The experimental diets were prepared according to the method of Chapman et al (1959) by including corn starch (80%), corn oil (10%), non-nutritive cellulose (5%), mineral mixture (4%) and vitamin mixture (1%). The test diets were prepared by incorporating raw and differentially processed seeds as protein source at 10% level in the diet and the control diet was prepared by incorporating casein as a protein source. The control and test diets were fed to respective animal groups along with water ad libitum for 28 days and the growth performance of the experimental animals were analyzed.

Feed Intake (FI): Dry feed fed (g)/animal/28 days

The protein content of the diet was determined by micro-Kjeldahl method (AOAC 1990) and Feed Efficiency Ratio (FER) and Protein Efficiency Ratio (PER) were calculated according to the method of Chapman et al (1959).

The nitrogen balance studies were conducted for 14 days with 60 numbers of male albino rats of 50 ± 5 g body weight. The rats were randomly separated into six groups with 10 animals in each group and individually housed in polypropylene metabolic cages. The animal groups were fed with control casein diet, raw and differentially processed seeds included test diets separately and one batch was also fed with protein free basal diet for the determination of endogenous and metabolic nitrogen loss in faeces and urine. After 9 days of acclimatization period, the urine and faeces of the experimental animals were collected for five days and pooled separately. The nitrogen content of the urine and faeces were estimated by micro-kjeldahl method (AOAC 1990) and the values of True Digestibility (TD) and Biological Value (BV) of seed proteins were determined by following the method of Chick et al (1935) and Net Protein Utilization (NPU) by Platt et al (1961), whereas, the level of Utilizable Proteins (UP) was calculated by Gupta et al (1979) method.

Where,

NI = Nitrogen intake of the animals.

NF1= Nitrogen excreted in the faeces of the animals fed with test diet.

NF2= Nitrogen excreted in the faeces of the animals fed with protein free diet.

NU1= Nitrogen excreted in the urine of the animals fed with test diet.

NU2= Nitrogen excreted in the urine of the animals fed with protein free diet.

Statistical analysis

Results were expressed as mean values ± standard deviations of three separate determinations. The data was subjected to a one-way analysis of variance (ANOVA) and the significance of difference between means at 5% was determined by Duncan’s Multiple Range Test (DMRT) using Irristat software (version 3/93).

Results and discussion

Proximate composition

The proximate composition of raw and processed seeds of Abrus precatorius were shown in Table 1.

The crude protein and lipid content of raw seeds (21.99% & 8.94%) were found to be similar when compared to certain common legumes such as Pisum sativum (21.9% & 2.34%); Phaseolus vulgaris (20.9% & 2.49%); Cicer arietinum (18.5% & 6.69%) and Lens culinaris (20.6% & 2.15%) as reported by Costa et al (2006). The protein and lipid content of autoclaved seed samples (22.04% & 8.95%) were higher than the raw and other processed seeds, which was in consonance with the earlier report in velvet beans (Siddhuraju and Becker 2005; Vadivel and Pugalenthi 2007). The reduction of the ash content in the soaked seeds (14%) when compared to raw seed samples might be due to the leaching of both micro and macro minerals into the soaking medium through the enhanced permeability of the seed coats during soaking treatment.

Antinutritional compounds

The effect of various processing methods such as soaking, cooking, autoclaving and roasting on the levels of antinutritional compounds of Abrus precatorius were given in Table 2.

Among the various processing methods employed, the autoclaving was found to significantly (p<0.05) reduce the maximum levels of various antinutritional substances such as total free phenolics (72%), tannins (73%), L-Dopa (84%), phytic acid (72%), oligosaccharides like raffinose (72%), stachyose (57%) and verbascose (67%), haemagglutinating activity (72%), trypsin inhibitor activity (65%) and amylase inhibitor activity (60%). Similarly, significant reduction of various antinutritional compounds during autoclaving treatment was reported for several under-utilized legumes like Bauhinia purpurea (Vijayakumari et al 2007); Dolichos lablab (Vijayakumari et al 1995); Mucuna monosperma (Vijayakumari et al 1996); Vigna aconitifolia and Vigna sinensis (Vijayakumari et al 1998).

Growth performance of the animals

The growth performance of the experimental animals fed with diets containing raw and processed Abrus precatorius seeds was illustrated in Table 3.

The FI value of the animals fed with diet containing raw seeds (161 g) was significantly lower (p<0.05) as compared to casein (292 g) and treated seeds (194-229 g) and earlier reports on Canavalia ensiformis (225 g) and Canavalia gladiata (244 g) (Bressani et al 1987), but higher than the vegetable peas (105-120 g) (Saharan and Khetarpaul 1994) and Canavalia maritima (93 g) (Seena et al 2005). The notable difference in the FI value of the rats fed with diet containing raw and differentially processed Abrus precatorius seeds was probably due to the difference between the diets in protein quality and levels of antinutritional compounds of the incorporated seeds. Maximum reduction on the levels of various antinutritional substances under autoclaving treatment might be related to larger FI values (229 g) of animals fed with autoclaved Abrus precatorius seeds inclusive diet.

The animal group fed with raw Abrus precatorius seeds included diet shows lowest BWG value (31.6 g) when compared to the control (96 g) and test animals fed with diet included with treated seeds (40-68 g). The BWG value of animals fed with diet containing raw seeds of the present study was in agreement with an earlier report on vegetable peas (23-28 g) (Saharan and Khetarpaul 1994). Among the different treated seeds, the autoclaved seeds significantly improve the BWG of rats (68 g), which was higher than the BWG values for pressure-cooked seeds of Canavalia ensiformis (28 g) and Canavalia gladiata (32 g) (Bressani et al 1987). Soaking and cooking processing methods were not demonstrated to have a beneficial effect on the growth rate of the animals. It might be due to the presence of heat resistant antinutritional compounds in the Abrus precatorius seeds, which are not destroyed completely by the soaking and cooking treatments.

The raw Abrus precatorius seeds exhibit poor FER (0.19) and PER (1.97) values when compared to control and processed seeds (Table 3), which might be due to the presence of high concentration of antinutritional substances and poor quality of proteins of the raw seeds. However, the FER and PER values of raw Abrus precatorius seeds were found to be higher than the faba beans (0.032 & 0.32) (Gupta et al 2005), but lower than vegetable peas (0.22 & 2.17) (Saharan and Khetarpaul 1994). Among the different treatments, the autoclaving resulted in significant (p<0.05) improvement of FER (0.29) and PER (2.96) of Abrus precatorius seeds. The PER value of autoclaved Abrus precatorius seed was higher when compared to Canavalia ensiformis (1.24) and Canavalia gladiata (1.24) (Bressani et al 1987). The results observed from the present study showed that the higher the FI, the higher the PER values obtained, which was coincided with an earlier report given by Bender (1956), in which he has pointed out that the PER determination is depends upon feed consumption.

The raw as well as processed seeds of Abrus precatorius exhibited lower PER values when compared to control casein diet. Usually much of the sulphur containing amino acids such as cysteine and methionine supplied in the diet were used to synthesize pancreatic enzymes (Fernandez et al 1996). This exacerbated the deficiency of sulphur containing amino acids in the legume seeds and manifested a lower production of body tissues. Rerouting of retained nitrogen in the animal group fed with raw Abrus precatorius seeds containing diet may explain why PER in this group was significantly lower than in control, despite the fact that protein content was similar among these groups.

Among the processed seeds, the soaked and cooked seeds were exhibited lower PER values (1.97 and 2.11, respectively), which might be due to the fact that soaking and cooking treatments are not effective in reducing the antinutritional compounds, which interfere with the protein quality of Abrus precatorius seeds. The lower PER value of roasted (1.98) seeds compared to casein (3.32) and autoclaved seeds (2.96) could be due to the fact that heat accelerates the millard reaction and makes the protein unavailable for digestion (Sagarbieri 1989). According to Friedman (1996), PER value below 1.5 describes a protein of poor quality; between 1.5 and 2.0 an intermediate quality and above 2.0 good quality. Hence, the seed proteins of raw Abrus precatorius was considered as intermediate quality seed proteins, whereas, the processed seeds possess proteins with good quality.

Protein quality

The protein quality such as True Digestibility (TD), Biological Value (BV), Net Protein Utilization (NPU) and Utilizable Proteins (UP) of raw and processed Abrus precatorius seeds were presented in Table 4.

The TD value of raw seeds was found to be lower (65%) when compared to control (92%) and treated seeds (69-78%) of the present study, but higher when compared to the TD value of Canavalia maritima (42.2%) (Seena et al 2005); Vicia faba (63.4%) (Gupta et al 2005); Bauhinia purpurea (46.4%) (Vijayakumari et al 1997) and comparable with vegetable peas (65.8-66.7%) (Saharan and Khetarpaul 1994). The autoclaving treatment significantly improved the TD level of Abrus precatorius (78%), which was higher than the values reported for Canavalia ensiformis (76.4%) (Bressani et al 1987) and Vicia faba (71.4%) (Gupta et al 2005).

The consumption of raw legume seed proteins was reported to increases the endogenous nitrogen loss through the shedding of intestinal mucosa (Sanoja and Bender 1983; Fairweather-Tait et al 1983), an effect that reduces the biological value of raw legume seed proteins. Further, the presence of various antinutritional substances, including trypsin inhibitors, which inhibits the complete digestion of protein and increases the excretion of endogenous faecal nitrogen (Nestares et al 1996) was also partly responsible for the decrease in the TD value of seed proteins of raw Abrus precatorius seeds.

Among the differentially treated seeds, lowest BV value was recorded for raw Abrus precatorius seeds (58%), which was lower than that of BV level of vegetable peas (62.9-63.1%) (Saharan and Khetarpaul 1994) and faba bean (60.4%) (Gupta et al 2005), but comparable with Bauhinia purpurea (57.2%) (Vijayakumari et al 1997). The NPU value of raw Abrus precatorius seeds was also lower (42.8%) when compared to casein (73.6%) and differentially treated seeds (46.2-56.4%), but similar with that of an earlier reports on vegetable peas (41-42%) (Saharan and Khetarpaul 1994) but higher than Canavalia maritima (16.8%) (Seena et al 2005) and Vicia faba (38.3%) (Gupta et al 2005). The autoclaved seeds exhibited highest level of NPU (56.4%) among the raw and different treated seeds, which was also higher than the previous report on Bauhinia purpurea (46.4%) (Vijayakumari et al 1997). The level of UP of raw Abrus precatorius seeds (10.2%) was higher than that of vegetable peas (8.4-8.6%) (Saharan and Khetarpaul 1994) and Bauhinia purpurea (7.2%) (Vijayakumari et al 1997). The highest level of UP (19.1%) was registered by autoclaved seeds when compared to raw and other processed seeds (10.2-19.1%).

Rats that received dietary casein as protein source were able to take full advantage of the nitrogen they retained to favour growth, probably as a result of the more balanced supply of amino acids provided by this diet. In rats fed with raw Abrus precatorius seeds, part of the nitrogen retained may have been rerouted for the synthesis of digestive enzymes such as trypsin and chymotrypsin (Liener 1994) to offset the effects of various antimetabolic substances present in the raw seeds, hence the seed proteins of raw Abrus precatorius seeds obtain poor TD, BV, NPU and UP values.

The protein quality parameters such as TD, BV, NPU and UP of the autoclaved Abrus precatorius seeds were higher than the raw and other processed seeds, which was in consonance with the previous study in Bauhinia purpurea (Vijayakumari et al 1997) and Vigna unguiculata (Dario and Salgado 1994). The decrease in the trypsin inhibitor activity and other antinutritional constituents as a consequence of autoclaving treatment would have been reducing the faecal nitrogen excretion in rats fed with autoclaved seeds included diet. Improvement in the protein quality after autoclaving treatment might be attributed to reduction on the levels of various antinutrients in addition to some other factors such as disruption of protein structure and increased accessibility of the seed proteins to enzymatic attack (Nielson 1991).

The fact that no such decrease in faecal nitrogen excretion was occurred in the rats fed with diet containing roasted seeds may have been due to the fact that dry heat treatment cause isopeptide formation (Dutson and Orcutt 1984; Kirk 1984). This reduces the protein quality, as isopeptides are not hydrolyzed in the intestine, are resistant to proteolytic enzymes and are thus excreted in faeces. As a result, digestibility and availability of some amino acids are reduced (Kirk 1984) and thus gave poor values of BV, NPU and UP for roasted gila bean seeds. Geervani and Theophilus (1980) also observed that wet heat processing method improves the protein quality of Cicer arietinum and Vigna radiata to a greater extent than dry heat methods.

Conclusions

• The results of the present study indicated that the autoclaving treatment was suitable and more effective in reducing various antinutritional compounds of Abrus precatorius seeds with out affecting the nutritional quality.

  
  

• When considering the biological value, the autoclaved Abrus precatorius seeds exhibited better animal growth performance and protein quality than the other processed seeds. Hence, such economic and potential processing method could be adapted for the versatile utilization of Abrus precatorius seeds as an alternative/additional protein source in the diets of human beings/animals, which will clearly reduce the over-dependence on common legumes for increasing protein requirements, especially in the developing countries.

Acknowledgements

Authors are grateful to the University Grants Commission for giving financial support to a Major Research Project [Sanction No. F. 3 – 42/2004 (SR) dt. 12.01.2004] and thankful to the Management and Administrative authorities of Karpagam Arts and Science College for their encouragement and support.

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Received 18 September 2007; Accepted 21 September 2007; Published 1 November 2007

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